D-8700 Wurzburg, F.R.G

J. Physiol. (1987), 391, pp. 193-207 193With 10 text-figuresPrinted in Great Britain

COLD STIMULATION OF TEETH: A COMPARISON BETWEEN THERESPONSES OF CAT INTRADENTAL Ad AND C FIBRES

AND HUMAN SENSATION

BY E. JYVASJARVI* AND K.-D. KNIFFKIFrom the Physiologisches Irstitut der Universitdt Wiirzburg, Rontgenring 9,

D-8700 Wurzburg, F.R.G.

(Received 23 December 1986)

SUMMARY

1. In nembutal-anaesthetized cats, the responses of intradental Ad and C fibres torapid cooling of the crown of canine teeth were studied.

2. Single-unit recordings were obtained from a total of eighty-six intradental Adand C fibres. The mean conduction velocity of Ad fibres was 13-9 m/s (n = 43; range:3-6-26-0 m/s), that of C fibres was 1-3 m/s (n = 43; range: 05-2-2 m/s).

3. In the intact tooth none of the identified Ad or C fibres showed any ongoingactivity in the absence of intentional stimulation.

4. 84% of the Ad fibres (thirty-six out of forty-three) and 88% of the C fibres(thirty-eight out of forty-three) were excited by cold stimulation of the canine tooththey were innervating.

5. For all cold-sensitive Ad fibres the responses to rapid lowering of the toothtemperature were rather uniform. After an initial high-frequency discharge duringthe most rapid change in temperature, the discharge rate fell as the rate of change oftemperature became smaller, and firing stopped completely when the temperaturehad reached a steady level. No firing occurred as the tooth temperature returned toits initial value.

6. A good linear correlation (r = 089) was found between the initial dynamicdischarge of responding Ad fibres and the maximum rate of change of temperatureachieved in a particular experiment.

7. The response behaviour of C fibres to rapid cooling of the tooth was also ratheruniform but different from that of Ad fibres. For C fibres no initial dynamic responsephase was observed. After a mean latency of 7-3 s the fibres began to dischargeregularly at a low rate at a time when the change of tooth temperature was alreadysmall.

8. The firing rate of the C fibres had a weak linear correlation (r = 06) with thestatic tooth temperature achieved. No discharge was observed as the temperaturereturned to its initial value.

9. For eleven cold-sensitive Ad and C fibres the receptive fields were determinedby mechanical stimulation of the exposed pulp tissue. For Ad fibres the receptive* Present address: Department of Physiology, University of Helsinki, Siltavuorenpenger 20 J,

SF-00170 Helsinki, Finland.

PHY 391

E. JYVASJARVI AND K.-D. KNIFFKI

fields were located at the pulp-dentine border, those for C fibres were located muchdeeper in the pulp and tended to have higher mechanical thresholds.

10. When a human tooth was stimulated by a similar cold stimulus as used in theanimal experiments, the sensation evoked and rated by the subjects using a fifty-point categorical division procedure was described as a sharp, shooting pain beinglocalized strictly to the tooth tested. Sensations other than pain were not perceived.

11. Comparing the pain ratings of human subjects with the response character-istics of feline intradental Ad and C fibres to cold stimulation ofa tooth, it is concludedthat Ad fibres with the properties found in the cat are probably responsible for thecold-evoked dental pain sensation in man.

INTRODUCTION

It is well known that cold applied to the teeth is a potent stimulus in producingdental pain in man (Hensel & Mann, 1956; Naylor, 1964; Trowbridge, Franks,Korostoff & Emling, 1980; Ahlquist, Edwall, Franzen & Haegerstam, 1984). Thequality of the perceived sensation depends on the clinical status of the tooth testedand the magnitude of the stimulus used, but is usually described as a sharp, well-localized pain. In a recent study it has been suggested that in addition to a painsensation, a cold sensation can also be perceived in teeth (Griisser, Kollmann &Mijatovic, 1982).Previous studies in animals have shown that cold stimulation of teeth evokes

responses in dental nerves (Wagers & Smith, 1960; Funakoshi & Zotterman, 1963;Yamada, Suzuta & Higuchi, 1968) but little information is available about theresponse properties of single intradental Ad and C fibres to cold stimulation of teeth(Matthews, 1977; Kollmann & Matthews, 1982; Narhi, Hirvonen & Hakumaki,1982 a). The contribution of cold-evoked activity in intradental Ad and C fibres to theperceived sensation in humans is not known.The purpose of this study was to record the neuronal discharges of single intra-

dental afferent Ad and C fibres to cold stimulation of the tooth they were innervatingin the cat and to compare these responses to the perceived sensation in man usingsimilar stimulating conditions. Some of the results have been presented in pre-liminary reports (Jyviisjiirvi & Kniffki, 1985, 1987).

