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Immobilisation stress-induced antinociception in rats: Possible role of serotonin and prostaglandins
83
European Journal of Pharmacology, 50 (1978) 83--85 © Elsevier/North-Holland Biomedical Press
Short communication IMMOBILISATION STRESS-INDUCED ANTINOCICEPTION IN RATS: POSSIBLE ROLE O F S E R O T O N I N AND P R O S T A G L A N D I N S SALIL K. BHATTACHARYA, PRAKASH R. KESHARY and AJIT K. SANYAL *
Department of Pharmacology, Institute of Medical Sciences, Banaras Hindu University, Varanasi-221005, India Received 13 April 1978, accepted 24 April 1978
S.K. BHATTACHARYA, P.R. KESHARY and A.K. SANYAL, Immobilisation stress-induced antinociception in rats: possible role of serotonin and prostaglandins, European J. Pharmacol. 50 (1978) 83--85. Immobilisation of rats for 1, 2 and 4 h induced duration-related antinociception. Antinociception induced by 4 h immobilisation was significantly inhibited after pretreatment by drugs known to inhibit synthesis of serotonin, induce degeneration of serotonergic neurones and inhibit prostaglandin synthesis. The results indicate that immobilisation stress induces autoanalgesia, which may be dependent on the availability of endogenous serotonin and prostaglandins in rat brain. Immobilisation
Antinociception
Serotonin
1. Introduction Recent research indicates that stress-induced hyperemotionality effectively elicits antinociception in the rat. Thus, acute or intermittent repeated electroshock increased in the pain threshold associated with enhanced endogenous opioid peptide receptor occupation in the rat brain. This behaviourally activated antinociception has been termed autoanalgesia (Chance et al., 1977, and references cited therein). In this communication we provide the first report of immobilisation stressinduced antinociception in the rat, together with preliminary data on the possible mechanisms involved.
2. Materials and methods Wistar strain albino rats {100--150 g) of both sexes were used. F o o d was withheld 18 h before and water immediately before immobilisation. Antinociceptive activity was deter* Correspondence to A.K.S. at the above address.
Prostaglandins
mined by the radiant heat rat tail-hot wire technique {Davies et al., 1946) using a Techno analgesiometer. The increases in the latent period of the tail flick response, with a cut-off time of 30 sec, between the pre- and postimmobilisation periods were measured; statistical analysis was done b y means of Student's t-test. The rats were immobilised by tying the fore and hind limbs separately and then together, and finally securing them by means of adhesive tape. The rats were p u t inside individual adjustable, cylindrical metallic restraint chambers and finally the tail was taped so as to make the animal completely immobile. The experiments were conducted at ambient temperature of 25 + I°C and between 9.00 am to 6.00 pm. The drugs used in the study, with dose and pretreatment time in parentheses were: p-chlorophenylalanine methyl ester hydrochloride (100 mg/kg, i.p., once daily for 3 days, the last injection being given 24 h before experimentation), 5,6-dihydroxytryptamine creatinine phosphate (75 pg/ rat, intraventricularly, 48 h), diclofenac sodium (10 mg/kg, i.p., 4 h) and indomethacin (15 mg/kg, i.p., 4 h ) . N o n e of these drugs had
84
S.K. BHATTACHARYA ET AL.
TABLE 1 Antinociceptive effect of immobilisation stress (1, 2 and 4 h) and the effect of PCPA, DHT, diclofenac and indomethcin on 4 h immobilisation-induced antinociception. Groups
Unrestrained control Immobilisation (1 h) Immobilisation (2 h) Immobilisation (4 h) PCPA + immobilisation (4 h) DHT + immobilisation (4 h) Diclofenac + immobilisation (4 h) Indomethacin + immobilisation (4 h)
n
10 5 5 15 5 5 5 5
Increase in latent period of tail flick response (sec) Mean
S.E.M.
