VIECHIVE.F

FISHERIES AND MARINE SERVICE

Translation Series No. 4170

Chronic optico-neuropathy due to environmental exposure of organophosphate pesticides (Saku disease). Clinical and experimental study

by S. Ishikawa

Original title: Kogai to Me. Yukirin to Me. Mansei Yukirin Chudokusho no Ekigaku no Rinsho oyobi Jikkenteki Kenkyu

From: Nippon Ganka Gakkai Zasshi 77(10): 1851-1886, 1973

Translated by the Translation Bureau (YH/RR) Multilingual Services Division

Department of the Secretary of State of Canada

Department of Fisheries and the Environment Fisheries and Marine Service

Halifax Laboratory Halifax N.S.

1977

90 pages typescript

•

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DEPARTMENT OF THE SECRETARY OF STATE

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TRANSLATED FROM - TRADUCTION DE INTO - EN

Japanese English

PLACE OF PUBLICATION LIEU DE PUBLICATION

Japan

PERSON REQUESTING DEMANDÉ PAR

JAN 2 0 1978 Allan T. Reid

J,/ F v i d

DIVISION MULTILINGUES

AUTHOR - AUTEUR

Satoshi ISHIKAWA

TITLE IN ENGLISH - TITRE ANGLAIS

Chronic optico-neuropathy due to Environmental Exposure of Organophosphate

Pesticides (Saku Disease) ---Clinical and Experimental study---

TITLE IN FOREIGN LANGUAGE (TRANSLITERATE FOREIGN CHARACTERS) TITRE EN LANGUE éTRANGéRE (TRANSCRIRE EN CARACTLRES ROMAINS)

Kogai to Me. Yukirin to Me. Nansei Yukirin Chudokusho no Ekigaku no Rinsho oyobi Jikkenteki Kenkyu

(34 figures, 10 tables) REFERENCE IN FOREIGN LANGUAGE (NAME OF BOOK OR PUBLICATION) IN FULL. TRANSLITERATE,FOREIGN CHARACTERS. RéFéRENCE EN LANGUE éTRANG2RE (NOM DU LIVRE OU PUBLICATION), AU COMPLET, TRANSCRIRE EN CARACTE'RES ROMAINS.

Nippon Ganka Gakkai Zasshi

REFERENCE IN ENGLISH - RéFéRENCE EN ANGLAIS

Journal of the Japan Ophthalmological Society

PUBLISHER - éDITEUR

1973

YEAR

ANNE

DATE OF PUBLICATION DATE DE PUBLICATION

VOLUME

77

ISSUE NO. HUME RO

10

PAGE NUMBERS IN ORIGINAL NUMéROS DES PAGES DANS

L'OM GMAL

383-418 (1851-1886) NUMBER OF TYPED PAGES

NOMBRE DE PAGES

DACTYLOGRAPHIéES

89

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1448482 Japanese L_ YH/RR JAN 2 0 1078

1.

Chronic optico-neuropathy due to Environmental Exposure

of Organophosphate Pesticides (Saku Disease)

-- Clinical and Experimental study --

Satoshi ISHIKAWA

Collaborators

Mikio MIYATA, Kazuo KONO, Shinji ONO, Atsumi YAMAZAKI, Haruo HIKITA, Hajime

SUZUKI, Hiroko fl1AI, Shichiro MIYAZAWA, Tetsuo TAMAI, Hiroaki KUBO, Ken OTO,

Kimiyoshi UONO, Masaya SEGAWA, Shigezo UGA.

i't.sticid..s of the organopho •pltate (()P) compounds, the %viclest and catastrophic. use throughout

ti •s v:tirld have possibly increased the yield or ail' icultural produce and controlled vectors of certain tiopical Medicially, some 0P have been used for management of glaucoma, esotropia and

iuyastlitinia ut avis. 'Fhis tis• ha; decreased because or the nail ow margin between therapeutic and

clo;es. ()1> are potent inhil)itors of carboNylic esterae cmzyntes including ctretylcholin esier- a‹.-s ;:rciP est . , rase) inc1 pseudockilmesterase. pharmarologic and toxiciolc.)gic action of OP are

irhibition of acetyltholine.terase of' nervous system as \yell as e.yes causing ciccumulation of' aretyl-

(hipline at the synapses. l'he overaccumulation of acetylcholine initially stitnulates and then ptiralyzes

1io.r1: • 111 cholinergic symip;vc. These synapses are mainly central and peripheral nervoUs s) -stem,

veil:vie nerve:, ganglionic synapi,..s ni‘ autonomic nervous system and some sympathetic. IICI'VC of attain°. nervous system.

Acute intoxication of 01' i.e, par:ithion, inalathion, cliazinon and F..PN Jas been \yell known and

• N:c.e-iivc• lireiatur,s are avail:tide. ()a the other band, only the several p:tpers are available ctliout their °Il i,: i n toxication in linie.o i s, ease,; OP clue to triorthc.icreqyl phosphate have been repot u•t1

.is the name of Jamaica ginger palsy. This 1 ompotnicl inhibited ptiteclocholinesterase rug! the cliserese

%\ as known In çair,e sti\l•re abdominal cramp ‘viill a se\ -ere peripheral. polynetirilis. The eyes ‘vere

ti•ly O n Co e .•I' l er minter:1 pestit or OK' OP are knov. ri to cause delayed peripheral neurnpathy.

r uports stigg est ed that the OP - inhihited ptirnttl and trite c - holinesterase enzymes clnrim; prolonged e mInsure rt ml their toxicity is milcler and slinVer. f\bnlit thOir chronic toxicities, no report is available al:rint the ocula• c:omplications. lsbikawa reported L typical ocular syndrome due to 01 al the Sakit

district in 1 !IGO. In the present report. the descriptitm %yin 1 i rnacle about Saku disease and se.veral

t,cular concplications will be discussed with the experimental evidence using 2 years experiment ‘vith

SEC 5-25T (6/76)

2 .

OP in the beagle dogs, This disease ha s now been increasing in a high rate throughout Japan dur., several years. Most cases initially visited the hospital with the chief complaint ur reduced past visi• t•aL was never corrected. N lost cases Sho ■ved reduc ed erythrocyte and serum cholinesterase acti v it y

ss•ell as ti V' residue of CH.' in their blood or urine over 0.01 ppm when detected hy ilanie photometric gaschromatography.

The cases were selected by a follosving diagnostic criteria proposed by the members of the japanes., Health and Welthre Department. Major signs: Ocular sign---1. reduced vision 2. narrowings of the peripheral visual field and/or central scotomas 3. abnormal refraction of myopic tendency with or wit1. out severe vertical cortical astigmatism. General 'sign—. 1. existense of the pyramidal signs 2. reduction of the proprioceptive sense and difficulty of the one foot standing 3. autonomic signs. dizziness, headache, nausea, vomitting, diarrhoea, constipation, perspiration, numbness, polyclipsia, impotency, vesicular and rectal signs etc. Laboratory—detection of 01' in the blood or urine over 0.01 ppm. Therapeutic—recovery of the signs with the administration of 2-PA N1 or atropine. Additional signs: Ocular sien---1. congestion or atrophy of the optic nerve 2. difficulty of the ocular smooth pursuit move-ment 3. abnormal ERG; General sign.-1, reduction of the cholinesterase activity mainly erythrocytes. 2. mild abnormalities of the hiver functions main])' LDH, Cl'K, CCLF, Alkaliphosphatase, etc.. 3. sensory disturbance of glove- stsel:mg type. .1. foot drop 5. ERG abnormalities.

These clinical signs and syniptoms were perfectly proved either physiological test or histopathologi!.:al study of the. geagle dog's experiment during two years' chronic intoxication using per-os ethylthiometon 0.5-1.5 ppm administrations.

Froni these data, the author concluded that the Saku disease was definitely produced by the chronic exposure to oreanophosphate pesticides either from clinical, epiderniological, laboratory and cmperimental studios.

RESULTS P.383

I. Clinical Symptoms

(1) Neuropathic Symptoms, etc.

As ISHIKAWA, 31 UONO, 32 and SEGAWA33 have reported, it is well known

that children and adults from the Saku area show various neuropathie P.384

symptoms. In Figure 10 the various neuropathic symptoms of a group of

100 children and 51 adult cases from the Saku area are compared to a

normal group of 100 children and 50 adults from the sanie Saku area. In

Figure 10 the ratios of symptom occurrence(%) are on the transverse axis,

children are represented by f s, adults by EMMA , patients are on the

left side of the graph, and the control group is on the right side. That

is, regardless of whether child or adult, there are very clear changes:

in the tendon reflexes there are clear aggravations and the emergence of

disease-like reflexes, in the sensory systems there are marked declines in the senses

3.

of position and of vibrations, and in coordinated movement4there is inabi-

lity to stand on one foot. Also observed were dysdiadochokineasia, muscle

atrophy, decreases in muscle strength, and inability to walk (adults).

These things are quite pronounced when compared to the control group.

A 2(2

test of symptoms of the autonomic nervous system showed sig-

nificant differences between children of the Saku area and a group of 100

control cases in excessive fluid intake, car sickness, excessive perspira-

tion, headaches, etc. These examinations were carried out in cooperation

with UONO and SEGAWA. Because there are no essential differences in the

ocular symptoms from those reported by ISHIKAWA and OTO, they will not be

described here. Figure 11 shows the symptoms of the autonomic nervous sys-

tem of the children and those of the control group, and for comparison Fig-

ure 12 shows the symptoms of patients with chronic organophosphate poisoning,

as found in farming areas by Metcalf. 82 It is interesting that reduced

vision comes third behind forgetfulness and languor. Then this is followed

by headaches, muscle aches and excessive perspiration. Because Metcalf is

a brain-wave researcher and neurologist, psychological symptoms such as

forgetfulness and reduced memory are number one, while fatigue and languor

are number two, and reduced vision is number three.

This appears to be due to the fact that no examinations other than

vision measurements were carried out; moreover, such results were obtained

because he did not carry out any modern neuroophthalmological examinations,

and he may have been unaware of them. Problems such as the above arise if

data are obtained by specialists.

(2) ERG

For a patient with the above symptoms, ERG (Electroretinogram) is

sometimes useful for diagnosis. This can be quite useful for diagnosis because

the ERG changes 9before and after PAM9 when a patient with reduced vision due

to acute intoxication has EPN and methyl- or ethyl-parathion found in the

urine or blood. Depending on the degree of intoxication, the supernormal

is sometimes normalized within 3 - 6 hours after PAM is used. 83 The chronic

intoxication cases will be described in this report.

The 2 cases used for illustration here have already been reported

by ONO et al. 83 Both were caused by ethyl- and methyl-parathion;

case 1 is due to direct spraying for 10 years and case 2 is due to ginseng

produced in the Sakudaira area, and Figure 13 shows 3 ERG (20 joules) graphs

for each of the two cases. At their initial examinations both patients

had 0.01 vision (upper one middle one is around 0.5, and lower is 1 .5) ,

which improved gradually upon long-term administration of PAN and Padrine*;

the ERG's were obtained with the same gain, time; the lower case can be con-

sidered to be a quite normal ERG. The upper ERG is often observed in chro-

nic-intoxication cases, and the following is clearly visible: time required

to reach the maximum in the a-wave, a positive "hump" at the a-wave maximum

which might be the overlap of a rhythmic small wave 01 , and delay of the

b-wave. These have commonly been observed to normalize gradually as vision

improves. These abnormalities in the ERG are fully reprodueible, as has

been reported by MAI for animal experiments using Wister rats. This will

be discussed in the section on animals.

(3) Smooth-Pursuit Movements (EOG)

Example 1: Figure 14A shows a case of a very strong inhibition of eye-

ball movement, and Figure 14B shows how treatment with PAN has restored it.

On the patient EOG, the top wave is the input frequency, a horizontal 40°

sine curve at o.5 Hz. Figure 15 shows the step-like waves found for both

eyes directed to the right (upper case) and sometimes when directed to the

*Translator's note: Transliteration of an unidentified term, actual spelling may differ. This also applies to asterisked words otherwise not explained in the remainder of the translation.

P.385

5.

left (lower case); these occur sometimes and sometimes they do not. In

one section, large saccadation is also observed. By January 18, 1973

ihe pattern has normalized, and by April 18, 1973 it bas become almost

normal. In this case, because there were some abnormalities in the

convergence , pupils, AC/A and A/A, it can be considered that there must

have been some abnormalities during an early period due to accumulation of

acetylcholine in the inner-brain stem eyeball motor center. This obser-

vation will be quite helpful for future investigations regarding the role

of the occurrence of step-like waves.

(4) Pupils and Pupilogram

For acute intoxication it is quite well known that intoxication by

parathion results in myosis of the pupils which become pin holes, but there

are also cases in the literature where parathion intoxication results in

mydxiasis of the pupils.84

For chronic intoxication it bas been reported

that the pupils are dilated or are of normal size.85

Figures 16 and 17 p.386

are infrared photographs taken by ONO in a dark room using various intensi-

ties of light stimulation. Figure 16A shows the extremely myotic pupils,

"pin-hole pupils", of a patient whose vision was rapidly reduced when he

was spraying malathion, and came to the hospital just before losing bis eye

sight. For such an acute case the pupils do not react to light stimulation,

and they often remain myotic even in a dark room. However, for a case of

chronic intoxication of malathion ( a farmer --- malathion was detected in

the urine and in the blood), the pupils were dilated in a dark room and

reaction to light was weak. As shown in Figure 16B, the differences are

not great even when the light intensities are varied. This is the most com-

mon occurrence found in the Saku patients. This is also similar to when

ethylthiometon was used on beagle dogs, and Figures 17A and 17B show the

A

6.

control case and the chronic-intoxication case. These observations become

clear if pupilograms are examined. p.387

While the details of the data are described in the literature, Figure

18 shows the diagrams. The top case is for light stimulation of one second,

the left side gives the pupil diameters, and their pupilograms are shown

for chronic-, normal- and acute-intoxication cases. For the acute case

the amplitude of the reaction to light is small, the latent period is short

and once myosis occurs restoration to the original state is slow. For the

chronic case there is a prolonged latent period, the amplitude is decreased,

and the myosis period is prolonged. These observations are very important

for the diagnosis of organophosphate intoxication. One of the most impor-

tant clinical observations is when the administration of Mydrin* P or M

does not readily produce dilation of the pupils.

