- Email: [email protected]
Behavioral self warming and cooling of spinal canal by rats
Physiology & Behavior, Vol. 28, pp. 489-495. Pergamon Press and Brain Research Publ., 1982. Printed in the U.S.A.
Behavioral Self Warming and Cooling of Spinal Canal by Rats I B. DIB, M. C O R M A R E C H E - L E Y D I E R
A N D M. C A B A N A C
Universit( Claude Bernard, L.A. C . N . R . S . 181 Facult( de M ( d e c i n e Lyon-Sud, Laboratoire de Physiologie, B.P. 12, 69600 Oullins, France R e c e i v e d 25 A u g u s t 1981 DIB, B., M. CORMARECHE-LEYDIER AND M. CABANAC. Behavioral self warming and cooling of spinal canal by rats. PHYSIOL. BEHAV. 28(3) 489--495, 1982.--Rats with a chronic thermode implanted in their spinal canal could bar-press to warm or cool their spinal cord. With a "cold" lever, they cooled their spinal canal less in a cold environment than in a warm environment. With a "warm" lever they behaved in the same way, i.e., warmed their spinal canal more in a warm than in a cold environment. In a two-lever situation they pressed the cold and the warm levers alternately in warm environment, but did not press either in cold environment. These results suggest that cold and warm spinal cord provided the rats with rewards of a different nature. Self warming
Self cooling
Spinal canal
Reward
E X P E R I M E N T A L cooling and warming of a thermode chronically implanted in the spinal canal is followed by the whole pattern of thermoregulatory cold and warm autonomic responses. These responses show the existence of a temperature sensitivity in the spinal cord. This is now well documented in several species [10, 12, 13, 14, 15, 18, 19]. That this sensitivity is able, also, to elicit various thermoregulatory behaviors has been shown in pigeon [15, 16, 17], frog [9], pig [3] and dog [6]. What would happen if the animal had access to a lever modifying not environmental temperatures as in the experiments referenced above, but the temperature of the deep sensor itself?. Such an experiment was carried out by Corbit [4], Corbit and Ernits [5] in rats and by Adair [1] in monkeys. In both experiments the thermode was chronically implanted in the hypothalamus. The animals used the lever to cool the thermode when it had been warmed by the experimenter. However, in a cold environment, pigs did not use operant behavior to warm their neutral hypothalamic thermodes [2]. Discrepancies may be due to the fact that hypothalamic temperature sensitivity in the rat is not symmetrical, as far as warming and heating by the experimenter are concerned. Rats seem to respond consciously to warming o f the hypothalamus but not to the cooling of it (Cabanac and Dib, publication pending). These observations led us to extend the experimentation to spinal cord temperature sensitivity. Would rats use a lever changing the temperature of their own spinal cord? Dogs did warm their spinal thermode but only in a warm environment [6]. This somewhat puzzling result also needed to be explored somewhat further.
Thermoregulatory behavior
GENERAL METHOD Rats were chronically implanted with a water perfused thermode placed in their spinal canal. By pressing a lever they could modify the temperature o f the water circulating in the thermode. This behavior was measured in warm, cool or cold environments while the water perfused was cold, at body temperature or warm. Experiments started 10 to 15 minutes after placing the rat in the apparatus, in order to let the animal stabilize its body temperatures and exploratory behavior.
Animals and Surgery Twenty four rats were used in the experiments. They were reared in individual cages. They weighed between 380 and 450 g. Each received a polyethylene U-shaped catheter of internal and external diameters 0.3 mm and 0.7 mm, respectively. Under 35 mg-kg-' barbiturate general anesthesia the thermode was introduced in the epidural space to 4.5 cm from T2, down to %. The free ends of the catheter were sutured to T2 then left protruding from the rat's neck. Each rat received 36,000 units o f depot penicillin subcutaneously. After surgery and recovery, the rats were apparently in good health, their motility was normal and they survived from 3 weeks to 7 months. Ten rats with apparent nervous trouble were discarded. The rats were petted and handled daily in order to minimize the effect o f future experiments.
Climatic Chamber During the experiments the rats were placed in a wire
~This work was supported by I.N.S.E.R.M. (Institut National de la Sant6 et de la Recherche M6dicale) A.T.P. 80-79-112, C.N.R.S. (Centre National de la Recherche Scientifique) L.A. N ° 181, et C.N.R.S.G.I.S. Institut de Physiologic Sensorielle.
489
490
D1B, C O R M A R E C H E - L E Y D I E R AND CABANA(
mesh cylinder 0.31 m high and 0.35 m diameter. This cylindrical cage itself was placed inside a slightly larger double wall cylinder made of copper. The space between the copper walls was filled with constantly stirred water at regulated temperatures. Hot, cool and cold wall temperatures provided, near the rat, air temperatures of respectively 35, 20 or 10°C. The wire mesh cage was equipped with two levers, L~ and L2, 55 mm long and 5 mm broad, 140 mm apart and 20 mm above the cage floor.
Thermode Perfusion During the experiments, tap water at 20°C was continuously circulated through the spinal thermode. The flow of water through the thermode was 8-11 ml-mn-L The actual temperature of the water was changed, just prior to entering the thermode, i.e., 15 mm above the rat's skin, using a spiral resistance heater R1 placed around the inlet. This resistor was fed by an adjustable rheostat (Fig. 1). By simple adjustment of the rheostat the experimenter could thus cool the thermode (31.1
FIG. 1. Experimental set for the measurement of self spinal cooling and warming. LI and Lz, levers; R~ and R2, heating resistors; Rh~ and Rh2, rheostats; C~ and C2, time meters; W, water; P, pump.