0.9 1.5 5.3 9.9 3.2 1.5 2.5 3.6
0.12 0.34 0.44 1.50 0.99 1.15 1.10 0.92
P
N.S. 1 N.S. 1 <0.001 1 <0.001 <0.001 2 <0.001 2 <0.001 2 <0.001 2
1 Indicates statistical significance between pre- and post-immobilisation data, except in unrestrained control where it indicates significance between first and second (after 4 h) data. 2 Indicates statistical significance in relation to the 4 h immobilisation group. Statistical significance was calculated by the paired t-test and unpaired t-test, respectively.
a n y significant a n t i n o c i c e p t i v e effect, p e r se, in t h e doses used. A c o n t r o l g r o u p was i n c l u d e d , w i t h u n - r e s t r a i n e d rats, t o n o t e t h e effect of repeated testing for antinociceptive a c t i v i t y , if a n y , on t h e l a t e n t p e r i o d o f tail flick r e s p o n s e .
3. Results I m m o b i l i s a t i o n f o r 1 h, 2 h and 4 h prod u c e d d u r a t i o n - r e l a t e d increasing a n t i n o c i c e p t i o n . T h e a n t i n o c i c e p t i o n i n d u c e d b y 4 h imm o b i l i s a t i o n was significantly i n h i b i t e d a f t e r pretreatment with p-chlorophenylalanine (PCPA), 5,6-dihydroxytryptamine (DHT), d i c l o f e n a c or i n d o m e t h a c i n , t o t h e e x t e n t o f 67.7%, 84.9%, 74.7% a n d 63.6%, r e s p e c t i v e l y ( t a b l e 1).
4. Discussion T h e results indicate t h a t i m m o b i l i s a t i o n i n d u c e d significant a n t i n o c i c e p t i o n , t h e intensity o f w h i c h was r e l a t e d t o t h e d u r a t i o n o f restraint. T h e role o f i m m o b i l i s a t i o n as a
stressor is well d o c u m e n t e d (Selye, 1976). The antinociception induced by 4 h immobilisation was i n h i b i t e d b y PCPA, a specific serotonin biosynthesis inhibitor (Koe and Weissman, 1 9 6 6 ) a n d b y D H T , w h i c h induces selective d e g e n e r a t i o n o f s e r o t o n e r g i c n e u r o n e s { B a u m g a r t e n et al., 1971), as well as b y t h e p r o s t a g l a n d i n ( P G ) s y n t h e s i s inhibitors, diclof e n a c and i n d o m e t h a c i n ( K u et al., 1975). I t t h u s a p p e a r s t h a t b o t h t h e availability o f e n d o g e n o u s s e r o t o n i n and t h e p r o s t a g l a n d i n s are prerequisites f o r t h e p r o d u c t i o n o f i m m o bilisation-induced antinociception. Central pain i n h i b i t o r y m e c h a n i s m s in t h e rat are k n o w n t o be s e r o t o n i n - m e d i a t e d . I n h i b i t i o n o f s e r o t o n i n b i o s y n t h e s i s or lesions in t h e m e d i a n f o r e b r a i n b u n d l e b o t h l o w e r the p a i n t h r e s h o l d , an e f f e c t w h i c h is associa t e d w i t h d e p l e t e d brain s e r o t o n i n levels. B o t h p a i n t h r e s h o l d a n d s e r o t o n i n levels were restored to normal by the serotonin precursor, 5 - h y d r o x y t r y p t o p h a n ( G r e e n and G r a h a m e S m i t h , 1975). M o r p h i n e a n t i n o c i c e p t i o n , in t h e rat, is also c o n s i d e r e d t o be s e r o t o n i n m e d i a t e d ( G r e e n and G r a h a m e - S m i t h , 1975). T h e r e are a n u m b e r o f r e p o r t s suggesting t h a t stress, including immobilisation stress,
IMMOBILISATION STRESS-INDUCED ANTINOCICEPTION
enhances rat brain serotonin turnover, though views to the contrary are also available (Green and Grahame-Smith, 1975). In recent communications from this laboratory, we have suggested that morphine first stimulates the biosynthesis of PGs (possibly PGE1), which in turn enhances serotonin turnover in rat brain to mediate morphine antinociception. We have also reported a serotoninmediated antinociceptive effect of PGE1 in this species (Sanyal et al., 1977; Debnath et al., 1978; Srivastava et al., 1978). It may be possible that a similar PG-serotonin interaction-mediated mechanism is responsible for the antinociception induced b y immobilisation stress. These suggestions require further study which is now in progress.
