Dorsal and median raphe serotonergic system lesion does not alter the opiate withdrawal syndrome

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Abstract

Previous pharmacological studies have implicated serotonergic brain systems in opiate withdrawal. To test the hypothesis that serotonin (5-HT) has a critical role in the development of opiate withdrawal, we have employed a near-total brain 5-HT system lesion technique (90% depletion) using 5,7-dihydroxytryptamine combined with induction of opiate dependence by implantation of morphine pellets or by repeated injections of increasing doses of morphine. The effects of serotonergic neuron lesion were examined on spontaneous opiate withdrawal (changes in circadian locomotor activity) and naloxone-precipitated opiate withdrawal syndrome (the somatic aspect). The antiwithdrawal properties of clonidine, an α2-adrenoceptor agonist currently used for clinical treatment for the somatic signs of opiate withdrawal, were tested also in the lesioned rats. Our findings show that serotonergic lesions in morphine-dependent rats did not alter either the spontaneous or the naloxone-induced withdrawal syndrome (with exception of jumping behavior). Moreover, clonidine alleviated the naloxone-induced withdrawal syndrome in lesioned as well as in sham-operated morphine-dependent rats. These results demonstrate that 5-HT systems are not directly responsible for the development of the somatic opiate withdrawal syndrome in morphine-dependent rats.

Introduction

“Opponent Process” theory postulates that dependence to opiates represents a novel homeostatic state resulting from neuroadaptive mechanisms that counteract the effects of opiates in the organism Koob and Bloom, 1988, Solomon, 1980. The acute opiate withdrawal syndrome is assumed to be induced by these neuroadaptations, which no longer are counterbalanced by the presence of the drug (Koob and Bloom, 1988). The aversive component of morphine withdrawal can be evaluated by measuring the intensity of the place aversion conditioning triggered by injections of very low doses of naloxone (Schulteis et al., 1994). The administration of higher doses of naloxone or abstinence from morphine enables study of the somatic component of opiate dependence, which can be measured by rating various somatic and behavioral patterns during the acute abstinence syndrome (Schulteis et al., 1994). The spontaneous opiate abstinence usually is reflected by the disruption of the circadian cycle in rats (Stinus et al., 1998).

An acute injection of morphine increases serotonin (5-HT) release in the rat forebrain (Tao and Auerbach, 1995), which modulates dopaminergic activity within the nucleus accumbens (Spampinato et al., 1984). 5-HT transmission also is involved in the chronic effects of morphine. Increased brain 5-HT release is associated with the development of physical dependence to morphine Javelle et al., 1997, Way et al., 1968. It has been shown that inhibition of 5-HT transmission, by partial lesion with 5,6-dihydroxytryptamine, attenuates morphine dependence (Ho et al., 1972). In this model, higher doses of naloxone were required to trigger the abstinence syndrome. Consistent with these results, Samanin et al. (1980) showed that drugs inhibiting central 5-HT function during the development of opiate dependence decrease the intensity of physical dependence and, more specifically, the jumping behavior displayed during acute opiate withdrawal (Cervo et al., 1983). Although these findings pointed to an involvement of serotonergic activity in opiate dependence, the conclusions were based mainly on the sole rating of jumping behavior. It is recognized nevertheless that the opiate withdrawal syndrome is a complex behavior that includes expression of numerous somatic manifestations, and jumping behavior alone is unlikely to account for the entire opiate-dependent state (Gellert and Holtzman, 1978).

In view of these results, the exact role of 5-HT systems in opiate withdrawal remains unclear. In order to answer this question, we have destroyed 5-HT neurons arising from the dorsal (DR) and median raphe (MR) nuclei by multiple injections of 5,7-dihydroxytryptamine (90% depletion). The effects of the lesion in morphine-dependent rats were examined both on the development of spontaneous opiate withdrawal by continuous evaluation of locomotor activity and the somatic component of opiate withdrawal by quantifying naloxone-induced opiate withdrawal symptoms. Finally, we tested the ability of clonidine to protect 5-HT-lesioned rats from an acute opiate withdrawal syndrome.

Section snippets

Animals

A total of 123 male Sprague–Dawley rats (IFFA-CREDO, Lyon, France) weighing 220–240 g at the beginning of the experiments were used. Animals were housed by four in cages located in a thermoregulated room (22 °C) with a 12:12 h light–dark cycle (light from 8:00 a.m. to 8:00 p.m.). Food and water were available ad libitum. These conditions were maintained constant throughout the experiments. All manipulations and observations were made during the light phase of the circadian cycle.

The experiments

Biochemical verification of 5-HT lesion

Results are presented in Table 1. Bilateral multiple injections of 5,7-DHT into the DR and MR nuclei and in the vicinity of the interpeduncular nucleus, where all the ascending serotonergic ascending fibers are localized, produced an almost total lesion of ascending serotonergic innervation. Almost complete depletion of 5-HT was observed in the cortex (−88%, P<.001), in the hippocampus (−86%, P<.001) and in the striatum (−99.6%, P<.001). NA was not affected in either hippocampus (+1%, NS) or

Discussion

In the present study, we tested the hypothesis that brain serotonergic systems are critical for manifestations of spontaneous and precipitated opiate withdrawal. Multiple intracerebral injections of 5,7-DHT induced a reliable specific and nearly total (90%) depletion of ascending 5-HT forebrain innervation.

The present results have shown that spontaneous opiate withdrawal, as measured by disruption of circadian locomotor activity observed during the first days of opiate abstinence, was not

Acknowledgements

This study was supported by grants from the University of Bordeaux 2, the Centre National de la Recherche Scientifique (CNRS), the Conseil Régional d'Aquitaine, the MILDT (no. 96C02), the Spanish–French Integrated Action (HF-97-220 and 97-160) and National Institutes of Health (USA) (DA04043 from the National Institute on Drug Abuse). This is publication number 14703-NP from The Scripps Research Institute. The authors would like to thank Mike Arends for his editorial assistance.

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