Training and Behavior Measurement The experiment started only after the rats had been trained to the experimental cage and to lever use. Training was considered to be obtained when this behavior was stable. Experiment I. A one hour session was sufficient for the rats to learn how to the press lever and self cool their spinal canal when it was heated by the experiment (40
pressing in periods of 30 min. Results presented here were averaged as interindividual means.
Recordings In addition to behavior recording, four temperatures were continuously recorded: three from the rat and one environmental. All sensors were thermocouptes, recorded on a potentiometric recorder with a sensitivity of 0.1°C. One thermocouple was placed 70 mm inside the colon (Tre), one was taped on the skin at the dorsal side at the basis of the tail (Ts), one measuring water temperature 20 mm prior to its entry into the thermode (Tth). Tth values given in the text correspond to extremes of variation in perfusion temperature. During thermode peffusion spinal cord temperature variations were less extreme than thermode temperature variations. Ambient temperature (Ta) in the climatic chamber was measured not far from the rat. The temperature of the experimental room was kept constant (20_+ l°C). Although entangled with 2 tubes (thermode perfusion and evacuation) thermocouples (Tre, Ts, Tin) and 4 wires feeding the coil resistors, rats were largely free to move. Details on the techniques permitting the connection of a freely moving
SPINAL C A N A L T H E R M O R E G U L A T I O N
491
SPINAL SELF-COOLING
TIME (s)
15 '¢
a
•
b
46
T a . 20°¢ 42 4O 0
,.c
38 36 34 32 3O 28 26
O
,
0
,
,
20
,
.
*
40
|
60 TIME
L
A
.
.
o ZO (min)
•
40
1
•
,
I0
o C
S
SPINAL
FIG. 2. Sample of thermode temperature recorded in a cool environment (a) and in a warm environment (b). At time 0 the rat was placed in the climatic chamber and the spinal thermode was heated at about time 10 min. Tin: temperature of the water perfusing the spinal thermode. The abrupt and deep depressions of Tm are the result of the rat's self stimulating behavior. On both instances the spinal thermode was peffused with hot water at almost 44°C; self stimulation resulted in spinal cooling.
rat to several pressure tubes and thermocouples have been published elsewhere [8]. E X P E R I M E N T I: SELF COOLING In this experiment, pressure on lever L, by the rat cooled the spinal thermode. This behavior was measured at two ambient temperatures (20°C and 35°C) and at three temperatures of the water perfusing the thermode: cold (33.1
Warm Spinal Perfusions in a Cool Environment (20°C) Figure 2a gives a typical example of the evolution of the variables during a session of spinal self cooling in a rat with a warm thermode (44
W
C
8
W
SELF-COOLING
FIG. 3. Average (_+S.E.) behavioral response of the group of rats placed in a hot (a) or cold (b) environment. The behavior recorded was the time spent pressing the lever to obtain a cooling of the spinal thermode. The water perfusing the thermode in the absence of the rat's behavior was either cold (C): 33.1-35°C, body temperature (B): 38.2-40.8°C, or warm (W): 41.2-44.5°C (*p<0.02, Student t-test with nearest column).
heating, it thus brought water temperature down to nearly 28.5°C. For the 30 min session, total lever pressing time was 60 sec. Heating of the spinal cord was followed by skin vasodilatation and decrease in rectal temperature. Seven experiments were performed on the six rats. Mean self cooling time for the group is shown Fig. 3b (column W).
Warm Spinal Perfusions in a Warm Environment (35°C) Figure 2b gives an example of this situation. It can be seen that the rat, after 30 sec latency, repeatedly pressed the lever and sometimes kept it depressed for several minutes. It thus lowered its spinal thermode temperature to between 28.5 and 26.5°C. In this session the rat pressed the lever 5 min 30 sec every 30 rain. In addition, Fig. 2 shows that bursts of lever pressing were followed by skin vasoconstriction. Figure 3a, column W shows the average self spinal cooling time of the six rats at 35°C ambient temperature and with a heated spinal canal.
Cold and Neutral Spinal Perfusions Figure 3 shows the mean self spinal cooling time of the
492
DIB, C O R M A R E C H E - L E Y D I E R AND CABANA(I
rats when ambient temperature was warm, 35°C (Fig. 3a) or cool, 20°C (Fig. 3b). Columns B and C refer to the temperature of the water perfusing the spinal thermode, at body temperature or cold (33.1
SPINAL SELF-HEATING 20'c
48
~5 %
46
i
44
J,
42
U o
40 3111 36 34 32
b
0 •
O
•
•
N
I
•
U
!
J
,
~
~
.
N 0 M TIME (rain)
.
.
.
.
4O
U
FIG. 4. Sample of thermode temperature recorded under various conditions. Tth: temperature of the water perfusing the spinal thermode. The deep changes of Tth are the result of the rat's self stimulating behavior. The spinal thermode was perfused with cold water at almost 34°C; self stimulation resulted in spinal heating.
recorded at two environmental temperatures, 20°C and 35°C, and three temperatures of the water perfusing the spinal thermode; cold (C: 31.1
Cold Spinal Perfusions in a Cool Environment (200(,") Figure 4a gives a typical example of the results obtained in these conditions. When the spinal canal was cooled to 33
Cold Spinal Perfusions in a Warm Environment (35°C) Figure 4b gives an example of this situation. It can be seen that the rat repeatedly pressed the lever and warmed its spinal thermode. This behavior appeared following 6 min latency after the onset of the cold perfusion, Maximal spinal temperature chosen by the rat was between 44 and 480CI and total lever pressing time computed for 30 rain was 4 min. Sixteen experiments were performed on seven rats whose spinal canal was cooled (31.1
493
SPINAL CANAL THERMOREGULATION
T I M E Is~
b
fl
OO
450
3OO
150
0
C
B
W
SPINAL
C
B
W
SELF - HEATI NG
FIG. 5. Same legend as Fig. 2, but here the rats warmed their spinal thermode when they pressed the lever (**p<0.01, Student t-test with nearest column).