References Baumgarten, H.G., A.L. Bjorklund, L. Lachenmayer, A. Novin and U. Stenevi, 1971, Long lasting selective depletion of brain serotonin by 5,6-dihydroxytryptamine, Acta Physiol. Scand. Suppl. 373, 1. Chance, W.T., A.C. White, G.M. Krynock and J.A. Rosecrans, 1977, Autoanalgesia. Behaviorally activated antinociception, European J. Pharmacol. 44, 283.
85
Davies, O.L., J. Raventos and A.L. Walpole, 1946, A method for evaluation of analgesic activity using rats, Brit. J. Pharmacol. 1,255. Debnath, P.K., S.K. Bhattacharya, A.K. Sanyal, M.K. Poddar and J.J. Ghosh, 1978, Prostaglandins: Effect of prostaglandin E I on brain, stomach and intestinal serotonin in rat, Biochem. Pharmacol. 27,130. Green, A.R. and D.G. Grahame-Smith, 1975, 5-Hydroxytryptamine and other indoles in the central nervous system, in: Handbook of Psychopharmacology, Vol. 3, Biochemistry of Biogenic Amines, eds. L.L. Iversen, S.D. Iversen and S.H. Snyder (Plenum Press, New York) p. 169. Koe, B.K. and A. Weissman, 1966, p-Chlorophenylalanine, a specific depletor of brain serotonin, J. Pharmacol. Exptl. Therap. 154,499. Ku, E.C., J.M. Wasvary and W.D. Cash, 1975, Diclofenac sodium (G.P. 45840, Voltaran), a potent inhibitor of prostaglandin synthetase, Biochem. Pharmacol. 26, 641. Sanyal, A.K., S.K. Bhattacharya, P.R. Keshary, D.N. Srivastava and P.K. Debnath, 1977, Prostaglandins: Antinociceptive effect of prostaglandin E 1 in the rat, Clin. Exptl. Pharmacol. Physiol. 4, 247. Selye, H., 1976, Stress in Health and Disease (Butterworths, London) p. 184. Srivastava, D.N., S.K. Bhattacharya and A.K. Sanyal, 1978, Effect of some prostaglandin synthesis inhibitors on the antinociceptive action of morphine, Clin. Exptl. Pharmacol. Physiol. (in press).
European Journal of Pharmacology, 50 (1978) 83--85 © Elsevier/North-Holland Biomedical Press
Short communication IMMOBILISATION STRESS-INDUCED ANTINOCICEPTION IN RATS: POSSIBLE ROLE O F S E R O T O N I N AND P R O S T A G L A N D I N S SALIL K. BHATTACHARYA, PRAKASH R. KESHARY and AJIT K. SANYAL *
Department of Pharmacology, Institute of Medical Sciences, Banaras Hindu University, Varanasi-221005, India Received 13 April 1978, accepted 24 April 1978
S.K. BHATTACHARYA, P.R. KESHARY and A.K. SANYAL, Immobilisation stress-induced antinociception in rats: possible role of serotonin and prostaglandins, European J. Pharmacol. 50 (1978) 83--85. Immobilisation of rats for 1, 2 and 4 h induced duration-related antinociception. Antinociception induced by 4 h immobilisation was significantly inhibited after pretreatment by drugs known to inhibit synthesis of serotonin, induce degeneration of serotonergic neurones and inhibit prostaglandin synthesis. The results indicate that immobilisation stress induces autoanalgesia, which may be dependent on the availability of endogenous serotonin and prostaglandins in rat brain. Immobilisation
Antinociception
Serotonin
1. Introduction Recent research indicates that stress-induced hyperemotionality effectively elicits antinociception in the rat. Thus, acute or intermittent repeated electroshock increased in the pain threshold associated with enhanced endogenous opioid peptide receptor occupation in the rat brain. This behaviourally activated antinociception has been termed autoanalgesia (Chance et al., 1977, and references cited therein). In this communication we provide the first report of immobilisation stressinduced antinociception in the rat, together with preliminary data on the possible mechanisms involved.
2. Materials and methods Wistar strain albino rats {100--150 g) of both sexes were used. F o o d was withheld 18 h before and water immediately before immobilisation. Antinociceptive activity was deter* Correspondence to A.K.S. at the above address.