METHODSAnimal experimentsThe experiments were carried out on twenty-nine adult cats (weight: 2-3-4-7 kg) with intact and

fully developed permanent teeth. The animals were initially anaesthetized with an intraperitonealinjection of sodium pentobarbitone (40 mg/kg). The right femoral artery and vein were cannulatedfor measurement of blood pressure and for injections, respectively. A tracheal cannula was insertedto allow undisturbed spontaneous ventilation during the experiments. A deep level of anaesthesiawas subsequently maintained by intravenous injections of supplementary doses of the sameanaesthetic as required. Arterial blood pressure and body-core temperature were monitoredthroughout the experiments and kept within normal physiological limits.The mandible was fixed to the maxilla with the mouth in an open position using dental acrylic

bite blocks cemented between upper and lower posterior teeth. A skin incision was made along theridge of the left mandible. The lower margin of the mandible was removed between the anteriormental and mandibular foramina to expose the mandibular canal. The inferior alveolar nerve was

194

INTRADENTAL A& AND C FIBRES AND SENSATION 195carefully freed from connective tissue with minimal disturbance of the circulation and cut as farcentrally as possible. The animals were placed on their back in a recording frame, the head wasfixed using a metal bar across the palate. Over the mandible the skin flaps were sutured to a metalring to form a pool which was filled with warm paraffin oil subsequently kept at about 37 'C.

A B

Polyethylene

Isolation~~ Islaioplate ~~~~block

Inferior alveolar nerve

5030 c

1 T(GC) *:Z'lo Ecr~ 1

10 s 5 sFig. 1. Schematic diagram showing the experimental set-up for the animal (A) and human(B) experiments. A, T indicates the measurement of the tooth temperature near the pulpusing a thermistor. A typical record ofthe tooth temperature (T) during cold application tothe tooth crown by applying dichlorodifluoromethane gas through the polyethylene tubeis shown in the lower trace. For electrical stimulation of the tooth, a constant-currentstimulator was used. Recordings of single-fibre activity (e.g. upper trace) were obtainedfrom fine filaments split from the inferior alveolar nerve. B, the test tooth was isolatedfrom the other teeth and the surrounding tissue by a block made of impression material.A typical record of the time course of the pain rating of a human subject to coldstimulation of an incisor tooth is shown below. The open arrows indicate the directed flowof the cold spray to the exposed part of the cat's (A) and human (B) tooth crown; thevertical arrows mark the onset of stimulation.

The left lower canine tooth was cleaned and dried. A 1 mm thick Teflon plate was cementedaround the neck of the tooth to isolate the tooth crown from the surrounding tissues (Fig. 1 A). Apolyethylene tube (diameter: 6 mm) was placed over the tip of the tooth crown and fixed in positionwith dental acrylic so that about 15% of the surface area of the tooth crown was within the tube.To measure the tooth temperature near the pulp, a thermistor probe was sealed into a cavity drilledapically to the plate. A shallow cavity just reaching the dentine was drilled between the tube andthe plate; a piece of cotton-wool soaked with isotonic saline was placed at the bottom of this cavity.For electrical stimulation a platinum wire electrode (diameter: 0-4 mm) was inserted into thecavity until it made firm contact with the piece of cotton-wool. The indifferent electrode wasattached to the lower lip of the ipsilateral side.