(5) Refraction and Astigmatism

These aspects are described in detail by TOKORO, et al.86

Also, because ISRIKAWA et al.31 have reported in detail on the refrac-

tive values for Saku patients after being treated with midorin and cyclogyl,

the data described here will be mainly on corneal curvature diameters. For

this investigation we have received the co-operation of the ophthalmology

group of Tokyo University. It is well known that Saku patients ordinarily

show strong astigmatism regularis and this can be observed withielacido's

keratoscope; Figure 19A shows a normal cornea with more or less uniform p.388

image widths and intervals, and the vertical and horizontal meridians cross

at right angles. However, for a Saku patient, a 9-year-old girl with astig-

matism regularis, the horizontal - 2.5D and vertical - 12.0D show an astig-

matic condition which appears like a rugby bail placed sideways, as shown

in Figure 19B (slightly.larger than actual size). Although this example

7.

shows only the left eye, the trend is that, in the first and second quadrants

the Placido images are quite squashed in shape, but there are only a few

squashed shapes in the third and fourth quadrants. Although the horizontal

meridian is almost a straight line running on the cornea,

the upper and lower vertical lines (first-second and third-fourth quadrants),

particularly the line in the third and fourth quadrants,are convex lines

from the left side of the figure, and the upper and lower lines cross with

an extremely deformed shape. That is, this showed that the cornea is de-

formed towards the front and has a pushed-forward shape. Next, data are

shown of the corneal curvature radii converted to diopters for results ob-

tained after the administration of dilating agents. Figure 20A is for the

"horizontal meridian" and B is for the "vertical meridian" corresponding

respectively to the horizontal and vertical axes in the figures. The ver-

tical axes show the number of eyes of patients who were diagnosed as organo-

phosphate intoxication cases, and female (middle), male (bottom) and nor-

mal (top) cases are shown. The analyzed data from these histogramSand

average values, standard errors, Fisher-test values, and significance

particulars were calculated. The normal eyes show almost symmetric binomial

curves, but on the horizontal axes there are slight leftward shifts in both

females and males, and on the vertical axes the diopter values are shifted

to the right, particularly in the females,

that is , astigmatism regularis occurrence is strong in

females. Results from animal experiments are shown in another section. It

is also similar for intraocular pressure and optic axis length. 86 With

the horizontal males and females and the vertical females the radius of curva-

ture is significantly smaller (at the 5% level) than that of the control group.

8.

(6) Organic Chlorides in the Blood

The organic chlorides DDT, BHC, Dieldrin , Heptachlor, and Endrin

etc. were measured by FID and ECD gas chromatography on 20 Saku and Kita-

zato patients, but no differences from the control group were found;conse-

quently these were eliminated as a cause of this disease. 25 Results on

mercury in hair, fluorine, etc.

reported. 25

were also the same as those already

Thus, the results obtained here were exactly the same as those reported

by ISHIKAWA25

, and simply confirmed their results. Astigmatism is the

result of corneal deformation and ciliary muscle contraction.

(7) Criteria for Diagnosis

Criteria for diagnosing this disease were investigated by a research

group started in 1971, and the efforts of this research group were adopted

by the Ministry of Public Welfare at their June 16, 1973 conference on

"Research concerning the determination of diagnosis criteria used on child- p.389

ren with reduced vision which frequently occur in certain areas". Table 6

shows this result. Its content is divided into mandatoryr and reference

items, with the mandatory items containing things such as ocular symptoms,

test observations, treatment effects and systemic symptoms. The important

thing here is that the diagnostic decision is a result of direct verifica-

tion of organophosphates in the blood or urine according to FPD gas chro-

matography. If this value is found repeatedly to be more than 0.01 ppm,

along with other clinical symptoms, there is probably no mistaking that it

is definitely this disease. If the amount of organophosphates are more

than 0.1 ppm in blood everytime, the patient definitely has this disease.

These symptoms are usually very severe ,and a high degree of optic and per-

9.

pheral neuropathy are seen so that everyday activities are almost

impossible to carry out. These diagnostic criteria were reviewed by re-

presentative neurologists from universities across the country and also by

Professor Walsh. It is hoped that from now on these criteria will be used

so that the detection of victims and the treatments can be promptly under-

taken.

(8) Diagnostic Steps for Organophosphate-Intoxicated Victims

Since the main complaints of a victim in many cases are reduced vision,

numbness in the legs, and difficulty in walking, diagnosis of this disease

requires-that various other diseases must be excluded; these other ailments

are ones such as brain tumors, Arachnoid inflammation, inflammatory retro-

bulbar optic neuritis,Leber'sdisease, infantile optic atrophy,-Devicts disease,

retinal pigment degeneration, various drug intoxications, various types of demyelinating

encephalopathy, vascular diseases, and others. Of the diagnostic steps

shown below, it is quite important that the environmental histories are

ascertained, particularly details about contact with agricultural chemicals.

That is, questions such as the following must be asked in detail:"are agri-

cultural chemicals used at home ?", "are insecticides used at home ?",

"are they used in the garden ?", "is there a nursery nearby and is there

always a strong odor ?", "are juices of the same fruits and vegetables drunk

throughout the year ?", "is water used from a well which is near farming

fields ?"7 and also questions regarding possible contact with organophos-

phates at their jobs. For other clinical examination methods various lite-

rature references58 should be consulted, but the measurement of residual

organophosphates58,77 in the blood and urine will be a determining factor

in the diagnosis. This point is an important additional detail which is

10.

given here in this report. The systematic diagnosis procedure is shown

below.

Diagnostic Steps

Reduced vision 1

Environmental history (contact with agricultural chemicals)

Field of vision, intraocular pressure,reflex and ophthalmo, ERG, flicker

dilation (cyclogyl)

ERG, dark adaptation, intraocular pressure, EŒ

Eye-ground examination, fluo*

Neuropathic examination, brain-wave test

sampling of blood and urine

Liver functions, blood, cholinesterase

Gas chromatography

Diagnostic decision

II. Animal Experiments (beagle dogs)

1. Changes in Refractive Systems

Although it is well known that eye lotions cause myopia91 , with regard

to organophosphates and the occurrence of myopia, particularly, for the

occurrence of myopia due to minute doses of organophosphates for prolonged

periods, the investigation reported here is the first one of this type.

Although it depends largely on the constitution of each dog, the number of

days, from the beginning of organophosphate administration to the clear

occurrence of myopia, is about 10 months after the start. This is shown

in Figure 21A. The dotted line and the O's represent the 5 control dogs, and

* Translator's*note: Evidently an abbreviation.

11.

the solid line and the O's represent the 5 treated dogs; these lines are

averages for horizontal and vertical axes, and also, for both the control

and treated dogs, the values are the averages of those obtained for the

left and right eyes. Here, in order to simplify the Figure, only 6 dif-

ferent experimental points are shown; and it was found that the organo-

phosphate-administered dogs contracted astigmatism with a 5% (Student's t-

test) risk ratio after 5 months, and with a 1% (the same) risk ratio after

10 months. The vertical data (bars) in the Figure show the standard errors.

Although the administration of organophosphates was stopped after 24 months

for the 5 treated dogs, there was no tendency of a return to normal once

myopia had occurred. As an average value the myopia stage is about 2 - 3

diopters, and this is very interesting because the value in diopters is

very similar to that of pseudo-myopia, which bas been receiving much atten-

tion these days in Japan.

2. Changes in the Horizontal and Vertical Axes and their Relationships

to Dose Response and Dose Duration

It is known that corneal astigmatism occurs in organophosphate-into-

xicated victims,86

but what is it like when the refractive values are looked

at: Figure 22A (horizontal refractive values) and Figure 22B (vertical

refractive values) are shown. In this Figure the values obtained before

the start of administration were taken as zero, with each measured value

shifted to 0 diopters, and these were plotted with the horizontal axis being

the number of years, the vertical axis in diopters (minus values are at the

top); the numbers on the right side are average amounts administered to

each dog per day in mg/kg; the average values for the control and the stan-

dard errors are also shown.

12.

As shown in Figure 22A, the highest myopic tendencies in the hori- p.391

zontal values are changes of about 4 diopters for the dogs given 1.0 mg/kg,

about 3.0 diopters for the dogs given 1.5 mg/kg, and about 1 diopter for

a dog given 0.5 mg/kg; this latter dog is not much different from the

normal control ones.

In Figure 22B the changes in the vertical axes show a more or less

similar trend, but the dogs administered with 1.0 mg/kg and 1.5 mg/kg,

one of each, show changes of about -4.75 diopters and -3.75 diopters, res-

pectively. These values are the ones after l'4vdrin* application, and the

values before are much larger. This Figure reveals that a much stronger

myopic effect occurs in both the horizontal and the vertical for the dog

administered with 1.0 mg/kg than for the dog administered with 1.5 mg/kg,

and the horizontal axis change is rather large for the 0.5 mg/kg administ-

ered dog. That is, because there are individual differences in the changes

in refractive values (occurrence of myopia), with the degree of reaction

being different depending on the dog, and also because a dog given a larger

dose does not necessarily show a stronger reaction, the determination

of doses is very significant in research on the refractive systems.

Futhermore, because even one of the two dogs administered with 0.5 mg/kg

shows myopia of about -2 diopters in both the horizontal and vertical,

safe amounts of ethylthiometon to prevent myopia are 0.05 mg/kg if it is

1/10 the amount,and 0.01 mg/kg/day (values are the same in ppm) if it is

1/50.

3. Changes in the Corneal Curvature Radii (converted to refractive

values)

Although there were no differences in the average values of the corneal

13.

curvature radii from those of the control for the horizontal axis, in

contrast there were apparent significant differences (5% for Student's t- test)

from those of the control in the vertical axis after 13 months from the

start, and this trend continued until around the 20th month. This is shown

in Figure 2113. The average values and standard errors are roughly shown.

Although the horizontal axis becomes flat for both the control and the

treated dogs as growth occurs, growth is halted in the vertical axis, with

the curvature radius hardly changing until about one year after the start

of administration. From this it was ascertained that the occurrence of

corneal astigmatism is most prevalent at around one year and then a normal

value is gradually reached. The advance of myopia and of corneal astigma-

tism, as described above, is typically shown in Figure 23. That is, it was

found that the advance of myopia and the amount administered are correlated;

although it is not clear for the 0.5 mg/kg case, for the 1.0 mg/kg (1.5 mg/kg)

administered case, as administration is continued the advance is gradually

more as time passes. Contrary to this, for the advance of corneal astigma-

tism the greatest astigmatism was detected after administration for about

one year, after which the level of the control case was gradually approached

as growth progressed. For these experiments on refraction the amounts

administered cannot be increased without limit, and it is necessary to give

optimum amounts. In general, it would be thought that stronger myopia would

occur as the doses are made larger, but this is not the case. As reported p.392

by Ishikawa, needed for precision are examinations of accelerated and sup-

pressed occurrences of the refractive system when Edinger-Westphal nucleus

or ciliary ganglion are stimulated.

L, Cholinesterase Values

Among the ChE values of the blood, there were no observed decreases

in the serum ChE for the dogs treated with ethylthiometon. Hikida et al. 79

have reported in detail on this point using various organs; the clearest

change was the blood-corpuscle ChE, that is, the acetyl-cholinesterase

value. This is shown in Figure 21C. On the vertical axis the % values

for each of the dogs is shown, with the value (average of three measure-

ments) of the initial period taken as 100%. The values for the 5 control

dogs ( 0-0 ) and for the 5 ethylthiometon-administered dogs ( )

are shown; the time,in months, is shown on the horizontal axis. Each

point shows the average value for the 5 dogs and the standard errors.

The value of acetyl-cholinesterase (AchtChE) decreased 50% after administ-

ration for 4 months and it is significant to less than 1% with the control.

When administration is continued, in the 5th month it decreases 75% on the

average, and after that the value remains constant. This Ach•ChE is well

correlated to the myopia of Figure 21A, and even in correlation tests of

different types, strong correlations were seen after the 5th month. That

is, when the administration of organophosphates was continued, it was found

that there was a strong correlation (under 1%) between the advance of myopia

and the drop in the Ach.OhE ratio.

5. Results from Pathological-Histological Investigations

(1) Ciliary Muscles

Readily apparent with an optical microscope were the following

observations on the ciliary muscles: light swelling of the muscle fibres,

apparent dropsical swelling of the ciliary epithelia, and vacuole degene-

ration. When these were further investigated with an electron microscope,

1 5.

the normal ciliary muscles have beautiful arrangements as shown in Figure

24A, but for the organophosphate-administered dogs stratified tectonic

substances with almost circular shapes were observed everywhere in the

muscle cells) as shown in Figure 24B for a dog with myopia of about 3

clapiers. Although similar shapes have been found in the retina after

the administration of chloroquine, this was the most often recognized and

the most specific finding in the ciliary muscles of organophosphate-admi-

nistered dogs. These were never found in the control group. This change

was more obvious in the administered dog which had myopia of about 4.75 D;

this is shown in Figure 24C. There is clear dropsical swelling in the

muscle cells, and the above mentioned stratified tectonic substances are

more clearly visible. Thus, for the ciliary muscles.l the degeneration is

possibly centered at the endoplasmic reticulum of the muscle cells, and

the dropsical swelling of the cells was found to be one of the most promi-

nent changes. The occurrence of myopia is the result of changes in the

cornea and of this phenomenon.

(2) Pigment Epithelial Cells

For the administered dogs there were some interesting findings in the

pigment epithelial cells of the retina. Figure 25A is a retinal pigment

epithelial cell of a control dog; here visual cell outer nodes* (upper left)

and also normal pigment granules can be seen. In the retina of the dog

administered with 1.5 mg/kg, a large breakdown of the pigment epithelial

cells is found, a beehive-like structure is observed, and also there are

breakdowns of the pigment with the observation of huge dark-colored struc-

tures which are different from the pigment granules. The Myelin-figur **

is the degenerated structure of mitochondria (m) or smooth endoplasmic reticula:

* Translator's note: Literally translated. ** Translator's note: a German term.

16.

In the ganglionic cells, vacuolation was more pronounced in the admi-

nistered dogs, but the changes were not very large when compared to the

control dogs.

(3) Optic Nerve

Initially for the optic nerve, the non-medullated nerve was examined

in the region of the papilla, and structures were observed which sug-

gested early deterioration. Figure 26A is a view of the non-medullated

nerve of the optic nerve of a control dog, and it is normal; but, as shown

in Figure 26B for the dog administered with 1.5 mg/kg, the non-medullated p.401

fiber of the optic nerve was surrounded by tectonic substances which appeared

to be deteriorated mitochondria, and these degenerated forms were clearly

visible. With an optical microscope the findings for the optic nerve are

much clearer.That is,as in the control dog, in the administered dog they

are seen as groups of bundles when hematoxylineocene and toluidine blue

dyes are used; but,particularly for the optic nerve of the administered

dog, these bundled groups are atrophied, the nerve fibers are atrophied

and are fewer in number, there is a vacuolization tendency, and there is

an increase of interstitial tissues; among the nerve systems, these changes

were the most prominent. These things are shown in Figure 27, with Figure

27A being the normal optic nerve and Figure 27B that of the dog administered

with 1.5 mg/kg. Such a pronounced morbid change as this, including the

changes in the ciliary muscles, was one of the severest ones in this ex-

periment. There are no such changes using quinoform.

(4) Outer-Eye Muscle

Changes in the outer-eye muscle have been reported in detail by Kono

et al.,87 and the changes were pronounced at the ganglionic junctions.

17.

In Figure 28A the ganglionic juncture of the normal control case is shown.

The muscle side and the nerve side are normal. The same section for the

dog administered with 1.5 mg/kg is shown in Figure 28B; here the expanded

vacuolization of the synapticfolding and irregularities in the mitochondria

of the sole plate were obvious.

(5) The Sciatic Nerve

The sciatic nerve, which is one of the large nerves in the peripheral

nerve system, will be described here. Figure 29A shows the normal sciatic

nerve, and it can be seen that each of the fibers is regularly arranged.

