Send reprint requests to: G. J. Grega, Ph.D.,1)epartment of Physiology, Michigan State Univen-�it:�-. East Lansing. Mich.

452

THE JOURNAL OF PHARMACOLOGY AND EXPF,RIMENTAL THERAPEtTICS

Copyright � 1975 by Tue Williams & \Viikins Co.

Vol. 193. No. 2Pr,,,ted in U.S.A.

PRESSURE-DEPENDENT FACTORS IN EDEMA

FORMATION IN CANINE FORELIMBS1

ROBERT L. KLINE.2 DANIEL P. SAK, FRANCIS ,J.

HADI)Y AND GEORGE J. GREGA

Depaitntent of Physiology, Michigan State University,

East Lansing, Michigan

Atcepteti for publication December 26, 1974

ABSTRACT

KLINE, ROBERT L.. 1)ANIEI� P. SAK, FRANCIS J. HADDY AND GEORGE 1. GREGA:

Pressure-depentient factors in edema formation in canine forelimbs. J. Pharmacol.

Exp. Tiler. 193: 452-459, 1975.

‘\c(’tvlcholine (10 jtg/unin) infused intra-arterially for 30 minutes into naturally perfused

fortiimbs increased forelimb weight 23 g, largely due to edema formation. The weight

g tin w�:is associated �vith marketllv elevated skin and skeletal muscle blood flows and

small vein pne�sures, suggesting tilat tile edema was attributable, in part., to a rise in

microvascuular pressures. Mechanically increasing venous pressure and blood flow to

similar levels for 30 minutes in l)ulmp-perfused forelimbs produced a weight gain of

27 g. Tile rate of weight gain for the acetvlcholine and mechanical alterations was nearly

itielltical. Acetvlcllolille and mechanical alterations both increaseti forelimb lymph flow

rate i)ut failed to increase lymph total protein concentration significantly. These studies

illdi(ate that ill the (log forelimb elevated microvascular pressures result in edema forma-

t’oui iW increasing the transcapiliary ilydrostatic l)re�ure gradient without producing an

inli)ortant� (iecrease in tile transcapillary colloid osmotic l)nessure gradient. Increased

l)nessture is not associateti with a large increase in microvascuiar permeability to plasma

proteins as is seen with the administration of high doses of histamine and bradykinin.

lle�tuitlv � have described pressure-depend-

eitt aul(i l)r(’��urt-ii1depeuldent mechanisms in-

vol�eii iii e(iennt formation in canine foreiimbs

l)U lO(allv atlininitered histamine (Grega et at.,

1972; Iladtiv et a!., 1972) and bradykinin

(Kline et (ii., 1973) . Pressure-independent fac-

tors. j.(� ., a direct effect on mierovascular per-

Received for publication October 15, 1974.1 This study was supported by a grant from the

National Heart and Lung Institute.2 Current address : Department of Physiology,

Health Sciences Centre, University of WesternOntario. London, Ontario, Canada N6A 3K7.

me�li)ility to l)laslna proteins, were found to be

important for edema formation during infusions

of iligh doses of these two vasoactive agents.

Forelimb weight. as well as lymph flow and

protein concentration increased dramatically(luring infusion of these agents, even under

constant flow conditions where microvascular

pressures were probably not elevated.It is difficult to assess the role of pressure-

dependent mechanisms in edema formation

when using agents which directly alter micro-

vascular permeability an(! thus influence trans-

vascular fluid ullovement. Forelimb microvascular

l)ressllres were undoubtedly also increased by1)0th agents (Gregs et a!., 1972; Kline et a!.,

1973 ) . Elevated microvascular pressures alone

1975 PRESSURE-DEPENDENT FACTORS IN EDEMA 453

will result in fluid filtration according to the

Starling-Landis hypothesis. What is not entirely

clear is whether or not, and if so, to what

extent elevated microvascular pressures affect

permeability to plasma proteins (Garlick and

Renkin. 1970 : Grotte, 1956 ; Haddy et ci.,

1972) . A decreased transmural coiloid osmotic

pressure gradient could also contribute to the

extent of fluid loss caused by increased micro-

vascular pressures.

This paper is an attempt to look more

closely at transvaseular fluid fluxes resulting

purely from pressure-dependent factors, under

simiiar hemodynamic conditions seen during

infusion of histamine and bradykinin. Hemo-

dynamic alterations in the forelimb were pro-

duced by mechanically elevating blood flow and

vein pressures or by local infusions of acetyl-

choline. Transvascular fluid fluxes were esti-

mated by continually monitoring forelimb

weight. Changes in microvascuiar permeability

to plasma proteins were evaluated by measuring

iympil flow anti protein concentration.