Prostaglandins
mined by the radiant heat rat tail-hot wire technique {Davies et al., 1946) using a Techno analgesiometer. The increases in the latent period of the tail flick response, with a cut-off time of 30 sec, between the pre- and postimmobilisation periods were measured; statistical analysis was done b y means of Student's t-test. The rats were immobilised by tying the fore and hind limbs separately and then together, and finally securing them by means of adhesive tape. The rats were p u t inside individual adjustable, cylindrical metallic restraint chambers and finally the tail was taped so as to make the animal completely immobile. The experiments were conducted at ambient temperature of 25 + I°C and between 9.00 am to 6.00 pm. The drugs used in the study, with dose and pretreatment time in parentheses were: p-chlorophenylalanine methyl ester hydrochloride (100 mg/kg, i.p., once daily for 3 days, the last injection being given 24 h before experimentation), 5,6-dihydroxytryptamine creatinine phosphate (75 pg/ rat, intraventricularly, 48 h), diclofenac sodium (10 mg/kg, i.p., 4 h) and indomethacin (15 mg/kg, i.p., 4 h ) . N o n e of these drugs had
84
S.K. BHATTACHARYA ET AL.
TABLE 1 Antinociceptive effect of immobilisation stress (1, 2 and 4 h) and the effect of PCPA, DHT, diclofenac and indomethcin on 4 h immobilisation-induced antinociception. Groups
Unrestrained control Immobilisation (1 h) Immobilisation (2 h) Immobilisation (4 h) PCPA + immobilisation (4 h) DHT + immobilisation (4 h) Diclofenac + immobilisation (4 h) Indomethacin + immobilisation (4 h)
n
10 5 5 15 5 5 5 5
Increase in latent period of tail flick response (sec) Mean
S.E.M.
0.9 1.5 5.3 9.9 3.2 1.5 2.5 3.6
0.12 0.34 0.44 1.50 0.99 1.15 1.10 0.92
P
N.S. 1 N.S. 1 <0.001 1 <0.001 <0.001 2 <0.001 2 <0.001 2 <0.001 2
1 Indicates statistical significance between pre- and post-immobilisation data, except in unrestrained control where it indicates significance between first and second (after 4 h) data. 2 Indicates statistical significance in relation to the 4 h immobilisation group. Statistical significance was calculated by the paired t-test and unpaired t-test, respectively.
a n y significant a n t i n o c i c e p t i v e effect, p e r se, in t h e doses used. A c o n t r o l g r o u p was i n c l u d e d , w i t h u n - r e s t r a i n e d rats, t o n o t e t h e effect of repeated testing for antinociceptive a c t i v i t y , if a n y , on t h e l a t e n t p e r i o d o f tail flick r e s p o n s e .
3. Results I m m o b i l i s a t i o n f o r 1 h, 2 h and 4 h prod u c e d d u r a t i o n - r e l a t e d increasing a n t i n o c i c e p t i o n . T h e a n t i n o c i c e p t i o n i n d u c e d b y 4 h imm o b i l i s a t i o n was significantly i n h i b i t e d a f t e r pretreatment with p-chlorophenylalanine (PCPA), 5,6-dihydroxytryptamine (DHT), d i c l o f e n a c or i n d o m e t h a c i n , t o t h e e x t e n t o f 67.7%, 84.9%, 74.7% a n d 63.6%, r e s p e c t i v e l y ( t a b l e 1).