Similar to that found in the optic nerve, the sciatic nerve of the dog ad-

ministered with 0.5 mg/kg showed the following in some sections, as seen

in Figure 29B: extreme randomness of the nerve fibers, increase in inter-

stitial tissues, doubling structure of nerve fibers, fattening of the medul-

lary sheath. The interstitial tissues were Schwerin cells and non-medullated

nerves, and so on.

(6) The Calf Nerve

Because the calf nerve is a sensory nerve of the peripheral nerve

system, the changes were very distinct, as expected. No abnormalities were

found in the control dogs. In most of the administered dogs extreme thick-

ening of the medullary sheath and increases in interstitial tissues were

clearly seen (Figure 30).

Even though these changes were prominent, in the early period of admi-

nistration this dog had difficulty in walkingeointing to a crippling effect;

but it recovered quickly and this effect was not observed after that. Given

next are the examination results on the calf nerve of a patient (whose main

complaint was visual difficulty), who was actually diagnosed by us as being

18.

a case of organophosphate intoxication. This is shown in Figure 30A, and

it was very similar to the experiments on beagle dogs in that extreme

thickening of the medullary sheath and increases in the interstitial tissues

were observed. Thus, even in nerve systems which look normal from the

outside, it is found that appreciable damage bas occurred and that degene-

ration is proceeding. Here, due to space limitations, the results found

for the other sections will not be described.

(7) X-ray Microanalyzer

(1) Recently, for toxicological research frequent use is made of

an electron microscope with an x-ray microanalyzer, which is used to deter-

mine the location of the toxic substances and to determine the content in

the cells. The result for a control case is shown in Figure 31A. This is

of the liver and the result shown is the complete area (photographed sec-

tion) which was scanned with these instruments; the bottom-left figure

shows the appearance of the element peaks for various materials from the

varied locations of the complete scan area. The right-band figure is the

observed result for point A as shown by the arrow. As described before,

K, Cu, Os, Cl, and Ca are respectively found. Figure 31 1 is also of the

liver, the results for the administered dog (1.0 mg/kg). The top-left

figure is the tested cell, and the arrows 1 and 2 are the positions where

analyses were carried out. The analysis photo shown to the right of the

figure shows the presence of Cu, P, S, Cl, K, and Ca; it can be seen that

the P and S peaks, which were absent in the previous photo, are respectively

larger. For the arrow points 1 and 2, the P and S, that is, phosphorus

and sulfur, are clearly larger. Thus, by utilizing these instruments in

this way, the presence of toxic substances in the cells of various organs

19.

becomes evident.

(2) Ciliary Muscles

Similarly, the investigational results with the x-ray microanalyzer

on the ciliary muscles, which have shown large changes, will be given.

These investigations were carried out with the cooperation of Mr. Miyazawa

from the central microscope examination room at Kitazato University.

Figure 32A is the result obtained for one section of the control-case

ciliary muscles. These data are the scans of one section of the ciliary

muscle, that is, one horizontal plane, and of the entire area scanned.

The bottom left side is the result obtained from scanning one section, and

on the right side is the result obtained from scanning the whole area. Cu

is the peak used as the mesh. Figure 32B is the data obtained from the

administered dog which was in the diseased condition. The bottom-left

figure is the result of a one-section scan, and the bottom-right figure

is the result of the complete-area scan. For the administered dog, incre-

ases in P, S, and Cl are clearly seen. With regard to the peaks which are

not detected in the control case, that is P and S, it is quite clear that

the P and S are those contained in the structural formula of ethylthio-

meton. When the above is all brought together, the pathological-histolo-

gical observations can be summarized as shown in Table 7. That is, these

changes are found in almost the entire body and in all the administered

dogs; although there are variations according to the amount administered,

changes were found in the central optical nerve, external knee-like struc-

ture, optic nerve, spinal cord, peripheral nerve, liver, pancreas , spleen,

alimentary canal, thymus, autonomie nerve ganglion, and in the muscles.

For the muscle system refer to the report by Kono. 87

p. LtOLi,

20.

(p ) ERG

Regarding changes of ERG due to organophosphate intoxication, experi-

ments with parathion and Baytex have been reported. 'mars original report81

should be consulted with regard to the detailed methodology; however, it

will only be mentioned that the above described clinical ERG findings are

definitely reproducible in the animal experiments. The animals used are

Wister rats, the ERG is at 2 and 20 joules and using scotopic ERG with 3

hours of dark adaptation. Just one Baytec injection was given hypodermically

with a microsyringe; after that the ERG was followed from the 4th to the

45th day, but the description here will be mainly of the 4th-day 20 ioule

ERG. The ERG is shown in Figure 33. In comparison to the control case,

the ERG's appear to be different between the 0.5 mg/kg (A) and the 50 mg/kg

(B) administered cases. That is, for administration of a small amount,

an extremely "supernormal ERG" is observed where the amplitudes of the a-

and b-waves and of the rhythmical small waves are increased, and the latent

period is shortened; in contrast, for the administration of a large amount,

a "subnormal ERG" is observed where the amplitudes are small and the latent

period of each wave is lengthened. In clinical cases these large ERG are

especially found in acute intoxication cases, as shown for the terminal

patient in Figure 13. It is well correlated with the subnormal ERG with

a positive hump in the a-wave. For each amount administered, Table 8 shows

the average ERG obtained for 3 ERG's on each of 4 animals using light sti-

mulation of 20 joules; the standard errors are also shown. Significant

differences are observed using the Fisher test with risk ratios of 5% for

and 1% for• go,K). It is clearly seen that the ERG is supernormal

when the amount administered is around 0.005 mg/kg - 5 mg/kg, and the ERG

21.

is subnormal when the amount is greater than 25 mg/kg. For amounts inter-

mediate between these ranges, the ERG's are near normal values, but in

some locations the ERG is not normal. By recording these ERG's by dif-

ferent methods, the abnormal can be distinguished from the normal; this

aspect will be published later. The typical ERG's from the above are

shown in Figure 34, which has each of the typical wave-shape figures super-

imposed. For chronic intoxication in both man and animals, many have a

subnormal ERG. When the amount administered is increased, a non-recordable

ERG results. The important finding determined from this experiment is that

because Baytex is weakly toxic with the LD50 value greater than 300 mg/kg,

an assessment of the acute intoxication experiment is obtained. From this p.405

data, it was found that a toxic effect on the optic-nerve system can already

be seen at 0.005 mg/kg. The toxicity of organophosphates on the sensory

system is very sensitive, Vio4 the LD50 value of 50 mg/kg. It should be

noted that this fact is very meaningful for determining the "toxicity asses-

sment" of toxic substances on the various organs. The cause of there super-

normal and subnormal ERG's is very much related to the control amounts of

acetylchblinesterase in the retina. 88 Regarding this aspect, refer to the

research of Miyata.

(9) Liver Functions

We will be publishing reports on the blood, the blood sugar values,

and various liver functions.

Literature Survey

(1) Optic-Neuropathy (Table 9)

In the past there have been quite a number of reports of investiga-

tions on the effects of organophosphates on the eyes and on the nerve

22.

systems. Narrowing down the large number of chronic and acute cases

reported to those of optic-neuropathy, the ones given in Table 9 are obta-

ined. Shown in chronological order are the respective chemicals, differ-

ence between chronic or acute cases, and reported clinical symptoms. These des-

criptions began in 193289 and by 1971 there were numerous reports. Since

there are too many reports on myosis and on ciliary-muscle spasms, these

are not included. For cases given here, the acute cases are the results

of a single contact with a large mount of organophosphates, the subacute

cases are the result of one or more contacts with organophosphates, and the

remaining cases not included in the above are the chronic cases which are

the result of a long period of contact with minute amounts of organophos-

phate.. The main chemicals reported on are the following: TOCP,89

para-

thion, 92 mipafox, 91 malathion, 101 DFP, 102 EPN, 97 TOAP, 99 phospholine

iodide. 105 The various organophosphates described here in this report are

parathion, malathion, diazinon , EPN, ethylthiometon, baytex, metasystox,

sumithion, and others. When surveyed historically the occurrences of the

following have been observed 94 : pupil disorder, control disorder, optic-

nerve engorgement, optic-nerve atrophy, myopia, astigmatism; the systemic

p.406

symptoms which have already been reported are peripheral neuropathy, reduced

memory and attention span, 103 abdominal-region symptoms 110 and autonomic

nerve symptoms. 108 hum an are all han symptoms. From these things it is

clear that disorders due to organophosphates appear in the brain, eyes,

peripheral nerves, muscle systems, and they show autonomic nerve symptoms.

(2) Less-Familiar Symptoms (Table 10)

Among the symptoms of organophosphate intoxication, the less-familiar

but important ones which have been reported in clinical cases as well as

23

in animal experiments are: reduced reabsorbing power of the tubuli

renales; disorder of amino acidmetabolism; reduced resistance towards viral

infectionsweakŒmo against stress; abnormalities in the function of supra-

renal gland; proneness to allergic diseases; increased susceptibility to aphtha,

ulcers, stomatitis, dermatitis; asthma; and cancer symptoms.

Furthermore, besides those we have already mentioned, the eye symptoms which

are found are cataracts $ embolism Of the centre artery of the retina, mydri-

asis, ptosis, supranuclear ophthalmoplegia, myopathy, myositis, etc. 117-123

These are only some of the disorders of the body caused by organo-

phosphates, and because of space limitations, the investigations on other

organs and the biochemical research have been deleted. Moreover, clinical

and basic studies of this kind, which are broad but precise, have not been

carried out abroad; this research is quite original.

DISCUSSION

1. Concerning Epidemiology

This research has clearly shown that after 1968, when indiscriminate

spraying of large amounts of malathion was begun in the Saku area, victims

began to appear. Before that time there was very little occurrence of the

disease because the usage of organophosphates was rare; but in 1970 (this

is two years after the spraying of large amounts of organophosphates) the

number of victims was three times more than in the previous year and a

correlation between the victims and the spraying is highly suspected. Because

of our warning the amount sprayed in 1971 was less than the previous year

and resulted in slightly fewer victims, but in 1972 and 1973 as the spraying

was increased, new victims were still appearing. Next, the monthly ratio

of patients examined throughout the year is concentrated during the period

p.407

24.

April to September, when large quantities of agricultural chemicals are

being sprayed, and the numbers decrease during October the busiest harves-

ting season; and it is interesting that the number of people with reduced

vision increases during November, a time when farmers are not as active,

because of residual organophosphates from other seasons in the water

and food. Furthermore, the results of the measurements of serum cholin-

esterase values for the patients (victims) showed that, in relation to the

normal value of 1.05± 0.20, the adult patients had lower serum values from

February to October, and in children they were lower from January to October;

these were confirmed to be reproducible by repeated measurements.

Although there may be general loss of health during the summer, the fact

that the serum cholinesterase values are the lowest in July is also consi-

dered to have a close relationship to the spraying of agricultural chemicals.

Although there were some fluctuations in the relationship between the resi-

dual amount of organophosphates in the drinking water for the population

of the area and the number of victims, it was very clear that the victims

were found concentrated in areas where the amount of organophosphates detected

was higher, and did not depend on the quality of water. Also, the fact

that there are almost no victims in mountainous areas where helicopter spraying

is not carried out, suggests that the presence of residual organophosphates in

the water is closely related to the occurrence of victims, Further, taken as

a wholei except for the Shokai region, the southern Saku area has a small

number of victims, and these are found mainly in the area east of the Chi-

kuma River, and because of the fact that this area bas volcanic-ash soil

while the west side of the river bas clay soil, one cannot discount the pos-

sibility that pollution by organophosphates in ground water appears more

25.

in the regions with volcanic-ash soil. Moreover, there are many apple

farming fields in the foothills of Mt. Asama, there is a section north of

the city of Komoro which is considered to be the water source for the Saku

area, and the drinking water in the southern Saku area is via a water sys-

tem from the Mt. Yatsuga area; these factors may have been responsible for

the differences in the number of victims. In any event, in areas where

large amounts of agricultural chemicals are sprayed,the figures ar5for

children,the occurrence of an average of 2.19 victims per 1,000 population,

while the detected organophosphatesare 0.75 ± 0.99 ppm; in other areas

where similar amounts of organophosphates are used the numbers would proba-

blybe not much different. Including the population in the mountainous re-

gions, the frequency of child victims, for the ca. 180,000 total population

of the Saku area, was found to be 17.2 per 10,000; it is possible that this

number will match those of other farming areas where about the saine amounts

of agricultural chemicals are used. Furthermore, an interesting observa-

tion was that the serum from a child victim from the Saku area showed

detected organophosphate amounts of 0.01 to 0.05 ppm in August 1972, but

the same patient showed an average of 0.1 to 0.5 ppm, about 10 times more,

in November of the same year. This is interesting because it appears that

the organophosphates used during the summer have been taken in via the

water, food, direct contact, etc.

2. Organophosphate Detection by Gas Chromatography

For the detection of organophosphates in the urine> there are the inves-

tigations by Saito and Kubo, and we have described here a method for detec-

tion in the blood, but even using highly sensitive gas chromatography as

the method for detecting organophosphates, it is impossible to detect all

26.

of the 80 some odd organophosphate agents presently in use. Moreover, it

has been proven experimentally by Miyata that substances such as organo-

phosphate metabolites (P=0, P=S), the decomposition products of the organo-

phosphates produced in the body, show strong anti-cholinesterase activity ;

because these metabolite peaks are not included in the actual measured

values of this experiment, addition of these peaks, that is analogs, would

result in much larger values. The values presently obtained by us are

quite accurate, but if the errors in measurement were .±10%, and because

the victims with the highest detected amounts have about 1.0 ppm and the

lowest detected amounts are 0.01 ppm, even if the values were 10% less, the

patients would have amounts between 0.1 and 0.001 ppm of organophosphates

in the blood; it can be considered that this will be a very important

problem in the future. From the investigation carried out at Kitazato

University, the organophosphates found in the urine were more than 1.0 ppm

for a person who had committed suicide and also large peaks between 1 and

10 ppm of sumithion , salithion and DDVP etc. were detected for a victim

who was totally blind and had difficulty in walking. Because the control

cases have less than 0.01 ppm, the values presently obtained for the Saku

victims are in between those of a serious victim and a normal person; they

are medium-level victims on the level-of-seriousness scale. Because dogs

administered with 1 mg/kg (1 ppm) of ethylthiometon have detected organo-

phosphate values in the blood which are greater than 0.1 ppm, a calcula-

tion from this indicates that if a victim bas 0.1 ppm concentration in the

blood, there are 0.01 ppm of organophosphates in the body in some form;

in comparison, because the average value of total organophosphates detected

27.

in drinking water, etc. is 0.75 ± 0.99 ppm, this is an indication of a

general agreement with the residual amounts. Further, the peaks of the

various organophosphate agents reported here are relative peaks, and from

this it cannot be determined whether only these particular agricultural

chemicals remain. That is, there is the possibility that these peaks

might coincide with those of others such as sumithion , diazinon, DDVP, p.408

ethylthiometon, kilbar* etc. which are sprayed in large quantities. In this

regard, we have clinical and experimental investigations in progress using

various other types of analytical methods such as thin-layer chromato-

graphy, gas-mass spectroscopy, ECD, etc, and the results will be reported

in the future. In any case, in order that disorders of the optic and peri-

pheral nerves do not occur, the organophosphate values must be suppressed

so that they are less than 0.01 ppm in the blood and less than 0.001 ppm

in the water and food.