Using a platinum wire electrode, functional single-fibre recordings were obtained from finefilaments dissected from the cut end of the inferior alveolar nerve. The neuronal activity wasamplified, filtered, audiomonitored, displayed on an oscilloscope screen and stored on magnetic

7-2

E. JYVASJARVI AND K.- D. KNIFFKI

tape using a FM system. Functional single fibres originating from the tooth pulp were identified byrecording single all-or-none action potentials in response to square-wave monopolar cathodalconstant-current pulses (amplitudes: 0-200 /uA; duration: 1 ms). The fibres were classified as Adfibres if their conduction velocity was between 2-5 and 30 m/s and as C fibres if the conductionvelocity was less than 2 5 m/s. Conduction velocity was calculated from the conduction distanceand the shortest latency to a suprathreshold stimulus. By the stimulation method used, theconduction velocity of the C fibres has a lower value than determined by direct axonal stimulation;yet the classification into Ad and C fibres can be made reliably (E. Jyvasjairvi & K.-D. Kniffki,unpublished observations).

After intradental fibres had been identified, their response behaviour to rapid cooling of the toothwas tested by applying dichlorodifluoromethane gas (Provotest) through the polyethylene tube. Theapplication time of the gas was controlled, so that the tooth temperature near the pulp did not fallbelow 2 'C. Response latency and duration were recorded. The pulp temperature was monitoredusing the thermistor output. Cold stimulation was repeated four to five times. Intervals betweenstimuli were usually 5-15 min. In eleven experiments the pulp was exposed afterwards andmechanically stimulated to determine the receptive fields of the recorded fibres.

Human experimentsSeven volunteers (four female and three male) took part in the experiments. The subjects were

verbally informed of the nature of the experiments and they were able to interrupt the experimentalsession at any stage if they wanted. Only intact, clinically healthy upper incisors were used. Thetest tooth was isolated from the other teeth and the surrounding tissues by a block made of Coltoflaximpression material (Fig. 1 B). A hole was cut into the block to expose from the tooth surface anarea, the size of which was approximately 150 of the whole tooth crown area. Altogether ten teethwere tested. Additionally one non-vital, root filled tooth was included to serve as a control forstimulus spread.To estimate the intensity of the perceived pain sensation to a cold stimulus applied to the test

tooth, a categorical division procedure (Heller, 1985) was used. Recently, this fifty-point magnitudescale (0: no pain; 1-10: very weak pain; 11-20: weak pain; 21-30: moderate pain; 31-40: strongpain: 41-50: very strong pain; 51 and more: intolerable pain) was found to be useful in psycho-physical pain experiments (G6bel & Westphal, 1984).A training session enabled the subjects to become familiar with the experimental surroundings,

with the cold stimuli applied, and with the rating procedure. Then the test tooth was stimulatedusing the same stimulation method as in the animal experiments. The stimulus was removed assoon as the subject indicated a response. Recordings of the ratings were obtained by using a visualanalogue scale, whose output voltage was displayed on an oscilloscope and stored on a magnetictape for subsequent analysis. Each test tooth was stimulated four to five times using intervals ofabout 15 min between the stimulation periods. After each trial subjects were asked about thequality of the sensation.

RESULTS

Animal experimentsGeneral properties

In the present series of experiments responses of a total of eighty-six singleintradental fibres were recorded. Only fibres that showed an all-or-none response tomonopolar electrical stimulation of the tooth crown using a stimulus of 1 ms durationand less than 200 ,tA amplitude, and no response to mechanical stimulation of theintact tooth or adjacent tissue were accepted to be intradental. Of these, forty-threewere classified as Ad fibres and forty-three as C fibres according to their conductionvelocity. In Fig. 2 typical responses of intradental Ad and C fibres to electricalstimulation of the lower canine tooth are shown. As the stimulus strength wasincreased above threshold, both units responded with a constant latency. For someC fibres, however, discrete shortenings in the latency at certain suprathreshold

196

INTRADENTAL Ad AND C FIBRES AND SENSATION

stimulus intensities were observed (cf. Matthews, 1977; E. Jyvaisjirvi & K.-D.Kniffki, unpublished observations).The conduction velocity was calculated using the shortest constant latency and

the distance between the stimulating cathode and the recording electrode. The meanconduction velocity (± S.D.) of Ad fibres was 13-9+ 6-4 m/s (range: 3-6-26-0 m/s) andthat of C fibres was 1-3 + 0 5 m/s (range: 0-5-2-2 m/s). The mean electrical threshold