Methods

Mongrel dogs of either sex, having an average

weight of 20 kg (range 17-24 kg) were used in

this study. All animals were anesthetized with

sodium pentobarbital (30 mg/kg) and ventilatedwith room air using a Harvard respiratory pump.

Hemodynamic measurements. The collateral-

free, innervated forelimb preparation was used to

investigate the hemodynamic aspects of prolongedinfusions of acetylcholine. A detailed description ofthe preparation has been published previously

(Grega at al., 1972). Briefly, the forelimb skin andmuscle above the elbow was sectioned by electro-

cautery, and the humerus was cut, leaving a neur-ally intact, vascularly isolated forelimb. Blood en-

tered the limb only through the brachial artery and

exited only through the brachial and cephalic veins.After heparin treatment (10 mg/kg), forelimb

pressures were measured from small arteries andsmall and large veins in both skin and skeletal

muscle. Small artery catheters were inserted in a

downstream direction, whereas small vein catheterswere inserted in an upstream direction. Pressures

were measured with low-volume displacement

Statharn transducers and recorded on a Sanborndirect-writing oscillograph.

The brachial and cephalic veins were cannulated

with a short section of large-bore polyethylene

tubing, and the outflow was directed into a reser-voir, which continuously returned blood to the

animal via the jugular vein. Blood flow through

skin and skeletal muscle was estimated by timed

collections of the cephalic and brachial venous out-flows, respectively. Forelimb weight was continually

monitored by placing the limb on a calibrated

I-beam balance. The addition of a 2-g weight

usually caused a pen deflection of 10 to 20 mm of

paper. Mean arterial pressure was measured from a

catheter in the lower abdominal aorta. Acetyicho-

line chloride was infused for 30 minutes into the

brachial artery, via a side branch, at an infusion

rate of 10 i�g of base per mm. The volume of theinfusate was 02 mi/mm. Pressures and flows were

determined twice during the control period, 2, 5,

10, 15 and 30 minutes after the infusion was mi-tiated and again 15 minutes after the infusion was

terminated. Total and segmental (large artery,

small vessel, large vein) vascular resistances in skin

and skeletal muscle were calculated as describedpreviously (Grega et a!., 1972).

In a second series of animals, the hemodynamic

aspects of prolonged increases in blood flow andvein pressures were investigated by mechanically

increasing these parameters in pump-perfused fore-

limbs. Arterial blood obtained from a femoral

artery was pumped into the brachiai artery with a

Sigmamotor pump. Initially the flow was adjustedso that the brachial artery perfusion pressure wassimilar to systemic pressure. After a control period,

blood flow and vein pressures were elevated simul-

taneously to simulate the conditions seen duringinfusion of acetyicholine, bradykinin (Kline et al.,

1973) or histamine (Grega et al., 1972) . Blood flowwas increased by speeding up the pump, while vein

pressures were adjusted to the desired level by

tightening screw clamps fastened to the venousoutflow catheters. Pressures, flows and limb weightwere monitored as above for 30 minutes underthese conditions, and 15 minutes after the neduc-

tion of flow and vein pressures to the control

levels.

Lymph studies. Samples of forelimb lymph were

obtained using a technique described previously

(Haddy et al., 1972) . Briefly, in essentially intact

foreimbs, the lymph vessels in the area of the

cephalic vein above the elbow were isolated andtwo or three of these were tied centrally. Another

lymph vessel was cannulated distally with a 10-cm

length of PE-lO tubing which had been beveled atthe cannulating end. Lymph was collected for 10-minute periods using miniature 0.5-mi graduated

cylinders. Forelimb small skin vein pres’nire and

aortic pressure were measured as in the hemody-

namic studies at the end of each 19-minute period.