3. Clinical Symptoms

(1) With regard to reduced vision from this disease, many factors

may be considered. The causes of it are the following: first, central

scotoma; second, myopia; third, corneal astigmatism; fourth, retinal dege-

neration; fifth, optic-nerve atrophy; and sixth, disorder of the central

optic nerve. Although each one of these were fairly clear in this beagle-

dog experiment, the symptom might appear with only one of the disorders or

with only some of the disorders, and so it is difficult to directly deter-

mine the disorder state of each of these for a victim. For example, in an

EPN experiment on a dog administered with 5 mg/kg the optic nerve atrophy

was more clearly seen than the changes in refraction.

28.

(2) Visual Field Stenosis

Although a variety of factors cause eccentric visual field stenosis,

in summary it may be considered to be due to the general lowering of visi-

bility in the retina and optic tract. These can be measured accurately

with a Tiibinger * perimeter and then determined, but retinal dis-

orders, as clarified in this report, are also important. The changes in

the ERG from the initial supernormal, then to the subnormal and finally to

the non-recordable is likely due to the degeneration of the retina; moreover,

disorders of the optic nerve arise quite clearly from the pathological

aspect, and also in the research by Honma et al. a high degree of drop-

sical swelling was found around the ciliary squamous body, and assuming

that this moves gradually to the retinal area, it would be the major cause

of retinal disorders due to dropsical swelling. If this continues,

it can lead to detachment of the retina, and indeed, such victims are found.

(3) Refractive Abnormalities

It is widely known that organophosphate agents cause myopia, and

myopia results mainly because of corneal astigmatism and dropsical swelling

of the ciliary muscles. Therefore, it was found in this experiment that

myopia already occurs with tlie administration of 0.5 mg/kg/day of ethylthio-

meton, which is an intakeof 0.5 ppm per day when converted to ppm; if the

safe amount is 1/10 of this value, it would be 0.05 ppm per day, and if the

safe amount is 1/50 of this value, intake per day must be less than 0.01 ppm

so that myopia does not occur. Thus, unless the tap water in the whole

country is found to contain less than certain amounts of organophosphates,

for instance 0.01 ppm of ethylthiometon, there is a good possibility that

29.

myopia will occur. In the Saku area the number of occurrences of myopia

among elementary and junior high school students bas increased rapidly

since 1954, the year the use of organophosphate agents began, and there

was a 2.5 fold increase from 1961 to 1965; this indicates that relating

the use of organophosphate agents to the astonishing number of myopic

patients in Japan will develop into a major problem in the future.

(4) Ocular-Movement Disorders

Although a distinct supranuclear paralysis due to organophosphate

agents has been reported by Ono and external-eye-muscle myopathy has been

reported by Kono, the ocular movements generally appear outwardly normal

with this disease. However, when tests on smooth pursuit movement with EOG

and on fixed sight quiver with a XYtracker werecarried out, distinct cha-

racteristic disorders were found in these victims. 125 As Metcalf has also

pointed out, these phenomena are disorders considered to be due to excess

accumulation of acetylcholinesterase in the ocular-movement nerve system

of the deep-brain stem or in the upper-central nerve system of ocular move-

ments. In fact, Hikida et al. also carried out biochemical measurements

of acetylcholinesterase in beagle dogs, and clearly found that the activity

values of acetylcholinesterase are extremely high in the brain-stem section

compared to any of the other brain systems. It is also high in the cere-

bellum, but this may be related to the presence of disorders in the equi-

librium function.

(5) Pupil Disorders

As reported by us, the pupil abnormalities are miosis for acute into-

xication cases, and mydriasis or normal pupils for chronic intoxication

30.

cases. For some symptoms, it has been repeatedly observed that pupils in

the mydriatic state become strongly miotic upon treatment, then become pin-

hole pupils, and finally return to normal size with continued treatment.

This effect is the same as reported by Davis. 111

(6) Optic Nerve and Retina

In animal experiments it is often observed that in the optic nerve

there is a clear pathological degeneration at the retrobulbar section, but

the pupil appears normal. Also, depending on the symptoms, there are cases

in which strong congestion, temporal discoloration and atrophy of the optic

nerve are found, and the papillae are found to be irregular. There are

even a number of cases of brothers who have the same intoxication, with

one showing congestion and the other atrophy. The simplest method for cli-

nically detecting abnormalities in the optic nerve disk is by decreases in

Flicker values, which are particularly sensitive in the red. The expansion

of Mariotte's blindspot and lengthening of the latent period of VER are

also useful at times. A case has been reported in which the completely

paled optic nerve returned to its normal color upon treatment; along with p.409

the regaining of sight, the color was totally improved. Describing the

above results collectively, in the early stages the optic nerve exhibits

congestion and temporal discoloration ewhile in the final stages atrophy

occurs and then loss of sight. Also, for the retina the changes are dif-

ferent and depend on the medications used and on the symptoms, but in many

cases the course taken is an early dropsical swelling condition followed

by atrophy and degeneration. Oto and Ichii et al. have reported in detail

on these long-term effects.

31

(7) Neuropathie Symptoms

Characteristic of this disease is that peripheral neuropathy occurs

mainly in the vision sensory system and in many cases it takes the form

of a dying back neuropathy114 which gradually rises upwards; also in this

research, degenerations at the sciatic nerve and calf nerve were the most

highly visible, experimentally and clinically. It can be assumed that

accentuation oor weakening of the tendon reflexes and the occurrence of

diseased-like reflexes are the result of intoxications of the central

nerve by this disease. Detailed patholegical investigations on the spinal

cord and on the central nervous system will be reported later.

32

4. Cholinesterase Values

As has been described, the cholinesterase values are measured by the

Shibata-Takahashi method or by the Michel method; if a line is drawn at

0.8ppH/hour, most values are normal. However, if this is measured in the

Saku area in July, more than 40% of the patients have values of less than

0.8. Since the presently so-called low-toxic organophosphates (baytex,

salithion , and kilbar*) often reduce only the blood-corpuscle cholineste-

rase, that is, acetylcholinesterase, it is very risky to clinically diagnose

this disease only by the serum values. The blood-corpuscle cholinesterase

is clearly lowered right after contacting the agricultural chemicals, with

recovery of activity usually slower than for the serum values; but because

recovery is observed within several days for a single contact, this disease

cannot be completely discounted even if this value is normal for patients

whose clinical symptoms are determined to be after effects. In the beagle

dog experiment the dogs administered with averages of 0.5 to 1.5 ppm show

decreases to less than 50% after 4 months of administration. By continuing

administration the value is lowered about 80%, but by stopping administra-

tion the positive clinical symptoms remain while the blood-corpuscle cholin-

esterase value returns to normal. We are presently carrying out repeated

investigations of blood-corpuscle cholinesterase values on a number of cases,

and will be publishing the accumulated data of more than 1,000. Also with

regard to the problem of relating the occurrence of this disease to indivi-

dual factors, here the isozyme of cholinesterase, in particular abnormali-

ties of the C-4 band zone have been found in patients, and this will be

solved in time.

33

However, the research of Hikida et al. showed that even if the serum

cholinesterase values in flowing blood are normal, the cholinesterase values

are generally found to be reduced to almost zero in the liver; thus, if by

a liver bioassay"tissue can be removed, and if the pseudo ChE values such

as its serum cholinesterase value in the liver can be measured, it can be used

as a definite diagnostic test for this disease.

5. Liver Functions

It is known that routine test methods used in general clinical exami-

nations show almost no abnormalities for cases of chronic intoxication due

to organophosphates. 126 However, there are some reports in which the fol-

lowing have been observed; increase of alkaline phosphatase2 1,129 increase

of creatine phosphokinase, 127 increase of lactate dyhydrogenase 115 ; also

found were lowering of vitamin A value in the blood, 128 increase in the

amount of total amino acids in the blood, 117 etc.; thus, there is the pos-

sibility that these can be used as supplementary tests for diagnosing this

disease. However, these findings also vary considerably and depend on the

organophosphate agent used. A definite diagnosis can be made if the amount

of organophosphate residues in the blood or in the urine are greater than

0.01 ppm. In many instances no abnormalities are observed in other data on the

blood, in the cerebrospinal fluid and in the electrocardiogram.

6. Brain Wave Observations

According to the reports by Metcalf82 and by Taneda et al. 130 of the

Department of Psychoneurosis, Kitazato University, many patients show abnor-

malities in their brain waves. For example, from 24 patients hospitalized

at Kitazato University (17 patients due to agricultural sprays, and 7

34

patients due to other causes) the findings are: slow-wave tendencies such

as abnormal e-waves, a sleepiness tendency, and 6 bursts, etc; and, in

general, abnormal epileptic brain waves are observed. Metcalf et al. have

also described the abnormalities in the brain waves as being quite meaning-

ful in diagnosing for chronic intoxication, and thus it indicates that

brain-wave findings for this disease will provide important information in

the future.

7. Diagnosis of Peripheral Neuropathy

We routinely perform measurements of the conduction rate of the upper-

and lower-limb motor nerves and sensory nerves. For this disease, many

cases are found in which there are delays in the conduction period of both

(nerves) or of one. Again, bioassay of the calf nerve can be used. In this

case the abnormalities can also be caused by other drugs, and this detailed

examinations are necessary to exclude factors other than phosphpUes.

TREATMENTS

If treatment of this disease is begun as soonas it is diagnosed, even

a serious victim, who has complete loss of sight and loss of reaction toward

light, can regain his sight if it is still at an early stage. The treat-

ments are based on the use of atropine, padrie,glutathione and PAM. The

treatment of serious cases, medium-level intoxication cases, and light

cases are described separately. p.410

1. Serious-Symptom Cases

PAM (500 mg) + atropine (0.5 mg) + 10% glucose (100 cc) are given once

a day by intravenous infusion. Protein foods rich in vitamin B12 are given

orally. Attention must be given to the following: PAN is quite effective

on organophosphate agents which possess the para-nitrophenyl group, that is,

35

ethyl- and methyl-parathion, EPN, etc., but it is not very effective in

cases where DDVP, baytex, and ethylthiometon are detected. Moreover, 3 - 4

days after the above treatment is begun the clinical conditions sometimes

become more impaired. On rare occasions there are occurrences of severe

diarrhea, paleness of the face, lowering of the blood pressure, kidney-

function disorder, etc. In these cases intravenous infusion of PAM is stopped

and 90 mg of uadrin* is administered orally. The patient will then recover

before long, and then administration of PAN is again started.

2. Unclear Symptoms

Padrin* (45 - 90 mg) + glutathione ornithate (600 - 1,000 mg)/day are

given orally. 500 mg of PAN is given 2 times a week, and the same amount

of glucose (as given above)/day is given by intravenous infusion.

3. Unclear Symptoms

Glutathione ornithate (300 mg)/day, and some vitamine, etce are added.

Use of these treatments definitely improves the clinical symptoms of

chronic intoxication. Even for those which extend over a long period of time,

improvements can be seen at about 3 months after the beginning of treatments.

However, in cases where there is peripheral neuropathy in the lower limbs

and difficulty in walking, mecamylamine (mebacine*) must be given. Also )

for difficulty in walking, One et al. in our group discovered the mecamyla-

mine treatment, and thus there are cases where a patient who had been con-

fined to bed was able to walk and return to society; we have already had

5 such patients here. There is also the case of surprising recovery of

sight from about 0.01 to 2.0. Also for arriving at a diagnosis, assessment

from these treatment effects (therapeutic trial) is also quite an important

131 aspect. Further, an interesting case is one in which, though

36

the optic nervewas atrophied, the sight improved.

Future Countermeasures

From the above, for intoxication of the human body by organophosphate

agents, it was found that in chronic intoxication a wide range of disorders

are centered around optic neuropathy and the autonomie nervous system and

this was confirmed by clinical and experimental studies; but future coun-

termeasures are : 1. The blind belief that low-toxicity organophosphates

are harmless must be changed, and besides only LD 50 values, it is also nece-

ssary to determine toxicity towards the sensory and nerve systems. 2.

The blood-corpuscle and serum-cholinesterase values are measured for children

with the main complaint of reduced vision; for children with low values,

it is necessary to carry out ophthalmological and internal tests near the

nerve systems. If these results are positive, the residual organophosphate

agents in the blood and in the urine must be measured for these positive

persons. 3. Once the diagnosis is made immediate treatment with atropine,

padrin*, glutathione, PAN, etc. must be carried out. 4 • The general prac-

titioner's knowledge of organophosphates needs to be strengthened. 5. Because

the analysis of organophosphates is not well advanced, efficient and accurate

analysis of organophosphate agents with gas chromatography must be carried

out at the national health laboratories of each area. 6. Tighter controls

which must be enforced are needed for the spraying of organophosphate agents,

which at present is in the hands of farmers and under very little controls.

Conclusions

Epidemiological, clinical, and basic investigations were carried out

mainly on 'victims that frequently appear around the Saku area' who have

what are known as organophosphate-agricultural-chemical symptoms of the eye,

37

and detailed results of these investigations were reported; also, the

'diagnostic criteria for chronic organophosphate intoxication ,132 , prepared

by the research group of the Health and Welfare Ministry, was introduced.

By examining the results of research to the present and the past literature

from abroad, the symptoms observed in Saku are determined to be those of

chronic organophosphate intoxication (mainly by malathion, kilbar*,etc.).

This disease occurs not only in Saku but in all parts of Japan, with the

present (July 1973) occurrence ratio being about 17.2 per 10,000 population.

It bas been reported here that recovery from this disease is quite good if

treatments are carried out with early detection, but if treatments are not

undertaken or unsuitable treatments are used, recovery prospects are quite

bad, and for serious cases optic-nerve atrophy or retinal degeneration accom-

panies this disease and leads to loss of sight.

This research was funded by a therapeutic research grant, 1971 and 1973,

from the Ministry of Health and Welfare (research on children with reduced

vision which occurs frequently in certain areas), and by the general research

fund A, 1973, from the Ministry of Education (toxicity of organophosphate

agents toward the visual senses -- its causes and its treatments).

EEM P77

I222

10

ET.:1=7. 'M. • 77.1

10

4,x (children) cm (adults)

20 10 3p

muscle atrophy

muscle fibrillary tic systems reduced muscle strength

f

ri j rg411-- e• I

Figure 10: Neuropathic Symptoms (victims from the Saku area, 100 children, 100 control cases, 51 adults, 50 control cases)

Tendon reflexes, disorders in the sensory nerve system, impe-diments of coordinated movements, disorders in the muscles, are all observed.

11!1.1: ,ff111':-/, /1' Yt1 100 f.%91, :M. 100 4'1, 1:%* À 511e, )t.t i50) e Fri! 5Ij,1

(%) 70

f

accentuated Tendon reflexes diminished

disease-like reflexes 4i1 er D !

Sensory systems

reduced sense of position reduced sense of vibrations numbness

victims 30

' 777.77:77.' 7,11

controls (%) 3 0

' '

'

1 17.7Z rstanding on one foot impossibljf" . -

coordinated movements Ldysdiadochokineasia

unable to walk

co

IUM77=11==

child victims adu:t r mq. viemms.

