Br. J. Pharmacol. (1992), 107, 15-21 '." Macmillan Press Ltd, 1992 Characterization of 8-OH-DPAT-induced hypothermia in mice as a 5-HTlA autoreceptor response and its evaluation as a model to selectively identify antidepressants 'Keith F. Martin, Ian Phillips, Mitchell Hearson, Michael R. Prow & David J. Heal Boots Pharmaceuticals Research Department, Nottingham NG2 3AA 1 8-Hydroxy-2-(di-n-propylamino)tetralin (8-OH-DPAT) dose-dependently induced hypothermia in mice. 2 The 5-HTIA receptor partial agonists, buspirone, gepirone and ipsapirone, also dose-dependently induced hypothermia. 3 The 8-OH-DPAT temperature response was antagonized by the 5-HT, receptor antagonists quipazine (2mg kg', i.p.), (±)-propranolol (10mgkg-', i.p.). (±)-pindolol (5mgkg-', i.p.), spiroxa- trine (0.5 mg kg-', i.p.) and metitepine (0.05 mg kg-', i.p.), but not by 5-HT2 (ketanserin) or 5-HT3 (MDL 72222, GR 38032F) receptor antagonists. 4 The response was also antagonized by the dopamine D2 receptor antagonists, haloperidol and BRL 34778. No other catecholamine or muscarinic receptors were involved in mediating the response. 5 Destruction of 5-hydroxytryptamine (5-HT)-containing neurones with the neurotoxin, 5,7- dihydroxytryptamine (75 bg, i.c.v.), abolished the response to 8-OH-DPAT indicating that the 5-HTIA receptors involved were located on 5-HT neurones. 6 Chronic antidepressant treatment down-regulated this 8-OH-DPAT response. In addition, chronic administration of anxiolytics and neuroleptics was also effective in this respect. Down-regulation was also observed following repeated administration of 8-OH-DPAT (0.5 mg kg-', s.c.), (± )-pindolol (10mg kg-', i.p.) and ketanserin (0.5 mg kg-', i.p.). 7 In conclusion, these data confirm that 8-OH-DPAT-induced hypothermia is mediated by 5-HTIA autoreceptors. They also indicate that the response involves D2 receptors. The present study also shows that a wide range of antidepressant drugs down-regulate this response although this property is not restricted to antidepressant treatments. Therefore, care should be exercised when interpreting data from this paradigm. Keywords: 8-Hydroxy-2-(di-n-propylamino)tetralin (8-OH-DPAT); hypothermia; mice; antidepressants; 5-HTIA receptors Introduction In 1983, Middlemiss & Fozard showed that 8-hydroxy-2-(di- Goodwin et al. (1985b) have reported that 8-OH-DPAT- n-propylamino)tetralin (8-OH-DPAT) was a selective agonist induced hypothermia in mice was selectively attenuated after at 5-HTIA receptors. Subsequently, it was found that repeated administration of antidepressant drugs and electro- peripheral administration of 8-OH-DPAT induced hypother- convulsive shock (ECS). However, as the types of antidepres- mia in mice and rats (Hjorth, 1985; Goodwin & Green, 1985; sant tested were limited and the authors failed to investigate Goodwin et al., 1985a; Gudelsky et al., 1986; Higgins et al., the effects of other psychotropic agents, their conclusion that 1988; Wozniack et al., 1988). The studies in mice have shown this adaptation was specific to antidepressants required fur- that this response is dose-dependent and sigmoidal (e.g. ther confirmation. Goodwin et al., 1985a). Pharmacological evaluation of the The aims of the work described here were two fold: first, to hypothermic response to 8-OH-DPAT led these authors to extend the pharmacological characterization of the 8-OH- suggest that it was specifically mediated by 5-HTIA receptors DPAT-induced hypothermia in mice and second, to deter- located on 5-hydroxytryptaminergic neurones. Goodwin and mine whether the adaptive down-regulation of this response his coworkers based this hypothesis on inhibition of the was specific to antidepressant treatment. response by ipsapirone (Goodwin et al., 1986), a partial agonist at 5-HTIA receptors (Martin & Mason, 1986), low doses of quipazine (Goodwin et al., 1985a) and abolition of Methods the response by destruction of 5-HT neurones (Goodwin et al., 1985a). However, these authors were unable to prevent Animals the effects of 8-OH-DPAT with a range of other 5-HTI-like receptor antagonists. Furthermore, haloperidol, a potent D2 Adult male C57/Bl/601a mice (Olac, Bicester) weighing dopamine receptor antagonist, was also found to inhibit this 25-30 g were used throughout this study. They were housed response (Goodwin et al., 1985a). The data suggest, there- in groups of 10 on a 12:12 h light:dark cycle (lights on fore, that the pharmacology of 8-OH-DPAT-induced hypo- 07 h 00 min) and had free access to food and water. The thermia in mice may not be as straight-forward as first ambient temperature was maintained at 20± 10C and thought. humidity was approximately 50%. Agonist studies Core temperature was measured by inserting the probe of a Author for correspondence. digital thermometer approximately 2.5 cm into the rectum 16 K.F. MARTIN et al. while lightly restraining the animal 10 min prior to (t- o), All drugs were