A

1 ms

B

liI

^1 M_{<_S~~~~~~~~~~~~1MAAApftvo5 ms

lFig. 2. Responses of intradental Ad (A) and C fibres (B) to single electrical current pulsesapplied to the lower canine tooth crown in the cat. In A, five sweeps are superimposed,showing the constancy of the latency. Stimulation parameters: A, 1 ms, 40 ,uA = 2-2 x T(threshold); B, 1 ms, 200 PuA = 1-4 x T. The stimulation artifact is indicated by the arrows;note the different time scale in A and B.

using 1 ms cathodal constant-current pulses for activating Ad fibres was 62-3 + 41-4uzA (range: 10-180 1sA) and that for C fibres was 1044 + 53.7 1tA (range: 25-200 ,sA).As long as the tooth remained intact in the course of the experiment, none of the

identified intradental Ad or C fibres showed any ongoing activity in the absence ofintentional stimulation. Even after repeated cold stimulation of the tooth (see below)the fibres never developed an ongoing discharge.While searching for single afferent intradental Ad and C fibres, units with ongoing

discharge were regularly recorded. These could either be identified mechanically aslow-threshold periodontal afferent fibres or they were unexcitable by the methodsused. The periodontal mechanosensitive units could not be activated by the arrange-ment of electrodes used for tooth stimulation (Fig. 1 A) unless the stimulus greatlyexceeded a duration of 1 ms and an amplitude of 200 ptA.

197


Sensitivity to cold stimulationOnly those single-fibre or multi-fibre recordings were accepted for cold stimulation

of the lower canine tooth crown, in which each action potential could reliably bediscriminated according to spike amplitude and form. Every attempt was made toget single-fibre recordings (Figs. 3 and 4) before testing the cold responsiveness of theunits.

A 143 r/sI I 11iiu 111lI 'i l !I

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9

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5 sFig. 3. Responses of three identified intradental Ad fibres to cold stimulation of the lowercanine tooth they were innervating. Recordings of the temperature (T) near the pulp areindicated in the lower traces in A, B and C. Above the individual recordings of theneuronal activity the value of the conduction velocity of the responsive Ad fibre is given.The arrows mark the onset of cold stimulation.

Ad fibres. In Fig. 3 the typical responses of three identified Ad fibres to coldstimulation of the lower canine tooth are shown. The responses of Ad fibres to rapidlowering of the tooth temperature were rather uniform. After an initial high-frequency short-latency discharge the firing rate decreased and stopped when therate of change of temperature became small (Fig. 3).

All responding A& fibres reacted in a similar manner (cf. Fig. 10B); they wereactivated with an average latency of 1-7 + 1F1 s (range: 01-3-9 s). The mean responseduration was 21-0+ 9-2 s (range: 3-42 s); the mean decrease of the tooth temperature

198


at the onset of activation was 4-9 °C (range: 01-12-6 °C); the mean initial pulptemperature being 29-2 + 1-7 °C (range: 26-1-33-7 TC).As a rule, the fibres discharged as the rate of change of temperature (dT/dt) was

negative and the discharge ceased as the rate of change became zero. The fibres neverdischarged as dT/dt became positive, i.e. as the tooth temperature increased. The

5- 0(8 1)

4

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X3

y=014+084xEX 1 < < r = 089

0 L

0 1 2 3 4 5Maximum temperature change (°C/s)

Fig. 4. Relationship between the mean discharge rate of Ad fibres and the maximum rateof change of temperature ((dT/dt)max) achieved in a particular experiment. Around thetime at (dT/dt)max the mean discharge rate was determined using a 4 s interval. n = 22.

initial dynamic discharge was correlated with the maximum rate of change of tem-perature (dT/dt)max. This relationship is shown in Fig. 4. With a greater (dT/dt)maxthe mean discharge rate was also greater. Using a linear least-square fit procedure,the relationship was y = 0'14 +084 x, where y represents the mean discharge rateand x the maximum rate of change of temperature achieved in a particular experi-ment; the correlation coefficient being r = 089.