After two control periods, acetyicholine chloride-was infused intra-artenially via a side branch of

the brachial artery for 30 minutes at a rate of 1(�

Infusion Period

�) Miii 10 Mm 15 lilin :5) ?iliri

(out rol , �

� -:;�u��� ,� I

.� �veig1it (g) t) . 0 �

0 �lin 2 �\Iii�

0 . () 7 . 4a

PA (111111 fIg) 1 it) 110 104

P5MA (mm Hg) M4 84 � 49’

‘�SSA (null Hg) � S2 � S2 � 52’

Pt4%IV (mm Hg) � 12.6 � 12.3 � 2$.9’P55v (mm Hg) 15 . I � 14 . 6 � 24 . 4a

P.�iv (mm Hg 7.5 � 7.3 � i6.5�

Pt.sv (nInA Hg) 4.M 5.1 10.3”

F1, (Ini/nAin/10() g) 9. 7 � 9.�.) 25.5”

F( (mi/nlin/iOOg) 12.6 � i3.() 25.0”

Postinfusion15 Mm

10.5’

10555”54”26 . 4”27 . 1”16 . 6’12 . 9”

28. 7’29.6”

14. 1”109

63”

62a27.2”26.6”16.0”13.1”27.7”

28.7”

17.5”101)

66’6i”24.0”25 . (1”

15.5”

12 . 5”

26.2”25.0”

23.2”105

71”

65’20. 1”

22.6”12.6”11.1”

21 . 9”

24 . 0”

16.7”101)

Si

549.1)

13.3

6.6

5.15.0

11.5

454 KLINE ET AL. Vol. 193

S i� < .05 compared to zero time.

�Lg of base per mm. Tile volume of the infusate

%%.#{149}�$ 02 mi/mm. Lymph was collected and pressures

were measured during a final 10-niinute. postinfu-

sit)n period. In PLIII1P perfusion stu(lies, lymph was(t)ilects(I and l)1�sst1��5 were mtasured before. dun-

ing an(I after tue mechanical elevation of blood flowand vein l)re�1r( to levels similar to those produced

in the hemodvnamic study. Vein pressure was dc-

vattti i)\ partially occluding venous outflow by

tightening a wire around the forelimb above the

elbow (excluding tile brachial artery).

Total protein concentration in 1mph was meas-

tired i)v a spectrophotometnic (Beckman DBspectrophotometer) method of Waddeli (1956) . All

data from the ilemodynamic anti lymph studies

%%�(��( analyzed using tile Student’s I test modified

for i)aired replicates. Comparisons between groups

�vtre i)erfonined using tile t test for nonpained data.

Results

I-Ie?nod!,n(1m 1C lIe(lSU1e in e iuts

Forelimb weight (tables 1 antI 2 ; fig. 1).

.\(tvicholine ( lOp.g/min for 30 minutes) in-

fa�d mt na-arterially into naturally perfuseci

forelimbs significant iv increased forelimb weigilt.

Aft(r all initial raputi weigilt gain (0-2 minutes)

forelimb weight contiulued to increase at a

ule:Inlv steaciv rate througilout the infusion

penio(l. Immediately after the infusion was

stopped. forelimb weight decreased rapidly by

all amount similar to that gained (luring the

first 2 minutes.

In pump-perfused forelimbs, the forelimb

weight response to prolonged increases in venous

pressure and blood flow was similar to thatseen in the acetyleholine experiments at ilatural

flow, i e., an initial rapid weight gain followed

by a steady rate of weight gain. Returning vein

pressures anti flow to the control levels resulted

in a rapid loss of weight similar in magnitude to

that gained from minutes 0 to 2. There was

110 significant difference between the rate of

weight gain nor tile total weight gaineti over

30 minutes for tile acetvlcholine and increased

pressure flow groups.

Forelimb pressures (tables 1 and 2) . Small

artery lresstlres in bOtil skin and skeletal

muscle were significantly decreased by acetyl-

choline. As iii a �)reviolls study with bradykinin

(Kline et ci!., 1973), these pressures gradually

returned towarci control during the infusion

period. Systemic arterial lresstine was unchangedtilroughout the infu�ion period. Small and large\.ein ijressures in botil skin and skeletal muscle

were significantly increaseti by acetylcholine.

The vein pressures remained greatly increased

throughout tile infusion period and returned to

TABLE 1

I?fJecl of aee!ylcholint ( JO /.uJ/?flifl) infused intra-arterially into collateral-free, innervated, naturally perfuse4

foreli;nbs

Pt = 7: ii�ean control forelimb weight = 506 ± 25 g. Abbreviations used are: PA, mean aortic pressure;

‘�SMA, nluscie small artery pressure; PSSA, skiui small artery pressure; PSMV, muscle small vein pressure;� skiti small vein i)Iesslule; PLMV, muscle large vein pressure; PLSV, skin large vein pressure; FB, bnachiai

venous OUtfloW (muscle) ; F�, cephalic venous outflow (skin).