4. Discussion T h e results indicate t h a t i m m o b i l i s a t i o n i n d u c e d significant a n t i n o c i c e p t i o n , t h e intensity o f w h i c h was r e l a t e d t o t h e d u r a t i o n o f restraint. T h e role o f i m m o b i l i s a t i o n as a
stressor is well d o c u m e n t e d (Selye, 1976). The antinociception induced by 4 h immobilisation was i n h i b i t e d b y PCPA, a specific serotonin biosynthesis inhibitor (Koe and Weissman, 1 9 6 6 ) a n d b y D H T , w h i c h induces selective d e g e n e r a t i o n o f s e r o t o n e r g i c n e u r o n e s { B a u m g a r t e n et al., 1971), as well as b y t h e p r o s t a g l a n d i n ( P G ) s y n t h e s i s inhibitors, diclof e n a c and i n d o m e t h a c i n ( K u et al., 1975). I t t h u s a p p e a r s t h a t b o t h t h e availability o f e n d o g e n o u s s e r o t o n i n and t h e p r o s t a g l a n d i n s are prerequisites f o r t h e p r o d u c t i o n o f i m m o bilisation-induced antinociception. Central pain i n h i b i t o r y m e c h a n i s m s in t h e rat are k n o w n t o be s e r o t o n i n - m e d i a t e d . I n h i b i t i o n o f s e r o t o n i n b i o s y n t h e s i s or lesions in t h e m e d i a n f o r e b r a i n b u n d l e b o t h l o w e r the p a i n t h r e s h o l d , an e f f e c t w h i c h is associa t e d w i t h d e p l e t e d brain s e r o t o n i n levels. B o t h p a i n t h r e s h o l d a n d s e r o t o n i n levels were restored to normal by the serotonin precursor, 5 - h y d r o x y t r y p t o p h a n ( G r e e n and G r a h a m e S m i t h , 1975). M o r p h i n e a n t i n o c i c e p t i o n , in t h e rat, is also c o n s i d e r e d t o be s e r o t o n i n m e d i a t e d ( G r e e n and G r a h a m e - S m i t h , 1975). T h e r e are a n u m b e r o f r e p o r t s suggesting t h a t stress, including immobilisation stress,
IMMOBILISATION STRESS-INDUCED ANTINOCICEPTION
enhances rat brain serotonin turnover, though views to the contrary are also available (Green and Grahame-Smith, 1975). In recent communications from this laboratory, we have suggested that morphine first stimulates the biosynthesis of PGs (possibly PGE1), which in turn enhances serotonin turnover in rat brain to mediate morphine antinociception. We have also reported a serotoninmediated antinociceptive effect of PGE1 in this species (Sanyal et al., 1977; Debnath et al., 1978; Srivastava et al., 1978). It may be possible that a similar PG-serotonin interaction-mediated mechanism is responsible for the antinociception induced b y immobilisation stress. These suggestions require further study which is now in progress.
References Baumgarten, H.G., A.L. Bjorklund, L. Lachenmayer, A. Novin and U. Stenevi, 1971, Long lasting selective depletion of brain serotonin by 5,6-dihydroxytryptamine, Acta Physiol. Scand. Suppl. 373, 1. Chance, W.T., A.C. White, G.M. Krynock and J.A. Rosecrans, 1977, Autoanalgesia. Behaviorally activated antinociception, European J. Pharmacol. 44, 283.
85
Davies, O.L., J. Raventos and A.L. Walpole, 1946, A method for evaluation of analgesic activity using rats, Brit. J. Pharmacol. 1,255. Debnath, P.K., S.K. Bhattacharya, A.K. Sanyal, M.K. Poddar and J.J. Ghosh, 1978, Prostaglandins: Effect of prostaglandin E I on brain, stomach and intestinal serotonin in rat, Biochem. Pharmacol. 27,130. Green, A.R. and D.G. Grahame-Smith, 1975, 5-Hydroxytryptamine and other indoles in the central nervous system, in: Handbook of Psychopharmacology, Vol. 3, Biochemistry of Biogenic Amines, eds. L.L. Iversen, S.D. Iversen and S.H. Snyder (Plenum Press, New York) p. 169. Koe, B.K. and A. Weissman, 1966, p-Chlorophenylalanine, a specific depletor of brain serotonin, J. Pharmacol. Exptl. Therap. 154,499. Ku, E.C., J.M. Wasvary and W.D. Cash, 1975, Diclofenac sodium (G.P. 45840, Voltaran), a potent inhibitor of prostaglandin synthetase, Biochem. Pharmacol. 26, 641. Sanyal, A.K., S.K. Bhattacharya, P.R. Keshary, D.N. Srivastava and P.K. Debnath, 1977, Prostaglandins: Antinociceptive effect of prostaglandin E 1 in the rat, Clin. Exptl. Pharmacol. Physiol. 4, 247. Selye, H., 1976, Stress in Health and Disease (Butterworths, London) p. 184. Srivastava, D.N., S.K. Bhattacharya and A.K. Sanyal, 1978, Effect of some prostaglandin synthesis inhibitors on the antinociceptive action of morphine, Clin. Exptl. Pharmacol. Physiol. (in press).