0 ' 10 20

I.

50 62

Forgetfulness, reduced, attention' span te Fatigue, languor

Reduced vision, xx

Headaches

Muscle aches

Excessive perspiration ,

1 1 victims (56 cases)

u,ri;) Li control (22 cases)

39

Figure 11: Autonomic Neurzpathic Symptoms of Saku-Area Victims

Z110 : lÂafo pecufir: lx

excessive fluid intake-- e t

car sickness e?

excessive perspiration-- - constipation

headaches diarrhea low blood pressure

dizziness nausea shakes

goose flesh

Figure 12: Clinical Symptoms of Chronic Organophosphate-Intoxicated Victims (from Metcall2

(Metcalf K.: ).

**Translator's note: unreadable print.

& 1 i jjIJ.5

D , E G h-• < Tr= re't t or, ;');

; .4P '

40

•

Figure 13: Two Examples of the ERG of Chronic Organophosphate- Intoxicated Victims --

top cases visual acuity 0.01 • P.:Dil--à", 1

ERG middle cases visual acuity 0.5 Case 1 Case 2

(during treatment) ...,--\ /....---„,,„ ir--....._ . lower cases visual acuity 1.5 —f _..../46.11.8. \,, f -------- 461224.

(after treatment) ---... / ---\,1

(initial examinations)

It can be seen that with treatment an abnormal ERG returns to complete normalcy.

—'-47.1.9.

. . -----`1 ■ --., .,.-------.,

472 19. ---- ' .----"---,' 4 7 217.

'•rr

V J \ .

T ___11 2')°

iTiOnmet

471.25

Figure 14 (A): Ocular movements 4 M (A) : h .•

before treatment, upper-viewing spasms

. . •

.., , , • . . .,

'...." . . '

... ,

-•. f..,'. u_----s-:..._ . _ , ;':,- ':•' .....<r-7:,:-;..- . r-7,. . , -...-,;

... -- - ' _ -- et'-'-.-77..‘ - • •.-e•Fs---: Ï-.-_.:.- --,'.._ • _. ,-. --, -,_ , ''...:' - .,- r• 1. -...-,-, '...- :-.."'-

,..,

"-:, 4e"-- '1-, ...:•: •., .„-- ''

t • ';.• ,......- ' --- l',.. -,, .- Z . , ,-;.-1...- • --,,,..,',_. • _....„.; i ,_ •

,,......'":".•• t - - - "'"•••''.. :: . - :4 .- ; ''' -- •••• • .-..e.

' ' -'•-•--' 4 ..---- i. • • > 't ',- - ...:,..) .-- - • -•

— • :,- ",-....;_.,— : • . <— •:.- '• , --'

-, ,..1 . .....,.,. ...-. . --

1

f;* 1 ,

Figure 14 (B): Complete recovery after i-e, 47 : u m,

treatment, observe the facial expression of complete recovery.

1 -

'

•

f

; R

L

47.5.30

R ,7") L -

100

1.000 tux

41

M- 1519 : tf;I:11,t*ÈY-IM _ _ UfjeI2-fè,1,- -co ts <

E 0 G 0.5 Hz

R

r\-\2 472.15.

/-\ ... • r---.. /,-,,,, 2,- R \_,/,r— \ J.: --`2\ •-•-\,'\ .7 ---)`'..__,j/e-

'‘, ," \„..__., \ r L ; ,, ." , .. .__,/ _.._ . • ,_.k• 48.4.18.

Figure 15: Smooth-Pursuit Movements

It can be seen that with treatments the wave shapes gradually normalize.

Figure 16 (A): Pupils (infrared photographs) DK---dark

Pin-hole pupils indicating a typical acute intoxication by malathion.

re Ism (A) : IL (*ci-me - () D K. — en pin hole

pupil

0K

100 lux

Figure 16

A case of Mydriasis

(B): Pupils eelGuA (B)

chronic intoxication with malathion.' iS observed in the DK.

D K

le 17 a (A) ; L. - TE

M17[11 (B) zr- 11" W. eit . . . _

e;-•

• t.-)K • —

M P e M51-‘:1-gl PS. ?L'57-;1-

g e

normal

42

Figure 17 (A): Pupils of a Beagle Dog

The pupils of a normal dog are shown.

Fiere:17 (B): Pupils of a Beagle Dog

An example of a chronically- intoxicated dog with ethylthiometon.

Figure 18: Pupilogram Diagrams

Shown are the pupil reactions for cases .which are chronically intoxicated, normal, and acutely intoxicated.

1Lell light stimulation

chronic intoxication

acute intoxication rt.=

• - - • n

1 97-, `,B) -a. t't ,

. ."..

] ma (A) : 11::e -;-1c..‘ 1 1- —

-

• .:•

•

Vf,e7 „

e't t

Figure 19 (A): Platide Photograph of a Normal Person

Figure 19 (B): Photograph of a victim;

notice that the horizontal and vertical lines are lent and do not cross at right angles.

m2ora (A): (D) In& (pr.f1) (93:17.)

FT" s

Th

'

1,1,11 2Cs,

to

o

7-21

p P 1t I r„-il 1 1 [11 ,,• 1,

s. 40 41 i2 4 3 45 45 .17 48 49 SO COItNEAL C-, ?...AT 1, ...7,R:ZONTAL MEMO.)

M. 2021 (B)

23

Il ,.•■•

't[L, :1 ,

jTi

10

L1114],_

›

ç Jh

44

Figure 20 (A): Corneal Curvature Radii (D) and the number of eyes (vertical axis) are shown.

The horizontal meridional lines are shown: top graph -- normal cases, middle graph -- victims (females), bottom graph -- victims (males).

Figure 20 (B): As above, but for vertical meridional lines.

I 4.'"-1

CCie:EAL 1- 1-FrE ( V t.P rr,41 I.0

X)1

45

Table 6. Diagnostic Criteria for Chronic Organophosphate Intoxication

Ministry of Health and Welfare

• Mandatory Items

Eye Symptoms (both eyes)* 1. Impaired vision (sometimes fluctuates) 2. field-of-vision stenosis or central scotoma 3. Abnormal refractions, many cases of myopia (sometimes normal in adults).

Systemic Symptoms 1. Weak symptoms of pyramidal tract (accentuation of reflexes in the

extremities, diminished in serious cases, occurrence of disease-like reflexes).

2. Impairment of specific sensory functions of the lower limbs, unable to stand on one foot with eyes shut (lateral swaying occurs a lot).

3. Presence of autonomic nerve symptoms**.

Examination and Treatment 1. Organophosphates are detected in the urine or in the blood (greater than

• 0.01 ppm). 2. Clinical symptoms are improved by atropine, PAM, glutathione, etc.

* In some cases eye symptoms are not seen in the early stages. ** diziness, headaches, nausea, lightheadedness, diarrhea, constipation,

numbness, perspiration, abnormal senses, thirst, impotence, bladder and rectal disorders, etc.

• Reference Items

Observations on the Eyes 1. In the eyeground, particularly in the optic nerve, there are observed

congestion (early stages), temporal discolorations (middle stage), and atrophy (late stages). Sometimes it takes the form of acute or chronic retrobulbar optic neuritis , or cortical blindness. Retinal blood vessels are expanded (early stages) and constricted (late stages).

2. Strong myopic astigmatism (in children) occur in many cases. 3. Abnormal smooth-pursuit movements. 4. ERG: supernormal (early stages)

subnormal (late stages) --- accompanied by prolonged latent periods for each wave.

5. Reduced central Flicker values.

Systemic Observations 1. Reduced blood-corpuscle- or serum-cholinesterase values. 2. Liver functions: LDH , CPK , CCLF (+), alkaline phosphatase value

blood: hemolysis occurs readily when blood is removed. 3. Glove and stocking type sensory disorder. 4. Bidstrup signs, foot drop**, difficulty in walking, musclé atrophy of

the whole body centered arount the extremities. 5. Abnormal brain waves, accompanied by neurosis. 6. Readily susceptible to allergic reactions, prone to viral diseases. 7. In the process other diseases can be eliminated. * When the hands are brought together, palm-to-palm, the fingers cannot

be bent backwards.

(continued)

46

Table 6 (continued)

** When walking with toes dropped down, only the big toe is bent backwards while the other four toes are still bent downwards. Sometimes it is the other way around.

For details, see Ganka Rinsho I Ho (Journal of Clinical Ophthalmology), 66:817, 1972.

6. 11U --PriaMPPecivigim.t

o

1. tti îàn) 2 . LI: 4) 'C., rql 3. 3!J. r7i :D; t • ( A. À fe: "e)

L (mmvAleplce,

2

3. tz.et.,$)

lk it**

(0.01 PFn

2. VThiifj 7 I- 1, , P A .?"

t 7.)

< r_D 7 , r-Y

•;- , 5 5 - ) fIi

**

F.:Z/11in 1. ;1«f*:, e E ( ) , 1. •

( (5t:m)

2. o 13.1 71 P. Z .h; t • ( .1 • ) 3. 4. ERG : supernounal ( ;11;);)

- subnormal ().JUI)

5 r14 7lj

1. 2. III- Li) 11i, CPKL CCLF (+),

7 2j V 7 7 , ifn 'dZ L. -•\' .;".

3. 4. Eidstrup -f -.v. , foot drop**,

5. k;',4 Un .Te;- , 5 i.,•; 6. 7it- 2r-* -- < ,

e 7. .7) ;5 -c: .

utr X 5 751 t.c •

*".` -c < . O (JQ)i: iL T 5. T f.+*.

: 81 7, FM47-1F-

25 20

Figure 21: Relationships between the number of months administered and Refractive values (A), Corneal curvature radii (B), and Acetylcholinesterase values (C) %, for beagle dogs. A,B show standard errors, C shows standard deviations.

m2im: e (A),freettp4 (33) (c) ABunwlm

47

-I. 11 • Refraction

ti 1

I 0 : _________

- 1

- 2

o Control • p

-3

10 15 20 11,,Jeleween

Cornea

I. 1; - 1 z 0 4: .1 1.. L -; •

."-----:-.------- I I rli i' --- -1

. i 11

, di ---,.....s: =-_—____ - •

.... :

II- •' i ; X. control

I 7., 37; o horizontal . . • vertical E

u • — right eye i 36; --- let 1 eye

0' . .

0 5 10 15 20 25 Tin,. in Month

AcelyIcholinesteras

% Cont rol o—o H E .

7... 150'

tt

= !

,

- 1 1

5 10 15 Tune in Month

o

years

index of refraction7.F in diopters

e WI

1-0 mg

1.5

amount administered

48

Figure 22: A (horizontal) and B (vertical) show the relationships between the amount administered and the occurrence of myopia according to dose response and dose duration.

ig220 A B (tri) ,

e.) dose response, dose duration L-

H 5i

A

- 4

Jii 4h - 3

index of refractionie in diopters

-2 4

1

C

----- Amount administered

1.0 m g

1.5

1.0

0.5

0.5

normal control

I. t7

+ 1

0.5 1 e

2

l.0

-2 0.5

0.5 T

o 1 J

+1 0.5 1 2

years

normal control

years

Progression of corneal --- astigmatism

)

A

Figure 23: Diagrams shaming the occurrence of m o.ia and astigmatism versus the number of years of administration.

23J : )5- 4r- fe. . it; P. 5'1'. 111

-,'..: ;)

Progression of myopia mg/Kg

49

2 0.5 1

(years)

Figure 24

CA)

50

CCI

51

52

Figure 25

„.

'igue 26.

',.1"ii).:/f/ - .). 4. • . ̀)e,,

rig e LI :e.

53 2.7 Figura

rt(10

'7'146'efe.1.-2Q10

y ',341>;\ Ç:ye •

-

,

\-y.;%!:erz?,0Ï-. rver .);):

(B)

Figure 28

54

55

1.,‘i

'

.„

. ■ •

er-e ,

'

; ?ie "1 0

(B)

; .e.

,

, ,f'ew

Figure 29

Figure 30

56

(Â) (B)

40 k

Figure 31

57

58

Captions for Photographs (Figures 2)-l'-31)

Figure 24: Ciliary muscles are shown: control case (A), 0.5 mg/kg administration (B), 1.5 mg/kg administration (C). As the amount administered is increased, the changes in the ciliary muscles become greater.

Figure 25 (A): Shown are retinal pigment epithelial cells (X 16200 of original figure) of a control dog.

(B): For a dog administered 1.5 mg/kg. Pigment epithelial cells containing canaliculi in a degenerated cell, Myelin figure are mitochondria,or smooth surfaced endoplasmic reticula which are degenerated, and large and abnormal granules are seen. Br. Bruch membrane,: cap capillary blood vessels, 0 S external nodes.

Figure 25

Figure 26 (A): Non-medullated nerve fibers in the papillae, Astrocytes contain a small number of dense bodies.

Figure 26 ( B): For a dog administered 1.5 mg/kg, degeneration is occurring, non-medullated nerve fibers are present.

Figure 27 (A): Toluidine-blue-dyed optical-microscope picture X580; • retrobulbar 5 mm is shown for a normal control case.

Figure 27 optic nerve,

Figure 27 (B): The same section of the optic nerve for a dog administered 1.0 mg/kg; compared to (A), there are observed very clear atrophy and degenerated images. These changes were observed in ail the administered dogs.

Figure 28 (A): Ganglionic juncture of the sokukin* of a normal external-eye muscle.

The same location in a dog administered 1.0 mg/kg; observe the vacuolization of the synaptic folding, and the expansion.

Figure 29 (A): Optical microscope view (toluidine blue, X 580) of the sciatic nerve of a normal dog.

Figure 29 (B): For the dog administered 0.5 mg; it is completely different from Figure 29A in that there is much destruction and considerable thicken-ing of the medullary sheath.

Figure 30 (A): The calf nerve (of an administered dog) is shown. The shrinking of fibers, the increase in tissues, and the thickening of the medullary sheath are very clear.

Figure 30 (B): The calf nerve obtained from a chronic organophosphate-intoxicated human victim. The main agricultural chemicals are malathion, sumithion. Note that the changes are the same as observed in animais.

Figure 31 (A): Shown here, and also in Figure 32, are the analysis with the X-ray microanalyzer. The upper part of the figure shows the sections tested, and the lower part shows the analyzed results. The arrow A is the location of the spot analysis, and ail of these are for the liver of a control dogi P --- phosphorus peak is not found.

Figure 31 (B): Analyses are carried out for the entire scan, and for point 1 and point 2. Note that in the observations of the liver, the phosphorus peak P-Kek is large for the phosphorus-administered dog.

* Translator's note: Unidentified term, lî-terally "bunch muscle". Could refer to fasciculus.

Figure 27

Figure 28

Figure 28 (13))

Figure 29

Figure 29

«•-•'7•••• .•:

-

-

_ •

• - • -

'7 .11 - - T. ,

,

,

59

Figure 32 (A): Ciliary muscle of a control dog; There is no P peak.

Cu-Ld is the mesh peak.

VaCC 40itV • 11- rub

ï„, 1-;.1 ••••• • • '••;

‘-‘ . • •

' ' ■ -•".-;\•••• •• "•<,' .

: • • •,'•i • `.••• .

• 2•:• • s. • • ,. •

• • : .