TABLE 2

Effect of prolonged increase in pressure and flow in pump-perfused, collateral-free, in ncrvated forelinib.s”

U = 7 ; mean (‘OlitEol forelimb weight = 520 ± 25 g. Abbreviations used are: PHA, hracilial artery perfusioti

jresstiie : all other ai)breviations 8$ listed in table 1.

I Pressure, I Flow

(I Mmii 2 Mm � Mm

( oi,trol- 3

O.()

101

Si)

SI)

10.6

1 1 .1)6.36.1)

I I . (1

I 7.6

-� weight (g)

PHA (nun Hg)PSMA (mm Hg)

‘�5SA (mm Hg)P5MI (fIlm Hg)P551. (mm Hg)PI.MV (mm Hg)P.5v (mm Hg)FR (mi/min/10() g)F� (11ul/uliin/iO() g)

0.0

I 0251

8010.211.6

6.3

6.611.6is. i

7 . 6�

l5S�134b

135b

25.lb25 . 5”

18.3”19.2”22.8”23.1”

10 Mm

I 3.9”157”128”

123’24.9”24.9”

17.8”

18.6”22.2”23.8”

10.2”

159”125”125”

24.1)”25.3”

17.8”

18.9”

22.4”24.1”

u; Mm

I 7 .

160”

I 27”126”

25. 4”24.5”17.9”

15.9622.5”23 . 7b

I’ostcontrol

30 i\lin � 4?; �%Iiz,

27.0” � 20.2”177b � 131”

142” � 103”141” 94

24.96 � 9.524.0” 10.317.96 � 5.518.2” � 5.6

22.2’ � 11.0

23.5” � 15.1

“ \ein iiesstures and 1)100(1 floiv were increased simultaneously by partially constnictitig venous outflowand increa.sitig 1)11�i) speed.

b p < .05 compared to zero time.

5-10 10-15

TIME PERIOD IN MINUTES

1975 PRESSURE-DEPENDENT FACTORS IN EDEMA 455

tile control level following termination of the

infusion.

Mecilanicaliv increasing venous pressures and

flow in l)IIml)-perfuse(i forelimbs produced

rai)i(i :111(1 significant increases in brachial artery

l)erfusioll l)resstlre aiid small artery pressures in

C

E0’

tn

FI;. 1. Rate of forelimb weight gain during edemaformation induced by mechanically increasing pres-sure and flow (1 P, I F), acetylcholine (Ach), his-tamine (Hist) , on bradykinin (Brady). Drugs wereadmirnstered intra-arteniailv at natural flow in thefollowing doses: Ach, 10 �&g/min ; Hist, 60 �zg/min;Brady, 10 �tg/min. Hist data calculated from Gregaet a!. (1972) ; Brady data calculated from Klineet a!. (1973). Numbers in parentheses are the num-her of animals pen group. � P < .05 when comparedto either �‘ P, I F or Ach.

both skin and skeletal muscle. The increased

pressures persisted throughout the experimental

period, and brachial artery perfusion pressure

and small artery pressure in muscle remained

elevated for 15 minutes after return of flow and

vein pressures to control values. Small and

large vein pressures remained elevated for tile

30-minute period of mechanically increased

blood flow and venous outflow resistance, tile

former pressure approximately at tile level seei�

during acetylcholine infusion.

Forelimb blood flow (tables 1 anci 2).

Acetyiciloline significantly increased blood flow

through both skin and skeletal muscle. The flow

remained elevated throughout the infusion

period, and returned to control levels during

the 15 minutes after tile infusion was termi-

nated.

In pump-perfused forelimbs, elevation of

blood flow resulted in increased flow through

both skeletal muscle and skin at levels slightly

below tilose seen during acetylchoiine infusion.

The increment in flow from control was greater

in skeletal muscle.

Forelimb vascular resistances (tables 3

and 4) . Acetylcholine significantly ciecreaseci

skin and skeletal muscle large artery, small

vessel and total resistances, and skin large vein

TABLE 3

Effect of acetijicholine (10 �iq/nlin)iflfi4N(’(l intra-arterially into collateral-free, innervated, naturally perfusedforel��,b.s

a = 7. Al)I)1eViati(,Iis useti are: H, resistance (mm Hg/nil/mill/lOt) g limb) ; RAS, skin large artery resist-

ali(P; I?5v)s, skin small vessel resistance; R�5, skill large vein resistance; RT5, total skiti resistance; RA�f,

miistie large artery resistance; R(g_v) M, muscle small vessel resistance; RVM, muscle large vein resistance

RTM, total nlllscle reSisI�liice RAT, total forelimb large artery resistance; R(s.v)T, total forelimb smallvessel resistance; HVT, total forelimb large vein resistatice; R�, total forelimb resistance.