warm environment; indeed their rectal temperature rose. Self spinal warming was also obtained in one dog [7]. In that case the results were identical to those obtained here in rats: increase in ambient temperature also increased the behavior of self spinal warming. Figure 5b, column C shows that, contrary to what was expected, rats did not self warm their spinal cord in a cold environment. This is paradoxical. Since they did so in a warm environment the cold signal alone is sufficient to arouse the behavior. One would, then, expect them to self warm more in a cold environment when a skin cold signal is added to the spinal cold stimulus. The cooling of the spinal canal was the same in both experiments, therefore the difference originated in the cold skin. Two possible explanations can be hypothetized to explain this behavior: (a) nervous input from a cold skin would inhibit the transmission of the cold spinal signal to the higher centres. This mechanism would also suppress self spinal cooling in a cold environment. (2) a cold skin would arouse a sensation marking the spinal sensation. Thus in a cold environment a spinal cooling would leave the rat unaware of it. Other negative results obtained in this experiment could be expected and serve as control.
E X P E R I M E N T III: C H O I C E
Warm and Neutral Spinal Perfusions Figure 5 shows the mean self spinal warming time o f the rats when ambient temperature was warm 35°C (Fig. 5a), or cool 20°C (Fig. 5b). Columns W and B refer to the water perfusing the spinal thermode, warm (41.4
Among the six experimental conditions combining 3 thermode temperatures and 2 ambient temperatures, only one case gave positive results statistically different from all other cases: rats self warmed their spinal canal in a warm environment when the thermode was cooled. This result shows the existence of a cold sensitivity, probably in the spinal cord. In this regard rats responded in the same way as pigs [31 and pigeons [17] but contrary to dogs [6]. Self warming was opposite to spinal cooling applied by the experimenter. The behavior was present in a warm environment in absence of a cool skin. Cold spinal canal therefore was necessary and sufficient to produce self spinal heating and was thus different from self spinal cooling (Experiment I). The temperature obtained by the rats was usually about 42.5°C at the entry of the thermode. Even if there was some heat loss between the thermocouple and the spinal canal one may assume that thermode temperature was higher than deep body temperature (38.5 to 39°C). The behavior therefore not only removed the cooling but also warmed the spinal canal. This may be the reason for the lever pressing: the rats would thus actuate their autonomic response and in turn better resist the
Experiment I showed that rats suppressed the warming of their spinal canal and self cooled it in a warm environment. Experiment II showed that rats suppressed the cooling of their spinal canal and self warmed it in a warm environment. Thus the rats evidence contradictory behaviors. In order to solve this contradiction we placed the rats in a situation of choice. In this experiment the rats had access simultaneously to two levers. Depressing the first lever warmed the water perfusing the spinal thermode. Depressing the second lever cooled it. When no lever was depressed water at the rat's body temperature circulated in the thermode. Nine rats served in this experiment. Each rat was placed for 90 min sessions at environmental temperatures of 10°C, 20°C and 35°C, 30 min each. In order to cancel out the influence of environmental temperature sequence, in half the sessions the sequence was reversed: 35°C, 20°C and 10°C. RESULTS AND DISCUSSION
In a warm environment, 20 experiments were performed on 9 rats whose spinal temperature was maintained at between 37.6 and 38.8°C. In a cool environment 20 experiments were performed on 9 rats whose spinal temperature was maintained at between 37.7 and 39.6°C. In a cold environment 13 experiments were performed on 8 rats whose spinal temperature was maintained at between 37.6 and 38.8°C. Figure 6 gives an example of a sequence where the experimental sequence started at 35°C Ta and was then cooled to 20 and 10°C. The change in ambient temperature was rapidly obtained by adding ice in the chamber wall. In sessions where Ta was raised equally rapid increase was obtained by adding hot water in the wall of the climatic chamber. Figure 7 shows the mean time spent pressing the lever by the group of rats. This figure shows that the rats bar-pressed a significant length of time in hot environment and little time in cold environment. The rats pressed on both levers, alternatively. Each press lasted 20 sec to 1 min and produced thermode temperature changes extending to 26°C and to
494
DI B. CORMARECHE-LEYD1ER AN D (:ABAN: ,~(' EXPERIMENT III :CHOICE
TIME ON COLD (s) TIME ON WARM (s)
302
90
125
47
TIME (sl 0 O
la=asCi Ia:20~C '~Ta:lo~C
41
,;ot
']9
37
35 .O
33
e"
31
TWO 29
27
25 i
i
20
1
t
40
1
t0
6
1
i
1
8~
TIME (min)
FIG. 6. Sample of thermode temperature recorded under various conditions. Tin: temperature of the water perfusing the spinal thermode. Data correspond to bar-pressing for cold and warm spinal rewards. The graph shows the fall and subsequent rise of Tth measured, but it does not show at what precise moment the rat pressed the warming lever. This is obtained from the counters (see Fig. 1). The spinal thermode was perfused with water at the rat's body temperature.
43°C. The latency for a temperature change after a press was between 10 and 15 sec. When in the hot environment a rat sometimes did not use either lever. If it started to use one, it usually immediately switched to the other, then came back to the first, and so on as long as the environment remained hot. As a result the time spent on both levers was comparable, as seen in Fig. 7. The increase of both responses at higher Ta (Fig. 7) may suggest that bar-pressing could reflect a higher activity at high Ta, the response would thus be non specific. This is unlikely because the time spent bar-pressing seemed to be adjusted not only to ambient temperature, but also to spinal stimulus temperature. Such adjustment is evident in Figs. 3 and 5 (results of Experiments I and II). The result of Experiment III confirms the results of both Experiment I and II. Since the rats bar-pressed alternately on warming and cooling levers in hot environment only, they produced, simultaneously, both the results of Experiment I and Experiment II. This behavior suggests that a conscious signal was perceived by the rat only when it started pressing one lever and was then condemned to run from the one to the other in order to correct the warm and cold spinal signals alternately.