. ,

''••• . • - : , •

- ." • " • s.• s • - • • ••••'

Cu-Lcr,

• J.

. -

,

•

•

,

•

_ . •

• • g . •

' .

-

- •

; -- • .

.' • ,

1 ---- C L.- K cr.

- - • :

• «7 7

. • , • • .

C u cf: I< c-

•

60

Figure 32 (B): Also of ciliary muscles, for a dog administered 1,0mg/kg.

In the bottom left figure the P and S peaks are from

ethylthiometon, and are larger than in A.

V'e,:iCe 430 kV

tab 4 x 60 -31 A

61

Table 7. The Pathological Results for Beagle Dogs (a summary)

1. Found are: pronounced changes in the 3rd zone of the central optic nerve, concent'ration of nuclei (Pyknose) is clear, and shrinkage of cells.

2. Vacuolization and partial darkening of the rim of the external knee-like bodies.

3. Degeneration of optic nerve, increase of glia, decrease of nerve fibers.

Spinal cord 0°11 0 Buidach funiculus glia increase, increase in Astrocytes.

5. Peripheral nerves motor nerves degeneration of motor end plate sensory nerves thickening of marrow sheath,demyelination

6. Liver: Increase of lipofuscin in the cells, spotty necrosis.

7. Pancreas: Irregular Island of Langerhans cells, and increases in their numbers.

8. Spleen: Siderosis, light atrophy.

9. Digestive organs: Degeneration of nerve endings, thickening of unstriated muscles.

10. Thymus: Decrease of Hassall's bodies.

11. Autonomic ganglion: Decrease in cholinesterase chromosome character.

12. Muscles: Neuromyopathy.

62

3311:' 4 -7- e ) E R G 6-.) (A , (Control) Lt) E R G

I ./;"! 0 . 3, 0.0U3sec

U., super ne; mal

subnormal

Figure 33: The changes in ERG (A,B) when baytex was administered (rats), and the ERG of a control. The time constants are 0.3, 0.003 sec. It is shown that for the adminis-tration of a small amount it is super normal and the latent period is shortened, and the administration of a large amount it is subnormal and the latent period is prolonged.

20-1,

: E R G JLL -(1 gi.2.) L 0). (lionnal)

m)pur (1›,U).subnorinal();

Figure 34; The ERG's are superimposed. Depending on the amounts administered, relative to the normal, supernormal for small amounts and subnormal for large amounts are observed.

Table 8 , The Results of ERG Measurements (4 days after administration)

20-joule light stimulation

latent period

time to a-wave apex

a-wave intervals

time to b-wave apex

control group 8 R ;!I 1M 1. (:".!:•5--;!': 4 H H) 20

• ; :0.0051,.z/J g 0.05ing/kg 0. 5 r.:..'> /kg I 25 r.u;/1:g

t7r 0 + 0 + O20±0 e.r*. ; I 9() _ ;.„ .1 ' Ç . ±o

2 90 ; ± 0 ltr'-• • + 0.1 1 _ _ 0

; a ::-.;17.1.«I'41.:1 5.65 -!- 6.15 . 5.05±- 0.25 ju:,,-;».00±0 _412. 0 3.14:c. 0 •-•-! _ .0. z,c.!••••••• !!!I , ;11.5,0 11.19 12.28 : 12.12 1 1 2 I 13.48 , 9 1.75 ;

- - - - - - - - ' 1 ±1.56 ±- 0.49 ± 31! +0.14 +1 7L +0 +1 2D

f•-••,•_.-7-; P i 3 1 ' 36.40 ,1 fb 47 32 OC1 31 4!; AI ±2.80! ±f . 1 + 1 .11 01 1

rythmical small waves

a-wave amplitude

b-wave amplitude

1 0, 10 50 ! 15 - 5 ' ' . !9 . 44 ±0 . 969 . 16 ± 0 . 2. 7 8.67±0.509.13.+0.509.25±0.8'1.5G+0.50 - --- ''' -;' i II, ' 5(1 ---- , ! , »,, , . - 0_ ! 13.16 ' 12. i',8 : 12.52 12.44 13.09

_ 15. '00 15."/ 5 ; 22.25 :

. . . _ . ;

- ! ±4.16 ± 1.-17 ± 0.14 +.- 6.22 :I- 1. 92 :-F, 1 . I'',5 + 0.75':":' + G. 89 ':""! _. - .- -

C:1, 1 20.77 ! 17.97 1 19.26 1 ). 7':. ■ lii. 25 '_.• 0 •. 5!.1 2" . W.) 3'2_ Or ; ; I ›.- 1 - ';-9 ,54 +0.25_ ._+ 0. 9 .5f, _ ±1 ' + 1 ._'i:-;:: . _± 1.0.1 _ -1: 1 . G5 + i ....)•-•'!!':', ; __o_ 31 . G3 • --2-5-. 22 , '2-.6.91 _ 28. 02 :_;;.(v) / 3! I 0 . 5, ? ; i ',._ '

±3.2 ±1.07. ±0.2,-- zi:- ..4.r,i,., -. f_ ; :; , i;r."!: -i- , . t•-,5 -_,...(i . ,-4. .„),- } - _ _ _ . . ._ _ .. . . , " • 5" 9 "")() Z.,;.)3. 0.) 'D 1 I . 0 ■.) I 0.^:3 . P'.' 9.',9 . 4 '•' I' :.' ) ;(..1 i . r ; ' .-/ 1:3.I.,D ' ; a :7.e. ;-'-' ;-- ..,--; -Q; - -, • -- pr, • , - •11 nn ...! ),•:, r

- • ,.., .; 5 -7 fO• ..., ..■ ,.....,, 7 '..... !..... ! L', -;-` •-:-. ;?,r,.(!.- ,•• . -.:- •!,-,. -,0 -: ,!, ± i ;;,.;-,•.., __,... -,,-,-i . -, - , 1 ., ,.„. 1129.00 1521.59 ! 1,110.0.1 • 3351.C..-. 2 217. 0!) Y4 ■ 7,0 17 0 1(.1'0 1115.00 ! ' '.••:. '-` + i78.5:', ± 2!': 00 - 1: 25.50 :I.: 501 ''.'''' --'r ''' --/- 3 /. "i -!c•' .1. !..;.'' -A 2: 0.3!, 0.0!!' ! '!"

'.-•-•:•- 1 ,=•0';', •••:- 5 .-) (::,-. :..I; .4: --:.' k.'_- 7.--5. :: -'.: - .;::::

** e*, *5g -- risk ratios with respect to significance to the control

standard deviation.

subacute reduced vision, peripheral neuropathy.

chronic reduced vision

64

Table 9. Reports of Eye symptoms and Optic-nerve symptoms due to Organophosphate Intoxication

Werden 193289 chronic irregular pupils, paralysis (TOCP) of accommodation, papillar

congestion, temporal dis-coloration, optic-nerve atrophy.

9 ;2 •:' 7_, Ft L.,

\Vercic.il 193';`"

"1 - C, P) 5 Ir. ‘: • ft-'

Grob 19 P.- ■"' .-

( -J- ?

Bi(!slrol) i'.!.» 1:.UF,

( -4 • r•t ) ?t,/_ 1.7

( , t

J.1 T, )

KnaBP (1) F P )

Goldman )

Petty

E N

Battistini ;;LL;; ,,i1. -LIP:I:,

(roc: p ) Albert ini -•;;; •

P) I:ïï fi

( T 0 P )

Heal )'

SteOurnscm

Grob 194990 acute (parathion)

Bidstrup 195391 (mipafox)

Summerford 195392 (parathion)

Parker 195593 acute (malathion)

Knapp 195694 subacute (DFP)

Upholt 195695 acute

GOldman 195896 acute (malathion)

Petty 195897 chronic (PEN,etc.)

reduced vision, sensory disorders.

astigmatism directus

reduced vision.

weakened muscle power, sensory disorders.

sensory disorders

• ■ . ■

reduced vision, congested papillae.

Battistini 195898 subacute (1) F P) (MCP)

G'crlioo )

reduced vision, tem-poral discoloration, central scotoma.

Albertini 195999 subacute (TOAP)

peripheral neuropathy

Gross 1959100 (TOCP)

subacute ascending parlysis, sensory disorders.

Healy 1959101 chronic (malathion)

ascending paralysis

Stephenson 1959102 subacute (DFP)

Gershon 1961103 chronic (parathion)

mydriasis

impaired memory, depression, schizophrenie reaction,unstableness

Hill 1962105 subacute (P. I)

Goldin 1964106 acute (malathion)

Holmes 1964107 (parathion)

chronic

occurrence of myopia

weakened muscles, sensory disorders.

unstableness, neuroses, no strength, sleepiness.

Saku Eye Disease

reduced vision, congestion of nose-side optic nerve, peripheral neuropathy, autonomic nerve symptoms.

Table 9 (continued) 65

Petry 1961104 chronic (parathion)

peripheral neuropathy, loss of strength, balance disorder.

PcirY 1 ..?51 -- „'••- . Ji)J,

. ) Goldin lit31'"" qq! Davingnon 1965108

(various organo- Holmes 1J I' ) ” Yzqi

51'; phosphates) •f•

Day ngnon Ls:: 41 ( ) rintmï

// Brm“) 15(■15''" a ).] n'T Brown 19oo109 chronic

• (various organo-Taberbbaw' 1 G';: 1:, phosphates)

“110 me,au Tabershaw l,uu

Y1, YU'S (parathion) pa \is Y. ,I)"" h. (-' )

Le num.Jc 1969 • J'Yi'UU4

82 ",''.;U Metcalf 1969 chronic

('d) (various organo- mrihn T,

i): • phosphates)

li

chronic

,1 • ) t,'I PPIAf.. 1.1 :. 1 ..1C MI

no power, disappearance of reflexes, balance disorder, sensory dis-order, autonomic nerve disorder.

optic nerve congestion, reduced vision.

abdominal symptoms, reduced vision, neur-otic symptoms, auto-nomic nerve symptoms.

reduced vision, auto-nomic nerve symptoms, ocular-movement disorder, abnormal brainwaves.

subacute

Davis 1969111 acute (parat

hion) dilation of pupils, miosis upon treatment.

Le Hunsec 1969112 subacute (parathion and others)

Ishikawa 1970 21 chronic (malathion and others)

Namba 1971113 chronic (various organo-

phosphates)

centered at retina, clogged artery.

In this country after 1971:

OtoLko, Ichil37 , Uono32 , Segawa33 , Muro114 , Shigeari,Kudo115 , Watanabe57 ,

..

Matsushima, Nakamura, Otsuka50 , and others51-56

Davis 1969117 1 J. qi '1'

7 , I I 1 )1 Weiner E;61'''' <

Pieuich 1972' tà1;';'. Le Hunsuc 1069'

Davis i()'' > Son ) 1971 F:ift' i!iS 1972"' renichel (J,`:') !;';):1 )972

1973"' iiE

Musphy 1969119

Stadt 1965120

Weiner 1961121

Fahmy 1969122

Pietsch 1972123

Le Hunsec 1969112

• • C,

/1-1v is 196:1" o ',.";; ) f

/4'; )(1 ).) 1,-

11 11

Gabliks 1969118 Cabliks r.)G.J""

:Murphy

66

Table 10. Less-Familiar Symptoms of Chronic Organophosphate Intoxication.

reduced reabsorption power of the kidneys and small ureter.

Disorder in amino acid metabolism,

Reduced resistance towards viral infections.

Weakmns against stress, abnormal functioning of suprarenal gland, allergies.

aphtha, ulcers, stomatitis, dermatitis ,

asthma

abnormal symptoms

cataracts

congestion of artery centered at the retina.

Davis 1969111

Solly 1971

Ono 197283

Fenichel (animals)

Kono 197311687

mydriasis

ptosis

supranuclear ophthalmoplegia

116 1972 myopathy

myositis

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which Frequently Occurs in Certain Regions), Rin Gan (Clinical Ophthalmology),

24: 835, 1970.

22) Satoshi ISHIKAWA, Ken OTO: Saku no Kibyo (The Unusual Saku Disease), Gan-ka

Rinsho I Ho (Japan Journal of Clinical Ophthalmology), 64: 731, 1970.

23) Satoshi ISHIKAWA: Saku Chiho ni Okeru Shiryoku Shogai Iwayuru Saku no Kibyo

(Visual-Power Disorders in the Saku Area, that is, the Unusual Saku Disease),

Igaku no Ayumi (Progress in Medicine), 73: 687, 1970.

24) Satoshi ISHIKAWA: Saku no Kibyo (Ganbyo) (The Unusual Saku Disease (Eye

Disease)), Medical Tribune, July 30, 1970.

25) Satoshi ISHIKAWA, Ken OTO: Shiryoku Shogai o Shucho to suru 'Saku no Ganbyo'

ni Tsuite (Concerning the 'Unusual Saku Disease' in which the Main Symptomsi_are

Eye Disorders), Nippon Iji Shinpo (Japan Medical Journal), 2425: 8, 1970.

26) Satoshi ISHIKAWA: Noyaku Ganbyo Oyobi sono Chiryo (Eye Diseases Caused by

Agricultural Chemicals and its Treatments), Chiryo (Journal of Therapy),

52: 1839, 1970.

27) Satoshi ISHIKAWA: Saku Chiho ni Tahatsu Shita Ganbyo (Eye Diseases which

Frequently Occur in the Saku Area), Igaku no Ayumi (Progress in Medicine),

75: 657, 1970.

72

28) Sota HONMA: mu no Jikkenteki NoyakuGanbyo (Canine Eye Diseases which are

Experimentally Produced by Agricultural Chemicals), Igaku no Ayumi (Progress

in Medicine), 7: 657, 1970.

29) Sachiko ICHII, Reiko MIYAGI: Osaka Kita-Kochi Chiku ni Mirareta Shoni no

Shishinkeien (Optic Neuritis in Children of the North Koch! Area of Osaka),

Igaku no Ayumi (Progress in Medicine), 76: 196, 1971.

30) Ken OTO, Satoshi ISHIKAWA: Saku Ganbyo no Seijin Rel (Adult Cases of the

Saku Eye Disease), Igaku no Ayumi (Progress in Medicine), 76: 84, 1971.

31) Satoshi ISHIKAWA et al.: Noyaku Ganbyo (Eye Diseases from Agricultural

Chemicals), Nippon Ganka Gakkai Zasshi, Rinjizokan go (Journal of Japan

Ophthalmological Society, Extra Edition), 75: 841, 1971.

32) Kimio UONO: Noyaku Gansho (Eye Symptoms Caused by Agricultural Chemicals),

Nippon Iji Shinpo (Japan Medical Journal), 2447: 48, 1971.

33) Masaya SEGAWA: Suchiku ni Tahatsu Shita Shiryoku Shogai Rei no Rinsho Shin-

keigaku teki Kenkyu (Clinical Neurological Research on Visual-Power Disorders

which Frequently Occur in Some Areas), Nippon Iji Shinp.2 (Japan Medical Jour-

nal), 2453: 43, 1971.

34) Sadaya FUJINO: 'Saku no Ganbyo' no Shiya ni Tsuite (Concerning the Field

of Vision of the 'Saku Eye Disease'), Shinryo to Hoken (Diagnosis and Insur-

ance), 13(1): 45, 1971.