(‘uiitrol(�mtrol

- :�

2.6

5.3

I.’

12.0

:� .05.7I). 5

12.21.3

:� .8

0.3

5.6

HAS

lt(S_V)S

H1-5HTS

RAM

11(5_I) M

M

M

RAT �

R(S-V)T

RVT

RT

Infusion Period

I) Miii � 2 Mm 5 Miii 10 Mi,� 15 i�1in 30 Mm

2.6 2.1 2.0 1.9 1.9 1.6”5.0 1.1” 0.9” 1.4#{176} 1.4” 2.3#{176}1.0 0.7” 0.6#{176} 0.5#{176} 0.5#{176} 0.5#{176}

11.6 3.9#{176} 3.5#{176} 3.8#{176} 3.9” 4.5#{176}:L() 2.3#{176} 1.6#{176} 1.6#{176} 1.6#{176} 1.6#{176}

8.5 0.8#{176} 1.1#{176} 1.3#{176} 1.6#{176} 2.4#{176}0.5 0.5 0.3 0.4 0.3 0.3

12.0 3.6#{176} 3.1#{176} 3.3� 3.6#{176} 4.3#{176}1.3 1.1 0.9#{176} 0.8#{176} 0.8#{176} 0.8#{176}3.7 0.4#{176} 0.5#{176} 0.6#{176} 0.7#{176} 1.1”

0.3 0.3 0.2 0.2 0.2 0.25.4 1.8#{176} 1.6#{176} 1.7#{176} 1.8#{176} 2.2#{176}

Postinftision45 Mi,i

2.4S .0

0.5

11.12.7

11.6#{176}0.4

14.8#{176}1.24.6#{176}0.36.2”

a 1� < .05 when comI)ared to zero time.

TABLE 4

Ez’Ject of prolonged increase in pressure and flow in pump-perfused, collateral-free, innervated forelimb.l”

H = 7. Abhreviat H)IiS 11.5 listed iii table 3.

I Pressure. I Flow

2 Miii .5 Miii 10 Miii 15 Miii

HAS

H(5-V)S

‘ITS

RAM

11(5-1’) II

‘Iv si

HTM

HAT

H 51)T

HIT

‘IT

Control- :� �Iiii

4.50.3

6 . 1

1 .1)

750.5

9.9

0.7

2.7

0.23.7

:10 Mm

Postcontrol

4iS I�1un(I Mm

1.3

4.40.3

6.0

I .11

7.50.5

9.9

0.7

2.7

U. 2

:3.7

1.45 . 5’,

0.37 . 5’,

1.45.0”

0.4

6.960.7

2.5

0.2

3.4

1.55 . 2”0.37.961.44 . 8’

0.36.6”0.7

2.3”

0.13 . 3”

1.55.1”0.37.0”1.44 . 8b

0.36. 6b

0.7

2.3”0.13 . 3b

1.65�3b

0.37.1”

1.64.7’0.4

6. 6”0.52 . 3’0. 13.3”

1.76.3”

8.3”

1.65 .

0 . :�7.5”

0.8

2.7

0. 13.8

2. 1”5.7”

I) .35.0”

2 . 5”

10.6”(1.4

13.6”1.1”:3.7”

0.2

5.0”

(1 \rej,i Jre.”.stire aiitl 1)100(1 floss- were iiicrea.seti simultaneously by 1)artialiy constricting venous outflow

anti increasing I)t1lllP speed.

“ l� < .05 wheui (on�1)ateti It) ZdIi) tin�e

456 KLINE ET AL. Vol. 193

TABLE 5

Effect oflocally administered acetylcholine (10 �g/nhin)

at natural flow

Abbreviations used are: PA, mean aortic pressure;P851., small skin veiii pressure; FL, lymph flow;

TP,. lymph total 1)rotein ; ACh, acetyicholine.

a = 6.

a Vein pressure and arterial pressure measured at

the end of each 10-minute period.

b p < .05 compared to controls.

1975 PRESSURE-DEPENDENT FACTORS IN EDEMA 457

resistailce. Total and small vessel resistances

were lowest during tile 2- to 5-mm period,

gra(iuailv increased from 5- to 30 minutes, but

still remained well below the control level at the

en(i of the infusion. Fifteen minutes after stop-

J)ing the infusion, muscle small vessel resistance

was significantly increased, leading to increased

total muscle and limb resistances.