Cffibres. The response characteristics of the intradental C fibres to cold stimulationof the tooth they are innervating were also uniform but were completely differentfrom those of the Ad fibres. In Fig. 5 the cold-evoked discharge of three identifiedsingle intradental C fibres are shown. In contrast to the discharge behaviour of theAd fibres, no dynamic-response phase at dT/dt < 0 was observed. As a rule, a regular,low-frequency firing started as the rate of change of temperature was small andterminated as the rate of change of temperature became slightly positive.The average response latency was 7-3 + 2-5 s (range: 3-2-13-0 s); at that time the

mean tooth temperature had decreased to 10-1+ 3-2 0C (range: 18-4-7-2 °C); themean response duration being 52'0+ 15-5 s (range: 25-93 s). As in the Ad fibres, nodischarge was observed in C fibres as the tooth temperature returned to its initialvalue.As shown in Fig. 6 the mean discharge rate (y) during the period when the rate of

change of temperature was about zero correlated linearly (y = 1I- 0-05 x; r = 0-6)with the final temperature (x) achieved in a particular experiment. When the endtemperature was lower, the mean discharge rate was higher.

199

200 E. JYVASJARVI AiND K.-D. KNIFFKI

A 1 7 m/s

28

18 T (OC)

B 2-1 m/s

_ \ _ ~~~~~~~~~~27tk T (°C)

_ ~~~~9

C 0.9 m/s

27t - - -]12T (°C)

5 sFig. 5. Responses of three identified intradental C fibres to cold stimulation of the lowercanine tooth they were innervating. Above the individual recordings of the neuronalactivity the conduction velocity of the responding C fibre is given. Each of the lowertraces in A, B and C display the tooth temperature (T) near the pulp during the recording;arrows mark the onset of cold stimulation.

1 5

CD

E 1 0 *Ea .0

CX ~~* 0..* 0

.C 0.050 y= 1.1-0.05x*-c

.~05

2 4 6 8 10 12 14

End temperature (°C)

Fig. 6. Relationship between the mean firing rate ofC fibres in response to cold stimulationof the lower canine tooth and end temperature achieved in a particular experiment. Themean discharge rate was determined after the temperature change became zero. n = 26.

INTRADENTAL Ad AND C FIBRES AND SENSATION 201

Effectiveness of cold stimulation. Of the Ad fibres 84 %, thirty-six out of forty-three,and of the C fibres 88%, thirty-eight out of forty-three, were excited by the coldstimulation of the tooth they were innervating. In Fig. 7 the distribution of theconduction velocities of the recorded eighty-six intradental Ad and C fibres are shownas a histogram. As can be seen there was no correlation between the responses of the

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Conduction velocity (mis)Fig. 7. Distribution of the conduction velocities of eighty-six intradental Ad and C fibrestested by cold stimulation of the lower canine tooth they were innervating. Open columns,cold-responsive C fibres; filled columns, cold-responsive As fibres; hatched columns, non-responsive fibres. n = 86.

Receptive area tomechanical stimulation

Fig. 8. Projection to the sagittal plane of the location of mechanically determinedreceptive fields of eleven Ad and C fibres which responded to cold stimulation of the intactlower canine tooth crown. Note that the receptive fields of Ad fibres were located moresuperficially than the ones for C fibres which were located deep in the pulp. Filled circles,C fibres; open circles, Ad fibres.

intradental C fibres to cold stimulation of the tooth and their conduction velocity.For the Ad fibres Fig. 7 shows that those which did not respond to the cold stimuliapplied to the tooth had a conduction velocity above 10 m/s.

In eleven experiments after repeatedly eliciting responses to cold stimuli, the pulpwas carefully exposed, and by probing with a dental needle, the receptive fields of four

E. JYVASJARVI AND K.-D. KNIFFKJ

AM and seven C fibres were determined. Fig. 8 is a summary of the location of thesereceptive fields. Receptive fields of Ad fibres were located superficially at the pulp-dentine border, whereas those of C fibres were located deeper in the pulp. Afterexposing the pulp and searching for receptive fields, one Ad and four C unitsdeveloped an ongoing discharge.

A C50 -_

C0LS' t 1IB D50 -

tt,5 s

Fig. 9. Time course of the pain ratings of four human subjects to cold stimulation of incisorteeth. The arrows mark the beginning of application of the cold stimulus. Stimulus wasremoved as soon as the subject indicated a response.