35) Satoshi ISHIKAWA: Yukirin no Yakuganlyo (Eye Diseases Caused by Organophosphate

Agricultural Chemicals), Ganka Rinsho I Ho (Japan Journal of Clinical Ophthal-

mology), 65: 387, 1970.

36) Kimiyoshi UONO: Noyaku Ganbyo (Eye Symptoms Caused by Agricultural Chemicals),

Ganka Rinsho I Ho (Japan Journal of Clinical Ophthalmology), 65: 393, 1971.

37) Sachiko ICHII, Reiko MIYAGI: Osaka-fuka Kita Kochi Chiku ni Mirareta Shoni

no Ganbyo ni Tsuite (Eye Diseases of Children in the North Kochi Area of Osaka),

73

Ganka Rinsho I Ho (Japan Journal of Clinical Ophthalmology), 65: 400, 1971.

38) Masaya SEGAWA: Suchiiku ni Tahatsu Shita Shiryoku Shogai Rei ni Kansuru

Rinsho Shinkeigaku Teki Kenkyu (Clinical Neurological Research on Visual-

Power Disorders which Frequently Occur in Some Areas), Ganka Rinsho I Ho

(Japan Journal of Clinical Ophthalmology), 65: 407, 1971.

39) Emiko KONOEi Taeko ARAI: Senhoku Sen-nan Chiku ni Mirareta Shoni no Gansho

(Eye Symptoms Found in Children of Sen-hoku and Sen-nan), Ganka Rinsho I Ho

(Japan Journal of Clinical Ophthalmology), 65: 413, 1971.

40) Ken OTO: Saku Ganbyo no Seijinrei ni Tsuite (Concerning Adult Cases of the

Saku Eye Disease), Ganka Rinsho I Ho (Japan Journal of Clinical Ophthalmology),

65: 418, 1971.

41) Sadaya FUJINO: Iwayuru 'Saku no Ganbyo' no Shiya (Field of Vision of the

so-called Saku Eye Disease), Ganka Rinsho I Ho (Japan Journal of Clinical

Ophthalmology), 65: 422, 1971.

42) Kanjiro MASUDA, Masaya TAKASE: Saku oyobi Moriguchi Chiho ni Mirareta Kibyo-

jida no Gan-atsu ni Tsuite (Concerning the Intraocular Pressure in Children

with the Unusual Disease found in the Saku and Moriguchi Areas), Ganka Rinsho

I Ho (Japan Journal of Clinical Ophthalmology), 65: 425, 1971.

43) Sota HONMA: mu no Jikkenteki Noyaku Ganbyo (Canine Eye Diseases which are

Experimentally Produced by Agricultural Chemicals), Ganka Rinsho I Ho (Japan

Journal of Clinical Ophthalmology), 65: 431, 1971.

44) Ken OTO: Yukirin Noyaku Ganbyo no Seijinrei ni Tsuite (Concerning Adult Cases

of Eye Diseases caused by Organophosphate Agricultural Chemicals), Nippon

Ganka Gakkai Zasshi (Journal of Japan Ophthalmological Society), 75: 1944, 1971.

45) Ken OTO, Satoshi ISHIKAWA: Yukirin Noyaku Ganbyo no Seijinrei ni Tsuite

(Concerning Adult Cases of Eye Diseases caused by Organophosphate Agricultural

Chemicals), Medical Tribune (submitted for publication).

74

46) Kameya IMAIZUMI, Kenzo ATSUMI: Noyaku to Me (Agricultural Chemicals and

the Eyes), Ganka (Ophthalmology), 13(7): 717-724, 1971.

47) Satoshi SHIGEARI, Takao MURO, Yoshiichi YOSHINO, Yukihiko SHIMADA: Noyaku

Chudoku ni Tsuite (Intoxication by Agricultural Chemicals), Naika (Internal

Medicine), 27(5): 870-881, 1971.

48) Nippon Noson Igakkai Noyaku Chudoku Kenkyu-han (Japan Association of Rural

Medicine, Research Group for Agricultural-Chemicals Intoxication): Noyaku

Chudoku Rinshorei Chosa (Clinical Investigation of Intoxication by Agricultural

Chemicals), Nippon Noson Igakkai Zasshi (Journal of Japan Association of Rural

Medicine), 18(4): 164-167, 1970.

49) Ken OTO: Yukirin Noyaku Ganbyo no Shindan to Chiryo (Diagnosis and Treatment

of Eye Disease Caused by Organophosphate Agricultural Chemicals), Ganka (Ophthal-

mology), 14(1): 17-26, 1972.

50) Hitoshi OTSUKA: Futatabi Yukirin Noyaku Chudoku Ganbyo ni Tsuite (Once Again

Concerning Eye Diseases Resulting from Intoxication by Organophosphate Agri-

cultural Chemicals), Ganka (Ophthalmology), 13(7): 715, 1971.

51) Tatsuo ANAMICHI, Keikichi IKUTA, Tetsuya KITASHIMA: Folidol Chudoku Kanjya no

Ganshojyo (Eye Symptoms of Folidol Intoxicated Victims), Ganka Rinsho I Ho

(Japan Journal of Clinical Ophthalmology), 49(7): 539, 1955.

52) Hyoei SUZUKI: Yukirin Noyaku Chudoku ni Yoru to Omowareru Gan Shogai no Ichi

Rei (One Case of an Eye Disorder which Appears to be Due to Intoxication by

Organophosphate Agricultural Chemicals), Ganka Rinsho I Ho (Japan Journal of

Clinical Ophthalmology), 50(5): 421, 1956.

53) Masaichi TATSUMI: Akitakennan ni Okeru Kyodo Bojyo-han no Kenko Chosa

(Medical Examinations in South Akita Prefecture by the Joint Insect Control

Group), Nippon Noson Igakkai Zasshi (Journal of Japan Association of Rural

Medicine), 17(4): 241, 1969.

75

54) Haruo WAKATSUKI: Noyaku Folidol Chudoku ni Yoru Kyusei Ikkasei Shitsumei

(Acute and Transient Loss of Sight Due to Intoxication by the Agricultural

Chemical Folidol), Ganka Rinsho I Ho (Japan Journal of Clinical Ophthalmology),

48(6): 419, 1954.

55) Ichiro HARA, Kazuo HASHIMOTO, Kazuo NOGI, Daikichi OBATA, Mitsuo TAKAHASHI,

Kimiji HASHIMOTO, Toshifumi OSHIMA: Nansei Yukirin Chudoku ga Utagawareru

Shinkeishogai no Ichirei (One Example of a Nervous Disorder which is Suspected

to be due to Chronic Organophosphate Intoxication), Osaka Furitsu Koshueisei

Kenkyusho Kenkyu Hokoku (Reports of the Osaka Municipal Institute of Public

Health), 9: 36, 1971.

56) Toshiichi WAKATSUKI: Noyaku Chudoku (Intoxication by Agricultural Chemicals),

Nippon Noson Igakkai Zasshi (Journal of the Japan Association of Rural Medicine),

17(4): 211-213, 1969.

57) Shinobu WATANABE: Kyusei Noyaku Yukirin Chudoku Sho no Kessei chu Noyaku

Yukirin Kenshutsu to Sono Igi (The Detection of Organophosphate Agricultural

Chemicals in the Serum for Acute Intoxication by Organophosphate Agricultural

Chemicals, and its Meaning), Igaku no Ayumi (Progress in Medicine), 80(11):

696-697, 1972.

58) Masayuki SAITO, Hiroaki KUBO: Tokutei Chiiku ni Tahatsu suru Shiryoku

Shogai-Jido no Chosa Hokoku--Seikagaku Teki Tachiba (Investigational Report

on Children with Visual-Power Disorders which Frequently Occur in Certain

Areas--from the Biochemical Viewpoint), Gan Rin (Clinical Ophthalmology),

66: 828, 1972.

75) Todai M2 Yukirin Kenkyu Gurupu (Tokyo University M2 Organophosphate

. Research Group): Yukirin Noyaku Chudoku o'Otte (Investigations

of Intoxication by Organophosphate Agricultural Chemicals), Nippon Iji Shin

Po (Junia tan) (Japan Medical Journal (junior edition)), 107: 24, 1971.

76

76) Todai M1 Yukirin Kenkyu Gurupu (Tokyo University M1 Organophosphate Research

Group): Yukirin Noyaku Chudoku (Intoxication by Organophosphate Agricultural

Chemicals), 1-44, 1972,

Gogatsu-sai Shiryo Panfureto (May Festival Materials Pamphlet).

78) Hichiro MIYAZAWA, Yoshiaki HORI, Tadao WATANABE: X-ray Microanalyzer no

Igaku, Seikagaku e no Oyo (Applications of the X-ray Microanalyzer to Med-

icine and Biochemistry), Dai 5 kai Nippon Rinsho Denshi Kenbikyo Gakkai (sho)

(the 5thc Meeting of the Japan Electron Microscope Society (abstract)), 1973

(Maebashi).

79) Haruo HIKIDA, Mikio MIYATA, Satoshi ISHIKAWA: Jikkenteki Mansei Yukirin

Chudokuken ni Okeru Korinesteraze no Seikagakuteki Kenkyu (Biochemical Inves-

tigation of Cholinesterase in Dogs with Experimentally Produced Chronic Organo-

phosphate Intoxication), Nichi Gan (Journal of the Japan Ophthalmological

Society) ) 77: 172-183, 1973 ,

80) Mikio MIYATA: Korinesuteraze (Cholinesterase), Dai 11 Kai Kita Nippon Ganka

Gakkai (the llth Meeting of the North Japan Ophthalmological Society (abstract)),

July 1973, Sendai.

81) Hiroki IMAI, Mikio MIYATA, Satoshi ISHIKAWA: Yukirin Noyaku 1 kai Toyo ni

Yoru Ratio no Momaku Shogai (Retinal Disorders in Rats Produced by just one

Administration of Organophosphate Agricultural Chemicals), Igaku no Ayumi

(Progress in Medicine), 86: 28, 1973.

83) Shinji ONO, Hiroko IMAI, Satoshi ISHIKAWA: Kyomiaru Mansei Yukirin Chudoku no

2 Rei (Two Interesting Cases of Chronic Organophosphate Intoxication), Rin Gan

(Clinical Ophthalmology), 27: 731, 1973.

85) Satoshi ISHIKAWA, Yuji HAKAMADA, Kazuo KURASAWA: Atarashi Denshi Dokokei

"Irusukoda" to sono Oyo (A New Electronic Pupilmeter, the "Iriscorder", and

its Applications), Denshi Igaku (Electronic Medicine), 17: 59, 1973.

77

86) Hitoshi OTSUKA, Kei TOKORO: Tokutei Chiiku ni Tahatsu suru Tokuyu no Gan

Shikkan no Shindan Kijin ni Kansuru Kenkyu (Toku ni Gan Kussetsu Yoso ni

Tsuite) (Investigations Concerning the Diagnostic Criteria for Specific Eye

Diseases which Frequently Occur in Certain Areas (Particularly with regard to

the factors of eye refraction)), Gan Rin (Clinical Ophthalmology), 66: 9, 1962.

87) Kazuo KONO, Hiroko Yukirin Choki Toyoken no Soshikigakuteki Kenkyu

(Histological Investigations of Dogs which were Administered with Organo-

phosphates for Long Periods of Time), Nichi Gan (Journal of Japan Ophthal-

mological Society), 77: 164, 1973.

•114) Takao MURO, Yukihiko SHIMADA, Yuzo YANO, Satoshi SHIGEARI: Yukirin • Yukienso

Chudoku ni Yoru Neuropathy (Neuropathy due to Intoxication by Organophosphates

and Organochlorides), Naika (Internal Medicine), 28: 509-519, 1971.

115) Naoyoshi KUDO: Yukirin Mansei Chudoku no Jittai to Rinsho (Actual and Clinical

Conditions of Chronic Organophosphate Intoxication), Dai 11 Kai Kita Nippon

Ganka Gakkai (sho) (the llth Meeting of the North Japan Ophthalmological

Society (abstract)), 1973 - 7, Sendai.

116)Kazuo KONO, Satoshi ISHIKAWA, Ryo TANAKA: Gankin Mahi o Shimeshita Tei ChE

Kessho (Low ChE Blood Symptoms which Show Ophthalmoplegia), Nippon Rinsho

(Japan Clinical Medicine), 31: 137-141, 1973.

125) Satoshi ISHIKAWA, Atsumi YAMAZAKI: Atarashi Gankyo Undo Sokutei Ho (Kosokudo

XY Torakka) (A New Ocular-Movement Measurement Method (the High-Speed XY

Tracker)), Noha to Kindenzu (Electroencephalography and Electromygraphy),

1: 23, 1972.

127) Satoshi ISHIKAWA: Unpublished Results.

130) Masao TANEDA, Toru KOGUCHI, Tomoaki HIDAKA: Yukirin Chudoku no Noha (Brain

Waves of Organophosphate Intoxication), Nippon Noha Kendenzu Gakkai (sho)

(Japan Electroenciphalography and Electromyography Society (abstract)), 1973.

78

131) Satoshi ISHIKAWA: Gan Kogai, Tokuni Yukirin Noyaku Chudoku no Shindan (Eye

Pollution, Particularly Concerning Diagnosis of Organophosphate Agricultural

Chemicals), Ganka (Ophthalmology), 14: 781, 1972.

132) Satoshi ISHIKAWA: Mansei Yukirin Chudoku -- Sono Shojyo to Shindan Kijyun

(Chronic Organophosphate Intoxication -- Its Symptoms and Diagnostic Criteria),

Nippon Iji Shinpo (Japan Medical Journal), 2572: 37, 1973.

Panel Discussion p.414

79

Sho OGATA: 1. In the early reports on Saku area patients, why is it

that the ChE values are not necessarily low?

2. It has been stated that PAN showed some effect on chronic organo-

phosphate intoxication, but Hiraki et al. say that it bas no effect on chronic

cases. What can be said about this point?

With regard to the questions from Professor Ishikawa, it is necessary

to have sufficient controls for the electron-microscope investigations. The

results of chronic intoxication by organophosphates using pure beagle dogs

were reported at the general meeting of the Japan Ophthalmological Society,

but again, this time acute and subacute intoxication experiments on beagle

dogs were carried out.

Concerning the experimental method, 6 beagle dogs were used, with 4 of

them administered with sumithion and the other 2 used as the controls. For

the group of administered dogs, 20 mg/kg of sumithion was administered orally

each day at the beginning, but because cholinesterase decreased rapidly and

impairment of the entire body occurred (one dog died), administration was

stopped; and when the cholinesterase returned to normal, administration

was begun again at a reduced amount of 5 mg/kg of sumithion; during the

following 6 months its effects on the whole body and on the eyes were observed.

The experimental results were: 1) The blood-corpuscle cholinesterase

values were rapidly reduced by the administration of the 20 mg/kg of sumithion

(to 20% of the former values), and after the administration was stopped,

there was recovery to about 50% in about 40 days; the readministration of

5 mg/kg again lowered the value to 20%. However, no outward abnormalities

were seen on the body as a whole.

p.415

80

2) Abnormalities were found in tests on liver functions.