[--'1
LEVER
CHOICE
SPINAL
SELF -HEATING
SPINAL
SELF-COOLING
FIG. 7. Mean results of Experiment 111. The rats could both self warm and self cool the water perfusing their spinal thermode. Columns show the mean time (_+S.E.) spent pressing the levers at three different ambient temperatures (T,). When the rat did not press the lever, water at its own body temperature was perfused in the thermode.
G E N E R A L DISCUSSION When a rat bar-presses to modify its own spinal temperature, three explanations of this behavior are possible: (1) the reward lies in the spinal canal, i.e., there is a sensation aroused from the spinal cord or its vicinity. (2) the reward originates from a modified skin sensation. Cutaneous afferent thermoreceptors travel in the spinal cord, a modified spinal temperature could modify the afferent message. (3) a thermal spinal stimulus arouses themoregulatory (and perhaps non thermoregulatory) autonomic responses, which could be rewarding in themselves or in modifying body temperatures in a direction opposite to the spinal stimulus. Let us look at the results on this basis. Self cooling of the spinal canal by the rats (Experiment I) confirms the existence of a warm sensitivity arousing autonomic and behavioral responses. Figure 3 shows that a warm skin temperature increased the spinal cooling, i.e., spinal and skin temperatures would have an influence of the s a m e n a t u r e . Thus it is not impossible that this behavior conforms to the first of the 3 possibilities listed above. The results of Experiment II show that rats corrected their spinal low temperature only at high environmental temperature, and this confirms preliminary results obtained in one dog [7]. Thus warm skin temperature was necessary, as if it was possible for a cold sensitivity of the spinal cord to exist only when the skin is warm. This would suggest that this sensitivity could conform to the second of the three hypothesis listed above. The main results of Experiment III confirm the results of both Experiment I and Experiment II at the same time. In addition, the fact that the same hot environment was able to arouse both self spinal cooling and self spinal warming simultaneously confirms the hypothesis that the rewards obtained were of different nature.
SPINAL CANAL THERMOREGULATION
495 REFERENCES
1. Adair, E. R. Preference for skin cooling during hypothalamic heating. Proceedings of the International Union of Physiological Sciences, XIII: 8, 1977. 2. Baldwin, B. A. and D. L. Ingram. The effect of heating and cooling the hypothalamus on behavioural thermoregulation in pigs. J. Physiol., Lond. 191: 375-392, 1967. 3. Carlisle, H. J. and D. L. Ingram. The effects of heating and cooling the spinal cord and hypothalamus on thermoregulatory behaviour in the pig. J. Physiol., Lond. 231: 353-364, 1973. 4. Corbit, J. D. Behavioral regulation of hypothalamic temperature. Science 166: 256-257, 1969. 5. Corbit, J. D. and T. Ernits. Specific preference for hypothalamic cooling. J. comp. physiol. Psychol. 86: 24-27, 1974. 6. Cormareche-Leydier, M. and M. Cabanac. Influence de stimulations thermiques de la moelle 6pini~re sur le comportement thermor6gulateur du chien. Pfliigers Arch. 341: 313-324, 1973. 7. Cormareche-Leydier, M. and M. Cabanac. Dog behaviour as related to spinal cord temperature. Experientia 32: 66--67, 1976. 8. Dib, B. Coninuous perfusion under pressure using several tubes in the freely behaving rat. Physiol. Behav. 24: 177-178, 1980. 9. Duclaux, R., M. Fantino and M. Cabanac. Comportement thermor6gulateur chez Rana esculenta. Influence du r6chauffement spinal. Pfliigers Arch. 342: 347-358, 1973. 10. Hales, J. R. S. and C. Jessen. Increase of cutaneous moisture loss caused by local heating of the spinal cord in the ox. J. Physiol. Lond. 204: 40P-42P, 1969.
I 1. Hamilton, C. L. Effect of food deprivation on thermal behavior of the rat. Proc. Soc. exp. Biol. Med. 100: 354-356, 1959. 12. Jessen, C., E. Simon and R. Kullman. Interaction of spinal and hypothalamic thermodetectors in body temperature regulation in the conscious dog. Experientia 24: 694-695, 1968. 13. Jessen, C. Panting in waking dogs by isolated heating of the spinal cord. Naturwissenchaften 54: 290-291, 1967. 14. Kosaka, M., E. Simon and R. Thauer. Shivering in intact and spinal rabbits during spinal cord cooling. Experientia 23: 385387, 1967. 15. Rautenberg, W. Die Bedentung der Zentralnerv6sen Therm6sensitivit~it for die Temperaturregulation der Taube. Z. vergl. Physiol. 62: 235-266, 1969. 16. Schmidt, I. Effect of central thermal stimulation on the thermoregulatory behaviour of the pigeon. Pfliigers Arch. 363: 271272, 1976. 17. Schmidt, I. Behavioral and autonomic thermoregulation in heat stressed pigeons modified by central thermal stimulation. J. comp. Physiol. 127: 75-87, 1978. 18. Simon, E., W. Rautenberg, R. Thauer and M. Iriki. Ausl6sung thermoregulatorischer Reaktionen durch lokale Ktihlung im Vertebralkanal. Naturwissenchaften 50: 337-339, 1963. 19. Simon, E., W. Rantenberg and C. Jessen. Initiations of shivering in unanesthetized dogs by local cooling within the vertebral canal. Experientia 21: 477, 1965.