Mechanically raisillg venous pressure and

blood flow resulted in differential resistance

responses in skin and skeletal muscle. Skin

small vessel and total resistances were signifi-

cantiy increased during anti after the 30 minutes

of elevated pressure and flow. Small vessel re-

sistance ill muscle was significantiy decreased

during the 30 minutes, leading to a drop in

total muscle resistance. After the return to

control contiitions. muscle small vessel and total

resistance were significantly increased. Large

vessel resistances were only minimally affected

(luring tile 30-minute experimental period. The

differential responses in skin and muscle resulted

in a sligllt decrease in combined forelimb small

vessel resistance aild total resistance during the

30-minute period.

Lymph Studies (tables 5 and 6)

Lymph flow rate, but not total protein con-

cent rat 1011 , was progressively and significantly

increased by the intra-arterial infusion of ace-

I).%� IL TPL

nil?’ flu �ii;n Jig ml! 10 mm c,/Ioo nit

Control 119 14 0.03 2.4

Control 122 14 0.03 2.5ACh, 10 fllili 124 98b

0�#{216}7b 2.7ACh, 20 mm 126 95b 0. 16” 2.8ACh, 30 miii 124 27” 0.296 2.5Po.st-ACh 123 14 0. 12” 2.3

TABLE 6

Effect of prolonged increase in pre.ssure and flow pt

pump-perfused forelirnbs

Abbreviations used are: PRA, bnachiai areteny

perfusion pressure; other abbreviations as in tai)le 5.

n = 8.

PBA#{176} PSSV#{176} F�. TI’L

u/ IlK)“7,’ “U mm Jig nit/JO t�iin

Control 10’2 13 0.02 2.3Control 1(14 13 0.02 2�3

I F, I P, 10 miii 162#{176} 28” 0. 096 2.5

I F, I P, 20 mm l57b 25” 0. 10” 2.7

I F, I P, 30 mm iss” 2S� 0. 14” � 2.7

PostS IF, IP 114 14 0.196 � 2.4

a Vein pressure and arterial pressure at the cud of

each 1(1-minute period.

6 � < � compared to controls.

tylcholine. Tile lymph flow rate remained dc-

skin and skeletal muscle infusion was termi-

nated. Skin small vein pressure was increased

and systemic arterial pressure was unaffected

as in the hemodvnamic studies.

Simiiar results were obtained when venous

pressure and blood flow were mechanically dc-

vated to levels resembling those seen ill tile

hemodynamic study. There was a significant in-

crease in lymph flow rate, with no significant

change in lrotein concentration. Skin small vein

pressure and brachial artery perfusion presstire

were increased as discussed previously.

Discussion

Acetylcholine ( 10 �tg/rnin) infused into the

brachial artery for 30 minutes in foreiinlbs

perfused at natural flow increased forelimbweight 23 g. The majority of the weight gain

occurred from minutes 5 to 30 and can be altri-

buted to an increased extravascular fluid volume,

since all segmental vascular resistances were

constant or rising from minutes 2 to 5 onward.

A constant or increasing vascular resistance

suggests that mean vessel caliber was either

constant or decreasing, hence vascular volume

changes cannot account for tile large increase

ill forelimb weigilt. In addition, Diana and

Shaciur (1973) have shown in a similar prepara-

tion tllat the slow, steady component of weight

458 KLINE ET AL. Vol. 193

gaul is a result of tn:lllseapiliarv fluiti nlovement

all�i hot tiit’ re�ult t)f a slow conlponent of

vascular pooling. Time weight gain was associated

Wltil :111 increase ill blood flow and small vein

pressure in i)Otil skiii an(i skeletal muscle. Tile

marked rise in sillail 1-cia pressures, which repre-

sent a minimum for cal)iliary hydrostatic pres-

su r(’ , suggests t ii:i t t he i Ilcrea�t’(i ext ravascuia r

fluid volume resulting from infusion of high

doses of aeetVlcilOline � attrihutetl to fluid

filtration due to a rise in microvascular l)ressures.

\Ieciia nicailv i licreasilig blood flow a 11(1 venous

pressure for 30 nlinutes in l)ulllp-perfused fore-

iifl1l)S resulted ill a forelimb weight gain of 27 g.