3) In the clinical observations on the eyes, abnormalities were not

found in the intraocular pressure or in the eyeground (color and fluorescent

photographs), and the measurements of the corneal curvature radii or refrac-

tions showed no strong astigmatic or myopic tendencies. 4) So that this

experiment could be continued, no tissue examinations were performed.

Shoichiro AKIYAMA: An important question is whether or not there

are chronic organophosphate-intoxicated victims in the urban areas.

From our series of studies on visual field stenosis we have doubts

about the presence of organophosphate intoxication in the urban areas, and

we ask for the speaker's suggestions on the following analysis.

(1) In a massive examination of elementary school children (3 blocs --

former farming areas, air-polluted areas, city-center areas -- each bloc con-

sisted of 6. schools for a total of 18 schools, and the number of children

tested was 18,571), we found 46 children (0.25%) with visual-field stenosis.

The ratios found were the highest in the city-center areas, then the air-

polluted areas, and the farming areas the lowest.

(2) As a treatment, padrin * had little effect.

(3) For a group of randomly chosen 25 children and 14 control children

the blood-corpuscle cholinesterase activity values were measured, but ail

cases showed normal values. Also, the serum cholinesterase values were all

normal.

(4) What were the ERG findings for chronic organophosphate-intoxicated

patients?

Sota HONMA: Previously reported,at the 24-26th meetings of the Japan

Ophthalmological Society, for dogs administered with organophosphates alone

(ethylthiometon, EPN) or in combination with organochloride agents &-BHC),

81

were changes in the following: clinical symptoms, eyeground observations,

spinal cord, peripheral nerve, rear-limb muscles and external-eye muscles.

Here, we have reported particularly on the histological findings of the

ciliary bodies in the eyes of the dogs.

As a whole, those showing the stronger symptoms of intoxication due to

the administration of agricultural chemicals had greater occurrences of cir-

culatory disorders of the ciliary bodies, and this finding tends to parallel

the changes in cholinesterase (blood corpuscle, serum) and the changes in

the refractive indexes of the eyes. From the above, it is believed that

organophosphate agents cause functional and organic disorders of the eyes.

Naoyoshi KUDO: 1) If the analysis of P in the blood is not possible,

eat else is important in the internal clinical examinations?

2) When organic-chloride agents are given to a patient with P + in the

blood, what kind of effects are brought about?

3) Are the blood-corpuscle ChE values and serum ChE values reliable

for chronic organophosphate intoxication?

The above questions are put to Professor Ishikawa.

Moreover, the speaker recently established the following things:

(1) In the blood the P is (-) for 20% of the inhabitants of farming

areas.

(2) The following things are characteristic for a person with P + in

the blood:

(a) There are large variations in the results of tests on liver function.

(b) It appears that there is no relationship between P + in the blood

and blood-corpuscle ChE values.

(c) Persons with P + in the blood are susceptible to hemolysis (about 10%).

82

(d) High values of LDH and ALP are found in persons with P + in the

blood (statistically significant).

(e) There may be a correlation between Bidstrup sign + and P + in

the blood. This is under investigation.

Michitsugu KAWAMOTO: When high concentrations of the organophosphate

agricultural chemical sumithion were orally administered to rats, very clear

corneal opacities were observed, and this symptom is believed to be the

cause of the acceleration of both the MMP type and the EMP-TCA type of corneal

sugar metabolism. Moreover, compared to the normal control cornea, definite

increases in the wet weights were observed for the corneas with opacities.

This may imply that the swellings have something to do with corneal opacity.

Very definite symptoms of keratitis are observed in the cornea and also in

the middle to inner zones of the retina; vacuolization of the cells and

sosuka** are found and examinations with the electron microscope show vacu-

olization of the nerve cell nodules; however, in general, definite changes

were not observed.

With regard to corneal opacity caused by high-concentration adminis-

trations of sumithion, the mechanism of this opacity is a very difficult

problem, but if there are any proposals, I would like to hear them.

Masao NISHIMURA: I would like to ask, from the epidemiological and

the hygienical points of view, about the relationships between the various

experimental results on organophosphate intoxication and the symptoms which

occur in the Saku area.

Masao NISHIMURA (additional): The main worldwide indicator used for

setting the criteria of organophosphate intoxication and safety is A ChE,hut if

Broféssor- Ishikawa ohtains-hetteT organapheephate sensitivity towards the eyes,

it becomes a major problem (for future research).Therefore,full study and scientific

** Translator's note: Unidentified term, leterally "transform into crude pores".

83

research, including those by people from other fields, are necessary.

2) In humans the concentrations for measurement by gas chromatogra- p.416

phy are too high. With only the retention times there are problems, and

are there not problems with regard to the types of organophosphates? More-

over it is also necessary to further study the aspect of detection in tap

water, and I hope that more epidemiological studies will be carried out.

3) It seems to be premature to reach a conclusion on the problem occur- .

ring in the Sakuarea only from the results of animal experiments.

Tsutomu UMEZAWA (additional): Mr. Oshima stated that a conclusion

cannot immediately be arrived at because there is a statistically significant

correlation between pollutants and subjective symptoms, while I would like

to say that, for the phenomenon which at present is believed to be a disorder

caused by photochemical reactions, reinvestigations must be carried out from

the beginning, including unknown substances.

I would like to hear from Professor Ishikawa with respect to this point.

Kazuo IWATA: Is there a possibility that organophosphate intoxications

will increase in the future, and why?

Harusaburo KUWASHIMA: A recent example of intoxicative degeneration

by drugs is the so-called SMON (subacute myelo-optico-neuropathy). When

the optic nerve is damaged by agricultural-chemical intoxication, as in the

results recently given by Mr. Sota HONMA, and if it is assumed that swelling

degeneration of the retinal ganglionic cells are found, the initial degener-

ation of the optic nerve stem should occur at the synapsis of the Tractus

opticus and the gaisoku hizajotai * ; bas this been examined? It is quite

natural that dalogis* are added to the atrophied image in later stages.

Keiichi NANBU: I hope that the clearly-defined method for the quan-

titative analysis of organophosphates in the blood will be published as soon

* Translator's note: Unidentified term, literally "lateral knee-shaped body".

84

as possible. It is distressing that the results are published only like

news.

Hiroyuki OSHIMA: For clinical-diagnosis examinations, particularly

with respect to refraction, mention was made of refractometry, ophthalmology,

supersonic waves, and phacometry; of these, phacometry does not give a

direct reading but requires complex calculations to arrive at the required

value; in this regard it cannot be handled in a manner similar to the other

tests. I would like to ask if phacometry was used only when information

from other tests was limited and other data were needed.

Hitoshi OTSUKA (additional): We have carried out animal experiments

on organophosphate intoxications with Professor Ishikawa. The changes in

the eye-refraction factors in dogs were investigated because we wanted to

establish our theory on myopia that abnormal tonus of the ciliary muscles

is transferred to the optic axis extension*. Although

this examination showed very definite disorders in the ciliary muscles due

to drugs, the results cannot be interpreted as proof of the theory of myopia

occurrence; so, I must say that it bas only been shown that it is possible.

Satoshi ISHIKAWA (response) to Professor OGATA: (1) Although cholin-

esterase (ChE) values can be used as one of the indicators of organophosphate

intoxication, for those such as acute parathion intoxication (Hiraki et al.),

from its value it is impossible to determine whether it is normal or abnormal

for chronic P intoxication. This is because we do not have the original values

for individuals who are first-time patients in the hospital. However, if

it increases during the therapeutic trial and does not go down afterwards,

its value can be considered to have been low; there are many such cases.

From approximately 1,000 Saku cases, the average serum ChE values were found

to be less than 0.8 only in July, while in the remainder they were all greater

* Translator's note: Literally translated.

85

than 0.8. The blood-corpuscle ChE is also similar. In this regard we would

like to assess these values after examining much greater amounts of data.

For acute intoxication cases the ChE values can be immediately compared.

(2) On what basis do you say that PAN is effective for acute cases

but not effective for chronic cases (particularly towards optic neuropathy)?

Hiraki et al. have not studied the data with respect to the optic nerve,

retina, pupil, peripheral nerves, etc.; and found in the literature from

other countries are cases where PAN was effective and also not effective,

even for P other than the para-nitrophenol group; thus, a verdict on this

should not yet be reached. The problem with therapy is that you can say

something about the results only after treating the patients, and you should

not decide only from the assertions of Hiraki et al. In our cases there are

also ones where PAM was completely ineffective. Please try it for yourself.

(3) Finally, I would like to comment on Professor Ogata's experiments.

First, for the liver functions GOT, GPT and ALP were given, but what are

the reasons for showing transaminase? This enzyme value is changed by hep-

atitis, liver cancer and heart-muscle diseases and is more difficult to

vary when compared to LDH, CPK, SDH and ALP. This is also the case for BUN.

Next, one dog died from the administration of 10 mg/kg of sumithion. Because

of sumithion for mice is high, is it correct to consider that sumi- the LD50

thion is of low toxicity, and is it really of low toxicity? With regard to

alkaline phosphatase, there is a report from abroad that it is high in chronic

cases. In your publication the experiments were carried out by using only H-E

staining; but, in the research on drugs such as P, 'disorders in the enzyme

levels' occur, and particularly for toxic substances, 'functional changes

rather than organic changes' occur in the early stages; thus, can one really

say anything because there were no changes in just the H-E? As given in our

•

86

reports here and yesterday, general histological examinations (LFB, H-E,

toluidine blue), histochemical examinations (ChE, SDH, LDH, trichrome), elec-

tron microscope findings,etc. must be studied and closely compared to the

control. If you say that it is difficult to judge even though the electron

microscope shows changes, I would like to examine those data. In any event,

because your data comes from the university where Professor Ueda resides,

it will have quite a powerful effect on the administrative officials in Japan

with respect to the problems of agricultural chemicals; thus, as a ophthal-

mologist, the data of Professor Ogata must be complete so that researchers

and specialists in the field will be fully convinced, and in the future you

must present complete results and methods which should be similar in scope

to the data studied and published by us. The present data are not enough.

To Professor AKIYAMA: 1. Because chronic organophosphate intoxication

is made up of a group of symptoms, it is difficult to attempt a discussion

of this disease by considering only the one aspect, the field of vision.

Therefore, it is quite uncertain and entirely unclear whether your cases

have definitely contracted this disease; I cannot answer you about your

cases because of the incompleteness.

2. The ChE values of the blood corpuscles and of the serum were looked

at only once on that day per individual, and the measurement method is also

unclear. The essential requirement is the detection of organophosphates

in the blood and urine by gas chromatography. More data are required.

3. The number of victims inthe urban areas is indeed smaller. However,

this disease was even found in the following: among children living near

prefectural pounds for stray dogs, a child who drank water from a well sit-

uated at a home built on a former farming field, a child with a grandmother

who used malathion on her bonsai plants at home. Ail of these cases occurred

p.417

87

in the urban areas.

4. Regarding the ERG, Imai et al. have reported in detail in the July

'Igaku no Ayumi' (Strides of Medicine), so please refer to it. A super-

normal ERG is observed for small amounts, and a subnormal ERG for large

amounts.

To Professor Kudo:l. The cholinesterase of both the blood corpuscles

and the serum are very difficult to assess. This is because we do not know

the normal values for each individual person. Data showing increases or

decreases in alkaline phosphatase values have been reported abroad and also

by us. The LDH appears to be higher. Depending on the case, the CPK also

increases. In the animal experiments there were many cases of unchanged

liver functions, even when histological changes were found in the liver.

The organic chlorides mask the P.

2. I have high regard for your detailed investigation of the victims

of Hirosaki.

To Professor Kawamoto: 1. Although changes in the cornea are very

important, we cannot say very much because no detailed investigations have

been done; however, it appears that the following have some bearing: ten-

sion in the eyelids, intraocular pressure, tonus of the external-eye muscle,

tonus of the internal-eye muscle,changes in the tissue itself of the cornea

etc. After further detailed studies, we should be able to answer more fully.

To Professor Nishimura: 1. The reason for the small amount of research

which has been carried out is that some agricultural chemical researchers

have incorrectly reported that organophosphates are harmless, decompose

easily, and leave no residues; furthermore, they have even said that chronic

intoxications do not occur. Also, researchers have been subjected to pressures.

2. In the clinical field some old data by Hiraki et al. states that

88

PAM is effective only for parathion and KEN. In research done abroad, there

are many reports of cases which were effective for chronic intoxications

other than for these two. In our own cases, there were both effective cases

and ineffective cases.

3. For chronic intoxications, if atropine is used first and then the

use of PAM is begun after a while, this appears to be more effective than

using PAM from the beginning. I suggest that this be tried on the actual

diseases. Then, I would like to consider it after I see your data.

4. The reason for using ekathin* is that this is used quite frequently

on the ground for fruits in the Saku area, and there is a preliminary report

by Professor Honda. Sumithion is not of low toxicity for dogs.

5. In the future I would like to carry out a more thorough investi-

gation on sumithion than the one by OGATA.

To Professor Umezawa: (1) The cause of the irritation df the

eyes by photochemical smog is not yet known. Measures to counteract it

are described in 'A Guide to Present-Day Treatments 1973'.

(2) For the photochemical reaction in other countries descriptions

of eye symptoms are rather few in number, and in my own experience in Los

Angeles there was a considerable change in the upper air passage mucous mem-

brane. In the case of Shakujii,it is surrounded by farming fields, and today

I have shown that in Canada and the United States the air-pollution symptoms are

due to - phosphorus. In this connection 'Me no chikachika'(Irritation of the

Eyes) is at the top of the list. Please consult it.

To Professor Iwata:l. This intoxication will never decrease as long as

the use of P is continued. Because victims are diagnosed as having apparent

myopia, amplyopia, or chronic retrobulbar neuritis, they are not treated,

and so it will become a major problem in the future. Similar to the Niigata

89

Minamata Disease, I would like to see more thorough consideration given to

shifting the thinking on the causes of recent illnesses from germs and viruses

to chemical substances.

2. From Professor Iwata's report I would like to hear more details

about the problem between the phenomenon of Ditchburn-Riggs* and field of

vision.

To Professor Kuwashima: (1) Pronounced changes were found in the

optic nerve, gaisokuhizajotai*, optic central nerve, and weak changes

in the ganglionic cells of the retina and in the optic cells. They are

greater near the central nerve than in the papillary area. Because I observed

this disease in its 2nd year, I don't know what the primary changes are.

(2) With quinoform there are no changes in the eyes. p.418

At present, although it has been carried out for two years with

•100 mg/kg, no changes have been observed.

To the people of the Kureha Chemical Industry_q11.: 1. I would like you to show me

the evidence and experimental data which show that organophosphate agents

are of low toxicity, harmless to the eyes, and decomposed rapidly. I would

also like to see your gas chromatography data on DDVP. Where have you re-

ported these?

2. Please read the gas chromatographic analysis entitled 'Organophos-

phate Analysis', as reported by Saito and Kudo.

3. Do you have evidence that low-toxicity organophosphates are harmless

to the eyes? Should this be commercialized as low-toxicity and harmless

without any data? Is it not more humane to save victims?

4 • At present in the farming areas there are 2.1 victims per 1,000 pop-

ulation who remain untreated, and it is necessary,on humane grounds, to change

* Translator's note: See note on Page 83.