Human experimentsThe aim of these experiments was to compare the intensity and time course of

discharges of cat intradental Ad and C fibres to a cold stimulus applied to a lowercanine tooth with sensory magnitude estimation in human subjects, when their owntooth was stimulated with the same kind of stimulus.

In general, the sensation evoked by the applied cold stimulus was described as asharp, shooting pain sensation, localized strictly to the tooth stimulated. No coldsensation or any other sensations were reported. Stimulation of the devital tooth didnot evoke any sensation at all. The typical time course of the ratings of four humansubjects to a cold stimulus applied to the test tooth is shown in Fig. 9. As can be seen,the intensity of the perceived pain sensation increased rapidly, reached a maximumwithin a few seconds and then decreased to zero. In sixteen trials in four subjects, a'shoulder' was observed where the rate of decrease in pain intensity was reduced fora few seconds before dropping steeply (Figs. 1B and 9A and C).The average latency between the onset of stimulation and the first perceived

sensation was 1-5+1 0 s (range: 0-2-6-0 s), and the mean duration of the perceivedsensation was 13-3 +4-6 s (range: 6-0-25-0 s). Repeated trials resulted in the sametype of perception with similar time courses. In Fig. IOA the averaged time courseof the ratings is shown. During the declining phase the 'shoulder' is just noticeableat about 10 s. The mean evoked activity of the Ad fibres is shown as an averaged

202


Human sensation

10 20 30 40Time (s)

50 60

'°r TT

A5 fibres

10 20 30 40 50 60Time (s)

C fibres

10 20 30 40Time (s)

50 60

Fig. 10. Comparison of human sensory ratings to cold stimulation of incisor teeth with theevoked discharges of single intradental Ad and C fibres by similar stimulation of lowercanine teeth in cats. A, mean time course of ratings of human subjects. The averagingprocedure included all seven subjects with their ten teeth being tested. Recordings asthose shown in Fig. 9 were A/D converted every second, and these values were averagedwith respect to the onset of the responses. n = 43. B, mean peristimulus time histogram(bin width: 1 s) ofthe evoked neuronal activity (impulses/s) ofAd fibres to cold stimulationof cat's canine teeth. The averaging procedure was performed using thirty-six fibres inforty-one trials. n = 41. C, similar histogram as in B (bin width: 1 s) for C fibre responses;thirty-eight fibres being tested forty-two times. n = 42. The bar underneath the onset ofthe responses in A, B and C expresses the mean latency + S.E. of mean of the responses,whereas those at the top of the columns indicate + S.E. of mean. The arrows indicate theonset of stimulation.

peristimulus time histogram in Fig. lOB, that of the C fibres is shown in Fig. 1OC.Note the marked differences in the intensity and time course of the responses of bothfibre types and the similarity in time course between human pain ratings and cat ASfibre responses.

DISCUSSION

The present results provide a comparison between the neuronal discharges of singleintradental AS and C fibres in the cat and the dental pain sensations in humansubjects in response to cold stimulation of the tooth. Because of methodological

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difficulties the response properties of intradental Ad and C fibres of intact teethcannot be studied by single-unit recordings in man. Therefore, since there are majorstructural similarities between human and cat teeth (Byers, Neuhaus & Gehrig,1982), an interspecies comparison was carried out.Most of the feline Ad and C fibres recorded were excited by cold stimulation of the

tooth they were innervating. This is in contrast to previous studies where the numberof cold-sensitive fibres was found to be small (Yamada et al. 1968; Matthews, 1977;Kollmann & Matthews, 1982; Nairhi et al. 1982 a; Nairhi, 1985). This difference mightbe due to the different methods of cold stimulation used in the various studies. It ispossible that with the methods previously used, the rate of change of toothtemperature was too slow and the temperature reduction was too small to excite thefibres. In some pilot experiments we used ethyl chloride to try to cool the tooth butthe pulpal temperature change was too slow and the temperature decrease too smallto evoke any reliable and reproducible responses particularly in C fibres. Obviouslya crucial factor in thermal stimulation of the tooth is the rapidity of the inducedtemperature change (Brannstrom & Astrom, 1964). Presumably this is why dichloro-difluoromethane and carbon dioxide snow are more reliable than pulp testing agentssuch as ice and ethyl chloride (Fuss, Trowbridge, Bender, Rickoff & Sorin, 1986).The present results revealed profound differences in the response properties of Ad