By similar resistance analyses, it �un be sho�vn

tilat tile nlajonitv of this weight gain was due

to transcapillary fluid moveillent, not to changes

in vascular volume. The rate of forelimb weight

gain in these Pn(’l):(rations �va nearly identical

to tilat seen in tile acetvlcholine series and was

aeconll)anied I)y Silllii:Lr increases in small vein

l)resstlrt’s. Again . the increa�ed extravascularfluid Voillllle can he eXl)l�tiIle(i bY fluid filtration

resultillg from increased nlicrovascular pressures.

Available evideitce (ioes not support a direct

increase in capillary i)ermeahiiity by acetyl-

(‘hOhille, SiIlCd tile permeability-surface area

I)rotiuct for 1)laslfla proteins or Dextn:in-1 10 waslint increased during intra-arteriai administra-

tiOll of acetylchohine, 5 pg/mill (Joyner et at.,

1974). In addition. acetylcilohille applied locally

to tile mesenteric vascuiatune after an intra-

\‘(‘llOils injection of colloidal carbon did not re-

stilt in a blackening of tile microvessels (North-

OVer and Northover, 1970) . Thus, the above

ht’modvnaimc tiata suggest that high doses of

:ieetvlchohine alld mechanically increased micro-

vascuiar pressures share a similar pressure-

dependent mechanism of etlenia formation, i.e.,iLli increased transcapillary hydrostatic pressure

gradient. However a decreased transcapillary col-

bid osmotic pressure gradient would also con-

tribute to the fluid movement out of the

vascuiature if increased micnovascuiar pressures

produced �ll increase in microvascular perme-ability to plasma proteins such that the con-

centration of protein in the interstitial fluid

rose.

To examine further tile possibility of a de-

creased transcapillary cohloid osmotic pressure

gradient during tile prolonged increases in

microvascular pressures seen in tile-c experi-

ments, lympil flow rate and total protein con-

centration were measured in the clog forelimb.

It. was assumed that lymph protein concentra-

tion as measured in these experiments reflects

the protein concentration in the interstitial

fluid (Haddy et ci., 1972) . Both acetylchohine

infusion and prolonged mechanical increases in

Vein pressure and flow markedly increased

lymph flow rate but had no significant effect

on lymph total protein concentration.

Joyner et a!. (1974) , using a dog hindlimb

preparation, also observed increased lymph flow

without increased protein concentration during

ultra-arterial infusion of acetylcholine (5 jig!

mm) . Similar findings have been reported for

certain other vasodilators such as methacholine,

l)apavcrine and isoproterenol (Joyner et a!.,1974) and for serotonin (Joyner et ci., 1974;

Merrill et at., 1974) which also probably in-

creases microvascular pressures. Haddy et al.

( 1972) reported an increased forelimb lymph

flow with a significant increase in lymph pro-

tein concentration during intrabrachial infusion

of acetyicholine (5 j.tg/min) and during me-

chanicaiiy induced increases in flow and venous

pressure, but the increase in lymph protein

concentration was very small.

These data suggest that increased micro-

vascular pressures have little if any effect on

interstitial fluid protein concentration, as deter-

mined by an analysis of lymph protein. Since

protein transport increases roughly in propor-

tion to lymph flow, a decreased transcapillary

colloid osmotic pressure gradient does not con-

tribute importantly to the fluid effiux. In addi-

tion, increased pressure is not associated with

a large increase in microvascular permeability

to piasma� proteins as is seen with histamine

and bradykinin.

Combining data from this study with those

presented previously for histamine (Grega et a!.,1972) and bradykinin (Kline et al., 1973)

permits a relative comparison of pressure-

dependent and pressure-independent factors in-

volved in edema formation by these vasoactive

agents. Figure 1 shows that, after initial vas-

cular volume changes (0-2 minutes) , there was

a marked difference in the rate of weight gain

caused by high doses of histamine and brady-

kinin compared to acetyichohine and mechanical

alterations, despite similar hemodynamic con-

ditions. In fact, vein pressures and flow were

1975 PRESSURE-DEPENDENT FACTORS IN EDEMA 459

fl1:lilltaill(’(i at a Iligher level throughout tile

experimental period for the acetylcholine and

mecilanicai cilanges reporteci in this study. It is

:tl)l)arellt tilat, Witil these doses of histamine

a 11(1 i)r:l(ivkillin , t he pressure-dependent mech-

allisfll colltributed less to their edemogenic

actioll tilan tile pressure-independent mech-

01115111, at least during tile first 10 minutes of

the infusion when nlost fluid filtration occurred.