Behavioral Self Warming and Cooling of Spinal Canal by Rats I B. DIB, M. C O R M A R E C H E - L E Y D I E R
A N D M. C A B A N A C
Universit( Claude Bernard, L.A. C . N . R . S . 181 Facult( de M ( d e c i n e Lyon-Sud, Laboratoire de Physiologie, B.P. 12, 69600 Oullins, France R e c e i v e d 25 A u g u s t 1981 DIB, B., M. CORMARECHE-LEYDIER AND M. CABANAC. Behavioral self warming and cooling of spinal canal by rats. PHYSIOL. BEHAV. 28(3) 489--495, 1982.--Rats with a chronic thermode implanted in their spinal canal could bar-press to warm or cool their spinal cord. With a "cold" lever, they cooled their spinal canal less in a cold environment than in a warm environment. With a "warm" lever they behaved in the same way, i.e., warmed their spinal canal more in a warm than in a cold environment. In a two-lever situation they pressed the cold and the warm levers alternately in warm environment, but did not press either in cold environment. These results suggest that cold and warm spinal cord provided the rats with rewards of a different nature. Self warming
Self cooling
Spinal canal
Reward
E X P E R I M E N T A L cooling and warming of a thermode chronically implanted in the spinal canal is followed by the whole pattern of thermoregulatory cold and warm autonomic responses. These responses show the existence of a temperature sensitivity in the spinal cord. This is now well documented in several species [10, 12, 13, 14, 15, 18, 19]. That this sensitivity is able, also, to elicit various thermoregulatory behaviors has been shown in pigeon [15, 16, 17], frog [9], pig [3] and dog [6]. What would happen if the animal had access to a lever modifying not environmental temperatures as in the experiments referenced above, but the temperature of the deep sensor itself?. Such an experiment was carried out by Corbit [4], Corbit and Ernits [5] in rats and by Adair [1] in monkeys. In both experiments the thermode was chronically implanted in the hypothalamus. The animals used the lever to cool the thermode when it had been warmed by the experimenter. However, in a cold environment, pigs did not use operant behavior to warm their neutral hypothalamic thermodes [2]. Discrepancies may be due to the fact that hypothalamic temperature sensitivity in the rat is not symmetrical, as far as warming and heating by the experimenter are concerned. Rats seem to respond consciously to warming o f the hypothalamus but not to the cooling of it (Cabanac and Dib, publication pending). These observations led us to extend the experimentation to spinal cord temperature sensitivity. Would rats use a lever changing the temperature of their own spinal cord? Dogs did warm their spinal thermode but only in a warm environment [6]. This somewhat puzzling result also needed to be explored somewhat further.
Thermoregulatory behavior
GENERAL METHOD Rats were chronically implanted with a water perfused thermode placed in their spinal canal. By pressing a lever they could modify the temperature o f the water circulating in the thermode. This behavior was measured in warm, cool or cold environments while the water perfused was cold, at body temperature or warm. Experiments started 10 to 15 minutes after placing the rat in the apparatus, in order to let the animal stabilize its body temperatures and exploratory behavior.
Animals and Surgery Twenty four rats were used in the experiments. They were reared in individual cages. They weighed between 380 and 450 g. Each received a polyethylene U-shaped catheter of internal and external diameters 0.3 mm and 0.7 mm, respectively. Under 35 mg-kg-' barbiturate general anesthesia the thermode was introduced in the epidural space to 4.5 cm from T2, down to %. The free ends of the catheter were sutured to T2 then left protruding from the rat's neck. Each rat received 36,000 units o f depot penicillin subcutaneously. After surgery and recovery, the rats were apparently in good health, their motility was normal and they survived from 3 weeks to 7 months. Ten rats with apparent nervous trouble were discarded. The rats were petted and handled daily in order to minimize the effect o f future experiments.
Climatic Chamber During the experiments the rats were placed in a wire
~This work was supported by I.N.S.E.R.M. (Institut National de la Sant6 et de la Recherche M6dicale) A.T.P. 80-79-112, C.N.R.S. (Centre National de la Recherche Scientifique) L.A. N ° 181, et C.N.R.S.G.I.S. Institut de Physiologic Sensorielle.
489
490
D1B, C O R M A R E C H E - L E Y D I E R AND CABANA(
mesh cylinder 0.31 m high and 0.35 m diameter. This cylindrical cage itself was placed inside a slightly larger double wall cylinder made of copper. The space between the copper walls was filled with constantly stirred water at regulated temperatures. Hot, cool and cold wall temperatures provided, near the rat, air temperatures of respectively 35, 20 or 10°C. The wire mesh cage was equipped with two levers, L~ and L2, 55 mm long and 5 mm broad, 140 mm apart and 20 mm above the cage floor.
Thermode Perfusion During the experiments, tap water at 20°C was continuously circulated through the spinal thermode. The flow of water through the thermode was 8-11 ml-mn-L The actual temperature of the water was changed, just prior to entering the thermode, i.e., 15 mm above the rat's skin, using a spiral resistance heater R1 placed around the inlet. This resistor was fed by an adjustable rheostat (Fig. 1). By simple adjustment of the rheostat the experimenter could thus cool the thermode (31.1
FIG. 1. Experimental set for the measurement of self spinal cooling and warming. LI and Lz, levers; R~ and R2, heating resistors; Rh~ and Rh2, rheostats; C~ and C2, time meters; W, water; P, pump.
Training and Behavior Measurement The experiment started only after the rats had been trained to the experimental cage and to lever use. Training was considered to be obtained when this behavior was stable. Experiment I. A one hour session was sufficient for the rats to learn how to the press lever and self cool their spinal canal when it was heated by the experiment (40
pressing in periods of 30 min. Results presented here were averaged as interindividual means.