dissolved in 0.9% saline and injected intra- immediately before (t0) and 20 min after (t20) an s.c. injection peritoneally (i.p.) with the exception of 8-OH-DPAT, buspirone, of saline or 5-HTlA receptor agonist and the difference in gepirone and ipsapirone which were given subcutaneously (s.c.). temperature between t0 and t20 was calculated. Antagonist studies Results Animals received either saline (10mgkg-', s.c.) or 8-OH- Effects ofS-HTIA receptor agonists on rectal temperature DPAT (0.5 mg kg-', s.c.) 30 min after an i.p. injection of saline or antagonist. Rectal temperature was measured When mice were injected with increasing doses of 8-OH- immediately before the first injection (t 30) and prior to the DPAT (0.05 to 5.0 mg kg-', s.c.), this 5-HTIA receptor second injection of saline or 8-OH-DPAT (to). A further agonist produced a dose-dependent decrease in temperature measurement was taken 20 min after the second injection (Figure 1). Time-course studies showed that the maximum (t20). The difference in temperature between to and t20 was hypothermic effect was observed 20 min after 8-OH-DPAT calculated. (data not shown). Similarly, the 5-HTIA receptor partial agonists, buspirone, gepirone and ipsapirone, also dose- Effect of chonic drug treatments dependently induced hypothermia in mice (Figure 1). Groups of mice received either 14 daily injections (i.p.) of Effects of5-HT receptor antagonists drugs or saline. The temperature decrease following an injec- tion of 8-OH-DPAT (0.5 mg kg-', s.c.) was determined 24 h The putative 5-HTl/5-HTIA receptor antagonists quipazine after the first and last treatments. (2mg kg-'), (±)-propranolol (10 and 20mg kg-'), (+)- pindolol (5 and 10 mg kg-'), spiroxatrine (0.5 and 5 mg kg-') 5-Hydroxytryptamine-containing neurone lesioning studies and metitepine (0.05 mg kg-') significantly attenuated the hypothermic response to 8-OH-DPAT (Table 1). Only quipa- The 5-HT neurotoxin, 5,7-dihydroxytryptamine (5,7-DHT; zine had any significant effect (a small increase cf. control) 75 pg) or saline-ascorbate (4 fl) was injected i.c.v. into when given alone (Table 1). Metitepine antagonized the re- halothane anaesthetized mice, pretreated with desipramine sponse at a dose of 0.05 mg kg-', which itself had no effect on (5 mg kg-', i.p.) to protect noradrenergic neurones, by the core temperature. However at a dose of 0.5 mg kg-', method of Heal (1984). The temperature response to 8-OH- metitepine caused a marked reduction in temperature DPAT (0.5 mg kg-1, i.p.) was measured seven days later as (3.85 ± 0.79°C; P <0.001). There was no further fall in described above. temperature induced by 8-OH-DPAT. The non-selective 5-HT receptor antagonist, metergoline had similar effects (Table 1). Noradrenaline-containing neurone lesioning studies Ketanserin (0.2 mg kg-'), a 5-HT2/5-HT,c receptor antagonist, did not significantly alter body temperature itself, Mice were given an i.p. injection of N-(2-chloroethyl)-N- nor did it affect the hypothermia induced by 8-OH-DPAT ethyl-2-bromobenzylamine (DSP-4, 100 mg kg-') 30 min (Table 1). after zimeldine (5 mg kg-', i.p.). Seven days later this proce- At moderate doses, the 5-HT3 receptor antagonists, MDL dure was repeated. The temperature response to 8-OH-DPAT 72222 and GR 38032F, did not alter the temperature re- (0.5 mg kg-', i.p.) was measured 12 days after the first DSP-4 sponse to 8-OH-DPAT (Table 1). injection as described above. Effects of catecholamine and muscarinic receptor Statistics antagonists Data were analysed by Student's unpaired t test and the null The ax-adrenoceptor antagonist, prazosin (0.1 mg kg-') did hypothesis was rejected when P <0.05. However, when two not attenuate the response to 8-OH-DPAT (0.5 mg kg-') treatment groups were compared with a single control group, (Table 2). However, at high dose (1 mg kg-'), it induced a the level of significance for rejection of the null hypothesis marked fall in core temperature itself; this was not altered by was set at P<0.01. 8-OH-DPAT (Table 2). The M2-adrenoceptor antagonist, Drugs 3- The following drugs were used (abbreviation if any and source shown in parentheses): N-(2-chloroethyl)-N-ethyl-2- co bromobenzylamine (DSP-4), 8-hydroxy-2(di-n-propyl- amino)tetralin HBr (8-OH-DPAT); ondansetron (GR 38032F), laH,3ax,5aH-tropan-3-yl-3,5-dichlorobenzoate (MDL 72222), (+ )-7-chloro-8-hydroxy-3-methyl-l-phenyl-2,3,4,5- tetrahydro-lH-3-benzazepine maleate (SCH 23390), spiroxat- 0 rine (Research Biochemicals Inc.), dothiepin HCl2, gepirone (HCl)2, ipsapirone (HCI)2, sibutramine HCI (Boots Phar- maceuticals), zimeldine (HCl)2 (Astra), exo-N-(9-[(4- fluorophenyl)methyl)-9-azabicyclo [3.3.1]non-3-yl)-4-amino-5- chloro-2-methoxybenzamide (BRL 34778) (Beecham), map- rotiline (Ciba-Geigy), metitepine, metergoline (Farmitalia), 0.05 0.5 5 0.1 1 10 0.1 1 10 0.1 1 10 erythro-( ± )-I-(7-methylindan-4-yloxy)-3-isopropylaminobutan 8-OH-DPAT Buspirone Gepirone.