and C fibres to cold stimulation of the intact tooth they were innervating.Upon rapid lowering of the tooth temperature the Ad fibres responded with an

initial high-frequency discharge during the period of the temperature change. Thefiring rate decreased and ceased as the rate of change of temperature became lowerand zero, respectively. The initial dynamic discharge showed a good correlation withthe maximum rate of change of temperature obtained. This response behaviour isconsistent with the hypothesis that thermal stimuli cause fluid movement in dentinaltubules which results in mechanical activation of the Ad nerve endings located nearthe pulp-dentine border (Brannstrom & Astr6m, 1964,1972; Brannstrom & Johnson,1970). For some of the cold-sensitive Ad fibres the receptive fields were determinedand were found to be located at the pulp-dentine border. These loci and the lowthresholds to mechanical stimulation are further indications that the cold stimuliapplied to the tooth were acting indirectly to activate the receptive membrane of theAd fibres by a rapid movement of the contents of the tubules at the pulpal end.The cold-sensitive Ad units never discharged again as the tooth temperature

returned to the initial value. This does not argue against the hypothesis oftemperature-induced fluid movement as the stimulus responsible for activating theAd fibres, since it may be that the rate of change of the temperature during therewarming period was too small. It should be mentioned, however, that in someexperiments radiant heat applied to the tooth was used to increase the rewarmingrate, but still not a single action potential was observed during these rewarmingperiods. It was also shown by electrical stimulation of the tooth that during therewarming periods the axons themselves were not blocked. Thus the possibility arisesthat there is rectification in the activation of intradental Ad fibres by cooling of thetooth. However, further experiments are needed to demonstrate such a phenomenon.

Another finding is in agreement with the hypothesis that a rapid movement of thetubule contents might be responsible for the activation of the Ad nerve endings in the

204


pulpo-dentinal border. This is that none of the recorded single Ad fibres responsiveto cold stimulation of the canine tooth showed an ongoing discharge without inten-tional stimulation. Therefore, it seems unlikely that the recorded responses originatefrom specialized cold receptors, as it is known that these have a background discharge(Hensel, 1981). Also the intradental Ad fibres ceased responding to steady-statetemperatures whereas cutaneous cold receptors are known to have a static dischargeover part of their sensitivity range (Hensel, 1981). After repeatedly eliciting responsesto cold stimuli applied to the intact tooth none of the Ad fibres developed aspontaneously occurring discharge. Repeated tooth stimulation produced no signifi-cant alteration in the sensitivity of Ad fibres as long as about 1 h was allowed forrecovery between stimuli, but there was a progressive decrease in the responses(frequency and number of spikes) with intervals of about 5-15 min. A similar findinghas been reported by Kollmann & Matthews (1982). It might be that a disturbancein pulpal blood flow results in the observed depressed excitability of intradental Adfibres (Edwall & Scott, 1971) or that partly reversible damage might have occurred,altering the displacement of the dentinal tubule contents. Recently, however, usingethyl chloride as a cold stimulus reproducible responses were obtained in humansubjects using short stimulus intervals (Alquist, Franzen, Edwall, Fors & Haeger-stami, 1986).The response behaviour of intradental C fibres to rapid cooling of the tooth they

were innervating was completely different from that of Ad fibres. The C fibres startedto discharge very regularly without an initial dynamic response phase at a time whenthe rate of change of temperature was already very low or had even reached a steadylevel. The firing rate was weakly correlated with the static tooth temperature. Itseems that the C fibres were activated at a certain temperature threshold and werenot responding as did the Ad fibres to the rate of change of tooth temperature. Thisargues against the hypothesis that the C fibres are responding to hydrodynamic forcesat the pulpo-dentinal junction. It is possible that the reduced absolute value of thetemperature had a direct effect on the receptive membrane of the free nerve endingsof the C fibres, but a direct effect of cooling on the axon itself cannot be excluded(Matthews, 1977). In contrast to the present results, Niirhi, Jyviisjairvi, Hirvonen &Huopaniemi (1982b) did not find any C fibre responses to cooling of the tooth. Thisdiscrepancy might be explained by the different methods used for cooling the intacttooth.For some cold-sensitive C fibres the receptive fields were found to be located deep