Recordings In addition to behavior recording, four temperatures were continuously recorded: three from the rat and one environmental. All sensors were thermocouptes, recorded on a potentiometric recorder with a sensitivity of 0.1°C. One thermocouple was placed 70 mm inside the colon (Tre), one was taped on the skin at the dorsal side at the basis of the tail (Ts), one measuring water temperature 20 mm prior to its entry into the thermode (Tth). Tth values given in the text correspond to extremes of variation in perfusion temperature. During thermode peffusion spinal cord temperature variations were less extreme than thermode temperature variations. Ambient temperature (Ta) in the climatic chamber was measured not far from the rat. The temperature of the experimental room was kept constant (20_+ l°C). Although entangled with 2 tubes (thermode perfusion and evacuation) thermocouples (Tre, Ts, Tin) and 4 wires feeding the coil resistors, rats were largely free to move. Details on the techniques permitting the connection of a freely moving
SPINAL C A N A L T H E R M O R E G U L A T I O N
491
SPINAL SELF-COOLING
TIME (s)
15 '¢
a
•
b
46
T a . 20°¢ 42 4O 0
,.c
38 36 34 32 3O 28 26
O
,
0
,
,
20
,
.
*
40
|
60 TIME
L
A
.
.
o ZO (min)
•
40
1
•
,
I0
o C
S
SPINAL
FIG. 2. Sample of thermode temperature recorded in a cool environment (a) and in a warm environment (b). At time 0 the rat was placed in the climatic chamber and the spinal thermode was heated at about time 10 min. Tin: temperature of the water perfusing the spinal thermode. The abrupt and deep depressions of Tm are the result of the rat's self stimulating behavior. On both instances the spinal thermode was peffused with hot water at almost 44°C; self stimulation resulted in spinal cooling.
rat to several pressure tubes and thermocouples have been published elsewhere [8]. E X P E R I M E N T I: SELF COOLING In this experiment, pressure on lever L, by the rat cooled the spinal thermode. This behavior was measured at two ambient temperatures (20°C and 35°C) and at three temperatures of the water perfusing the thermode: cold (33.1
Warm Spinal Perfusions in a Cool Environment (20°C) Figure 2a gives a typical example of the evolution of the variables during a session of spinal self cooling in a rat with a warm thermode (44
W
C
8
W
SELF-COOLING
FIG. 3. Average (_+S.E.) behavioral response of the group of rats placed in a hot (a) or cold (b) environment. The behavior recorded was the time spent pressing the lever to obtain a cooling of the spinal thermode. The water perfusing the thermode in the absence of the rat's behavior was either cold (C): 33.1-35°C, body temperature (B): 38.2-40.8°C, or warm (W): 41.2-44.5°C (*p<0.02, Student t-test with nearest column).
heating, it thus brought water temperature down to nearly 28.5°C. For the 30 min session, total lever pressing time was 60 sec. Heating of the spinal cord was followed by skin vasodilatation and decrease in rectal temperature. Seven experiments were performed on the six rats. Mean self cooling time for the group is shown Fig. 3b (column W).
Warm Spinal Perfusions in a Warm Environment (35°C) Figure 2b gives an example of this situation. It can be seen that the rat, after 30 sec latency, repeatedly pressed the lever and sometimes kept it depressed for several minutes. It thus lowered its spinal thermode temperature to between 28.5 and 26.5°C. In this session the rat pressed the lever 5 min 30 sec every 30 rain. In addition, Fig. 2 shows that bursts of lever pressing were followed by skin vasoconstriction. Figure 3a, column W shows the average self spinal cooling time of the six rats at 35°C ambient temperature and with a heated spinal canal.
Cold and Neutral Spinal Perfusions Figure 3 shows the mean self spinal cooling time of the
492
DIB, C O R M A R E C H E - L E Y D I E R AND CABANA(I
rats when ambient temperature was warm, 35°C (Fig. 3a) or cool, 20°C (Fig. 3b). Columns B and C refer to the temperature of the water perfusing the spinal thermode, at body temperature or cold (33.1
SPINAL SELF-HEATING 20'c
48
~5 %
46
i
44
J,
42
U o
40 3111 36 34 32
b
0 •
O
•
•
N
I
•
U
!
J
,
~
~
.
N 0 M TIME (rain)
.
.
.
.
4O
U
FIG. 4. Sample of thermode temperature recorded under various conditions. Tth: temperature of the water perfusing the spinal thermode. The deep changes of Tth are the result of the rat's self stimulating behavior. The spinal thermode was perfused with cold water at almost 34°C; self stimulation resulted in spinal heating.
recorded at two environmental temperatures, 20°C and 35°C, and three temperatures of the water perfusing the spinal thermode; cold (C: 31.1
Cold Spinal Perfusions in a Cool Environment (200(,") Figure 4a gives a typical example of the results obtained in these conditions. When the spinal canal was cooled to 33
Cold Spinal Perfusions in a Warm Environment (35°C) Figure 4b gives an example of this situation. It can be seen that the rat repeatedly pressed the lever and warmed its spinal thermode. This behavior appeared following 6 min latency after the onset of the cold perfusion, Maximal spinal temperature chosen by the rat was between 44 and 480CI and total lever pressing time computed for 30 rain was 4 min. Sixteen experiments were performed on seven rats whose spinal canal was cooled (31.1
493
SPINAL CANAL THERMOREGULATION
T I M E Is~
b
fl
OO
450
3OO
150
0
C
B
W
SPINAL
C
B
W
SELF - HEATI NG
FIG. 5. Same legend as Fig. 2, but here the rats warmed their spinal thermode when they pressed the lever (**p<0.01, Student t-test with nearest column).