It may be argued that acetVicilOiille, unlike

histanline. (loes ilot increase tile surface area

for exchange (Gabel (‘t at., 1964) , but probably

olllV ciecre:ises tile pre-/postcapillary resistance

ratio re�ulting ill increased ifow and pressure

ill existing microvascular channels. Thus, the

rate of weigilt gain is greater for histamine due

to a l)ossibie redistribution of blood flow and a

i:lrger �urfaee area (Baker and Menninger,

1974 ; Diana et a!., 1972) . However, increased

surface area alolle ill tile presence of elevated

microvascular l)ressures couici not explain the

large illcrease ill lymph protein concentration

SCCII �vitii ilistamine (Hadciv et at., 1972) and

hradvkinin (Kline et al., 1973). Reactive hy-

l)eremia after a I)eriod of prolonged ischemiaincreases blood flow all(i, very likely, micro-

vascular pressures and �voi.ild be expected to

increase surface area for exchange (Friedman,

1971) , bitt a significant� increase in lymph pro-

teill concelltration does not occur (Miller et at.,

1975L

Acknowledgments. The autilors wish to

tilank E. Gersabeck anti ,J. Johnston for their

assistance Ivitil tile experimellts anti M. Allen

for her hitIp ill �)re�)aring tile manuscript.

References

BIKER, C. H. AND MENNINGER, R. P. : Histamine-induced PeriPheral volume and flow changes.Amer. J. Phvsiol. 226: 731-737, 1974.

DIANA, .J. M., LONG, S. C. AND �AO, H. : Effect ofiiist:tiliilll’ on equival(’Ilt jore radius in capillaries

on isolated dog hindhmb. Microvasc. Res. 4:413-437, 1972.

DIANA, J. N. AND SHADrR, C. A. : Effect of arterialand venous pressure on capillary pressure andvascular volume. Amer. J. Physiol. 225: 637-650,1973.

FRIEDMAN, J. J. : ��Rb extraction as an indicator ofca�)illary flow. Circ. Res. 28 and 29: 1-15-1-20,1971.

GABEL, L. P., WINBURG, M. M., RowE, H. AND

GRANDY, R. P. : Tile effect of several pharma-cological agents upon Rb86 uptake by the per-fused dog hindlimb. J. Pharmacol. Exp. Ther.146: 117-122, 1964.

GARLICK, D. G. AND RENKIN, E. M. : Transport oflarge molecules from plasma to interstitial fluidand lymph in dogs. Amer. J. Physiol. 219: 1595-1605, 1970.

GREGA, G. J. KLINE, R. L., DOBBINS, D. E. ANDHADDY, F. J. : Mechanisms of edema formationby histamine administered locally into canineforelimbs. Amer. J. Ph�’siol. 223: 1165-1171.1972.

GROTTE, G. : Passage of dextran molecules across theblood-lymph barrier. Acta. Chir. Scand. suppl.211, 1-84, 1956.

HAnDY, F. .1., Scorr, J. B. AND GREGA, G. J. : Effectsof ilistamine upon iymp hprotein concentration

and flow in the dog forelimb. Amer. J. Physiol.223: 1172-1177, 1972.

JOYNER, W. L., CARTER, R. D., R.’,IzEs, G. S. ANDRENKIN, E. M. : Influence of histamine andsome other substances in blood-lymph transportof plasma protein and dextran in the dog paw.Micnovasc. Res. 7: 19-30, 1974.

KLINE, R. L., Sco’rr, J. B., HAnDY, F. J. AND GREGA,G. J. : Mechanism of edema formation in canineforeimbs by locally administered bradykinin.Amer. J. PhysioL 225 : 1051-1056, 1973.

MERRILL, G., KLINE, R. L., HADDY, F. J. .4ND GREG.’,,G. J. Effects of locally infused serotonin on canineforelimb weight and segmental vascular resist-ances. J. Pharmacol. Exp. Then. 189: 140-148.1974.

MILLER, G. L., KLINE, R. L., Scorn, J. B., HADDY,

F. J. AND GREGA, G. J. : Effects of ischemia on

forelimb weight and lymph protein concentra-tion. Circ. Res., in press, 1975.

NORTHOVER, A. M. AND NORTHOVER, B. J. : The effectof vasoactive substances on nat nlesentenic bloodvessels. J. Pathol. 101 : 99-108, 1970.

WADDELL, IV. J. : A simple ultraviolet .spectropiloto-metric method for tile determination of protein.J. Lab. Clin. Med. 48: 311-314. 1956.