in the pulp and compared with Ad fibres the mechanical thresholds were higher. Someof the cold-sensitive C fibres responded also to radiant-heat stimuli applied to theintact tooth crown (E. Jyvasjarvi & K.-D. Kniffki, unpublished observations). Thus,the intradental C fibres in some ways behave like the polymodal nociceptors foundin other tissues.

Like the cold-sensitive Ad fibres, the C fibres never discharged as the temperatureof the tooth returned to its initial value, even if the rewarming rate was increasedusing radiant heat. Electrical stimulation of the tooth showed that the axons werenot blocked. The mechanism of this rectification in the responses of intradental Cfibres remains unknown.As with Ad fibres, none of the C fibres developed an ongoing discharge after

205


repeatedly eliciting responses to cold stimuli applied to the intact tooth. But inmarked contrast to the Ad fibres, the responses of the C fibres were rather reproducibleeven if less than 5 min were allowed for recovery between cold stimuli.The response behaviour of AA and C fibres was remarkably uniform in each group

of fibres. This finding gives additional support to the proposal that a classificationinto Ad and C fibres can be reliably made by using monopolar electrical stimulationof the tooth (E. Jyviisjiirvi & K.-D. Kniffki, unpublished observations).

In the human experiments a cold stimulus similar to that used in the animalexperiments produced a sharp, shooting pain sensation, which could be accuratelylocalized to the tooth being tested. This is in agreement with the nature and qualityof the perceived sensation using cold stimulation of the tooth (Hensel & Mann, 1956;Naylor, 1964; Trowbridge et al. 1980; Ahlquist et al. 1984).

In the present experiments no sensation other than pain was perceived. This is incontrast to a recent study where cold perception in human teeth has been reported(Griisser et al. 1982). It might be that due to the rapid cooling of the teeth used inthe present experiments the evoked pain sensation has masked the sensation ofcold.No convincing evidence of a delayed, dull pain sensation usually associated with

activity of afferent C fibres was found in the present study thus contrasting thefinding of Hensel & Mann (1956). Only in a few cases the subjects described such a

sensation. In these cases the rating curves in the declining phase showed a shoulder,i.e. a period of time where the rate of decrease in the pain intensity was reduced. Butthis shoulder was also observed in some trials where the subjects did not report any

change in the quality of the sensation. Averaging the response curves of all trials, theshoulder is just noticeable at about 10 s in Fig.1OA. Interestingly, the occurrence ofthe shoulder and the onset of the evoked C fibre activity in the animal experimentswere coincident (Fig.10C). Whether there is a causal relationship, i.e. whether Cfibres are aptivated in humans at that time cannot be decided from the presentresults. It should be mentioned, however, that in a recent study recording intradentalnerve activity from dentin in human teeth, most probably fromAd fibres, afterintense cold stimulation some subjects reported a pain sensation after the recordednerve activity had returned to its base-line level (Ahlquist et al. 1984). This findingcould well be explained by the different response behaviour ofAd and C fibres shownin Fig. lOB and C.

In conclusion, a comparison of stimulus intensity, response latency, time course

and duration of the neuronal responses of the felineAd and C fibres with thecorresponding pain ratings of human subjects to cold stimulation of the toothsuggests that intradentalAd fibres with the properties found in the cat are probablyresponsible for the cold-evoked sharp, shooting dental pain sensation in man.

The authors express their gratitude to Mrs Christa Erhard and Mrs Petra Haumann forcompetent technical assistance in the laboratory, and to both of them and to Mrs Margit Schulzefor patient help in the process of preparing the Figures and the manuscript. We are grateful to DrW. Kollmann, Dr R. W. A. Linden, Professor Dr H.-D. Mierau and to Dr U. Proske for theircritical comments on the manuscript. The project has been supported by the Wilhelm Sander-Stiftung (grant No. 83.018.1/2).

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