warm environment; indeed their rectal temperature rose. Self spinal warming was also obtained in one dog [7]. In that case the results were identical to those obtained here in rats: increase in ambient temperature also increased the behavior of self spinal warming. Figure 5b, column C shows that, contrary to what was expected, rats did not self warm their spinal cord in a cold environment. This is paradoxical. Since they did so in a warm environment the cold signal alone is sufficient to arouse the behavior. One would, then, expect them to self warm more in a cold environment when a skin cold signal is added to the spinal cold stimulus. The cooling of the spinal canal was the same in both experiments, therefore the difference originated in the cold skin. Two possible explanations can be hypothetized to explain this behavior: (a) nervous input from a cold skin would inhibit the transmission of the cold spinal signal to the higher centres. This mechanism would also suppress self spinal cooling in a cold environment. (2) a cold skin would arouse a sensation marking the spinal sensation. Thus in a cold environment a spinal cooling would leave the rat unaware of it. Other negative results obtained in this experiment could be expected and serve as control.
E X P E R I M E N T III: C H O I C E
Warm and Neutral Spinal Perfusions Figure 5 shows the mean self spinal warming time o f the rats when ambient temperature was warm 35°C (Fig. 5a), or cool 20°C (Fig. 5b). Columns W and B refer to the water perfusing the spinal thermode, warm (41.4
Among the six experimental conditions combining 3 thermode temperatures and 2 ambient temperatures, only one case gave positive results statistically different from all other cases: rats self warmed their spinal canal in a warm environment when the thermode was cooled. This result shows the existence of a cold sensitivity, probably in the spinal cord. In this regard rats responded in the same way as pigs [31 and pigeons [17] but contrary to dogs [6]. Self warming was opposite to spinal cooling applied by the experimenter. The behavior was present in a warm environment in absence of a cool skin. Cold spinal canal therefore was necessary and sufficient to produce self spinal heating and was thus different from self spinal cooling (Experiment I). The temperature obtained by the rats was usually about 42.5°C at the entry of the thermode. Even if there was some heat loss between the thermocouple and the spinal canal one may assume that thermode temperature was higher than deep body temperature (38.5 to 39°C). The behavior therefore not only removed the cooling but also warmed the spinal canal. This may be the reason for the lever pressing: the rats would thus actuate their autonomic response and in turn better resist the
Experiment I showed that rats suppressed the warming of their spinal canal and self cooled it in a warm environment. Experiment II showed that rats suppressed the cooling of their spinal canal and self warmed it in a warm environment. Thus the rats evidence contradictory behaviors. In order to solve this contradiction we placed the rats in a situation of choice. In this experiment the rats had access simultaneously to two levers. Depressing the first lever warmed the water perfusing the spinal thermode. Depressing the second lever cooled it. When no lever was depressed water at the rat's body temperature circulated in the thermode. Nine rats served in this experiment. Each rat was placed for 90 min sessions at environmental temperatures of 10°C, 20°C and 35°C, 30 min each. In order to cancel out the influence of environmental temperature sequence, in half the sessions the sequence was reversed: 35°C, 20°C and 10°C. RESULTS AND DISCUSSION
In a warm environment, 20 experiments were performed on 9 rats whose spinal temperature was maintained at between 37.6 and 38.8°C. In a cool environment 20 experiments were performed on 9 rats whose spinal temperature was maintained at between 37.7 and 39.6°C. In a cold environment 13 experiments were performed on 8 rats whose spinal temperature was maintained at between 37.6 and 38.8°C. Figure 6 gives an example of a sequence where the experimental sequence started at 35°C Ta and was then cooled to 20 and 10°C. The change in ambient temperature was rapidly obtained by adding ice in the chamber wall. In sessions where Ta was raised equally rapid increase was obtained by adding hot water in the wall of the climatic chamber. Figure 7 shows the mean time spent pressing the lever by the group of rats. This figure shows that the rats bar-pressed a significant length of time in hot environment and little time in cold environment. The rats pressed on both levers, alternatively. Each press lasted 20 sec to 1 min and produced thermode temperature changes extending to 26°C and to
494
DI B. CORMARECHE-LEYD1ER AN D (:ABAN: ,~(' EXPERIMENT III :CHOICE
TIME ON COLD (s) TIME ON WARM (s)
302
90
125
47
TIME (sl 0 O
la=asCi Ia:20~C '~Ta:lo~C
41
,;ot
']9
37
35 .O
33
e"
31
TWO 29
27
25 i
i
20
1
t
40
1
t0
6
1
i
1
8~
TIME (min)
FIG. 6. Sample of thermode temperature recorded under various conditions. Tin: temperature of the water perfusing the spinal thermode. Data correspond to bar-pressing for cold and warm spinal rewards. The graph shows the fall and subsequent rise of Tth measured, but it does not show at what precise moment the rat pressed the warming lever. This is obtained from the counters (see Fig. 1). The spinal thermode was perfused with water at the rat's body temperature.
43°C. The latency for a temperature change after a press was between 10 and 15 sec. When in the hot environment a rat sometimes did not use either lever. If it started to use one, it usually immediately switched to the other, then came back to the first, and so on as long as the environment remained hot. As a result the time spent on both levers was comparable, as seen in Fig. 7. The increase of both responses at higher Ta (Fig. 7) may suggest that bar-pressing could reflect a higher activity at high Ta, the response would thus be non specific. This is unlikely because the time spent bar-pressing seemed to be adjusted not only to ambient temperature, but also to spinal stimulus temperature. Such adjustment is evident in Figs. 3 and 5 (results of Experiments I and II). The result of Experiment III confirms the results of both Experiment I and II. Since the rats bar-pressed alternately on warming and cooling levers in hot environment only, they produced, simultaneously, both the results of Experiment I and Experiment II. This behavior suggests that a conscious signal was perceived by the rat only when it started pressing one lever and was then condemned to run from the one to the other in order to correct the warm and cold spinal signals alternately.
[--'1
LEVER
CHOICE
SPINAL
SELF -HEATING
SPINAL
SELF-COOLING
FIG. 7. Mean results of Experiment 111. The rats could both self warm and self cool the water perfusing their spinal thermode. Columns show the mean time (_+S.E.) spent pressing the levers at three different ambient temperatures (T,). When the rat did not press the lever, water at its own body temperature was perfused in the thermode.
G E N E R A L DISCUSSION When a rat bar-presses to modify its own spinal temperature, three explanations of this behavior are possible: (1) the reward lies in the spinal canal, i.e., there is a sensation aroused from the spinal cord or its vicinity. (2) the reward originates from a modified skin sensation. Cutaneous afferent thermoreceptors travel in the spinal cord, a modified spinal temperature could modify the afferent message. (3) a thermal spinal stimulus arouses themoregulatory (and perhaps non thermoregulatory) autonomic responses, which could be rewarding in themselves or in modifying body temperatures in a direction opposite to the spinal stimulus. Let us look at the results on this basis. Self cooling of the spinal canal by the rats (Experiment I) confirms the existence of a warm sensitivity arousing autonomic and behavioral responses. Figure 3 shows that a warm skin temperature increased the spinal cooling, i.e., spinal and skin temperatures would have an influence of the s a m e n a t u r e . Thus it is not impossible that this behavior conforms to the first of the 3 possibilities listed above. The results of Experiment II show that rats corrected their spinal low temperature only at high environmental temperature, and this confirms preliminary results obtained in one dog [7]. Thus warm skin temperature was necessary, as if it was possible for a cold sensitivity of the spinal cord to exist only when the skin is warm. This would suggest that this sensitivity could conform to the second of the three hypothesis listed above. The main results of Experiment III confirm the results of both Experiment I and Experiment II at the same time. In addition, the fact that the same hot environment was able to arouse both self spinal cooling and self spinal warming simultaneously confirms the hypothesis that the rewards obtained were of different nature.
SPINAL CANAL THERMOREGULATION
495 REFERENCES
1. Adair, E. R. Preference for skin cooling during hypothalamic heating. Proceedings of the International Union of Physiological Sciences, XIII: 8, 1977. 2. Baldwin, B. A. and D. L. Ingram. The effect of heating and cooling the hypothalamus on behavioural thermoregulation in pigs. J. Physiol., Lond. 191: 375-392, 1967. 3. Carlisle, H. J. and D. L. Ingram. The effects of heating and cooling the spinal cord and hypothalamus on thermoregulatory behaviour in the pig. J. Physiol., Lond. 231: 353-364, 1973. 4. Corbit, J. D. Behavioral regulation of hypothalamic temperature. Science 166: 256-257, 1969. 5. Corbit, J. D. and T. Ernits. Specific preference for hypothalamic cooling. J. comp. physiol. Psychol. 86: 24-27, 1974. 6. Cormareche-Leydier, M. and M. Cabanac. Influence de stimulations thermiques de la moelle 6pini~re sur le comportement thermor6gulateur du chien. Pfliigers Arch. 341: 313-324, 1973. 7. Cormareche-Leydier, M. and M. Cabanac. Dog behaviour as related to spinal cord temperature. Experientia 32: 66--67, 1976. 8. Dib, B. Coninuous perfusion under pressure using several tubes in the freely behaving rat. Physiol. Behav. 24: 177-178, 1980. 9. Duclaux, R., M. Fantino and M. Cabanac. Comportement thermor6gulateur chez Rana esculenta. Influence du r6chauffement spinal. Pfliigers Arch. 342: 347-358, 1973. 10. Hales, J. R. S. and C. Jessen. Increase of cutaneous moisture loss caused by local heating of the spinal cord in the ox. J. Physiol. Lond. 204: 40P-42P, 1969.
I 1. Hamilton, C. L. Effect of food deprivation on thermal behavior of the rat. Proc. Soc. exp. Biol. Med. 100: 354-356, 1959. 12. Jessen, C., E. Simon and R. Kullman. Interaction of spinal and hypothalamic thermodetectors in body temperature regulation in the conscious dog. Experientia 24: 694-695, 1968. 13. Jessen, C. Panting in waking dogs by isolated heating of the spinal cord. Naturwissenchaften 54: 290-291, 1967. 14. Kosaka, M., E. Simon and R. Thauer. Shivering in intact and spinal rabbits during spinal cord cooling. Experientia 23: 385387, 1967. 15. Rautenberg, W. Die Bedentung der Zentralnerv6sen Therm6sensitivit~it for die Temperaturregulation der Taube. Z. vergl. Physiol. 62: 235-266, 1969. 16. Schmidt, I. Effect of central thermal stimulation on the thermoregulatory behaviour of the pigeon. Pfliigers Arch. 363: 271272, 1976. 17. Schmidt, I. Behavioral and autonomic thermoregulation in heat stressed pigeons modified by central thermal stimulation. J. comp. Physiol. 127: 75-87, 1978. 18. Simon, E., W. Rautenberg, R. Thauer and M. Iriki. Ausl6sung thermoregulatorischer Reaktionen durch lokale Ktihlung im Vertebralkanal. Naturwissenchaften 50: 337-339, 1963. 19. Simon, E., W. Rantenberg and C. Jessen. Initiations of shivering in unanesthetized dogs by local cooling within the vertebral canal. Experientia 21: 477, 1965.