J Physiol Sci (2012) 62:317–323 DOI 10.1007/s12576-012-0207-x ORIGINAL PAPER The effects of serotonin/norepinephrine reuptake inhibitors and serotonin receptor agonist on morphine analgesia and tolerance in rats Ercan Ozdemir • Sinan Gursoy • Ihsan Bagcivan Received: 22 July 2011 / Accepted: 28 March 2012 / Published online: 29 April 2012 Ó The Physiological Society of Japan and Springer 2012 Abstract Several studies have demonstrated that seroto- morphine tolerance. In conclusion, we determined that co- nergic and noradrenergic systems have important roles in administration of morphine with amitriptyline and venla- morphine analgesia and tolerance. However, the exact faxine increased the analgesic effects of morphine and mechanism underlying the development of morphine tol- attenuated the morphine analgesic tolerance. erance is not fully understood. The aim of this study was to investigate the possible role of serotonin/norepinephrine Keywords Amitriptyline Á Dihydroergotamine Á reuptake inhibitors (amitriptyline, venlafaxine) and sero- Morphine Á Morphine tolerance Á Venlafaxine tonin receptor (5-HT1A and 5-HT1B/1D) agonist (dihydro- ergotamine) in morphine analgesia and tolerance in rats. To constitute morphine tolerance, animals received morphine Introduction (50 mg/kg; s.c.) once daily for 3 days. After the last dose of morphine was injected on day 4, morphine tolerance was Opioid analgesics are commonly used to treat acute and evaluated. The analgesic effects of amitriptyline (20 mg/ chronic pain associated with surgical interventions or dis- kg; i.p.), venlafaxine (20 mg/kg; s.c.), dihydroergotamine ease states such as cancer. Opioids produce a diverse (100 lg/kg; i.v.) and morphine (5 mg/kg) were considered spectrum of centrally- and peripherally-mediated respon- at 15- to 30-min intervals (0, 15, 30, 60, 90, and 120 min) ses, including respiratory depression, sedation, nausea, by tail-flick and hot-plate analgesia tests. In this study, the euphoria, and itching [1, 2]. Longterm use of opioids can data obtained suggested that amitriptyline and venlafaxine be problematic due to the rapid development of profound significantly increased the analgesic effect of morphine and tolerance to the analgesic effects coupled with slow attenuated the expression of morphine tolerance. However, development of tolerance to many of the untoward effects dihydroergotamine significantly increased the analgesic of these agents. It is the inability to tolerate these unde- effect of morphine but did not reduce the expression of sirable side effects that eventually limits dose escalations and analgesic efficacy [2]. Morphine tolerance is a complex phenomena, involving & E. Ozdemir ( ) the serotonergic and noradrenergic systems [3, 4]. We have Department of Physiology, Cumhuriyet University School of Medicine, 58140 Sivas, Turkey recently demonstrated that the nitric oxide–cGMP signal e-mail: [email protected]; pathway plays a pivotal role in developing tolerance to the [email protected] analgesic effect of morphine [5]. The exact mechanisms underlying the development of morphine tolerance remain S. Gursoy Department of Anesthesiology and Reanimation, controversial. Among them, serotonin (5-HT) is particularly Cumhuriyet University School of Medicine, Sivas, well studied, and this transmitter has been shown to widely Turkey participate in pain pathways and to play a major role in mediating the analgesic action of morphine [6, 7]. It is well I. Bagcivan Department of Pharmacology, Cumhuriyet University School accepted that opioids establish part of their analgesic effect of Medicine, 58140 Sivas, Turkey through stimulation of the serotonergic system [8]. It has been 123 318 J Physiol Sci (2012) 62:317–323 reported that acute morphine administration enhances sero- blindly between 0900 and 1700 hours. Procedures and ani- tonin turnover as evidenced by an increase in its synthesis, mal handling met the guidelines of the National Institutes of release, and metabolism [8], in particular in projection areas of Health detailed in the ‘‘Principles of animal laboratory care’’ the dorsal raphe nucleus and median raphe nucleus [9], but (National Institutes of Health, 1996). The experimental upon chronic morphine administration, a decrease in the protocols were approved by the Cumhuriyet University release of 5-HT from the nerve terminals is observed [10]. Animal Ethics Committee (licence number: 87/Ethic). There have been controversial results about the role of 5-HT in the development of morphine tolerance. For example, Drugs serotonin activity evoked by systemic administration of L-tryptophan or 5-hydroxytryptophan was found to, respec- Amitriptyline, venlafaxine and dihydroergotamine (Sigma- tively, accelerate and attenuate morphine tolerance [11]. Aldrich, USA) and morphine sulphate (Cumhuriyet Uni- Tricyclic antidepressants (TCAs) have been used for versity Hospital, Turkey) were dissolved in physiological decades in the treatment of severe pain in non-depressed saline. Solutions were freshly prepared on the days of patients [12, 13]. Nevertheless, their analgesic mechanism of experimentation. Subcutaneous (s.c.) morphine (5 mg/kg), action is still unclear. It apparently involves a direct, central amitriptyline (20 mg/kg; i.p.), venlafaxine (20 mg/kg; s.c.), potentiation of the endogenous opioid system [14] or acti- dihydroergotamine (100 lg/kg; i.v.) were administered vation of a mixed analgesic effect mediated by serotonergic/ before the analgesia tests. noradrenergic pathways [13, 15] or combinations of these mechanisms. Sacerdote et al. [16] concluded that the anal- Induction of morphine tolerance gesic effect of clomipramine and amitriptyline, and their potentiation of morphine-induced antinociception, are rela- The animals were rendered tolerant to morphine using the ted to an activation of the endogenous opioid system medi- method by a previous study on the induction of morphine ated by serotonin. The TCA amitriptyline has potent tolerance [23]. For tolerance induction, groups of 6–7 rats serotonin/norepinephrine reuptake inhibiting activity and a were randomly chosen and treated subcutaneously (s.c.) potent local anesthetic effect [17, 18]. TCAs produce anal- with morphine 50 mg/kg, once a day for 3 days. To eval- gesia by various mechanisms involving N-methyl-D-aspar- uate the degree of tolerance, the analgesic effect of the test tate (NMDA) receptors, opioids, and substance P [19]. doses of morphine (5 mg/kg, s.c.) were measured by the Intrathecal amitriptyline injection provides spinal anesthesia hot-plate and tail-flick tests 24 h after the last morphine in rats [20]. These data suggest that TCAs might be clinically injection (day 4). In addition, to determine effects of valuable in pain management. Venlafaxine is a structurally amitriptyline, venlafaxine, and dihydroergotamine on the novel phenylethylamine antidepressant drug. In vitro, ven- morphine tolerance, morphine was applied with amitrip- lafaxine blocks the synaptosomal uptake of noradrenaline tyline, venlafaxine, and venlafaxine to the morphine-tol- and serotonin and, to a lesser degree, of dopamine [21]. erant animals on day 4. Finally, dihydroergotamine mesylate (DHE), a serotonin receptor (5-HT1A and 5-HT1B/1D) agonist, has been exten- Antinociception tests sively utilized and studied in the treatment of episodic and chronic migraine [22]. Hot-plate test In the light of these data, the present study aimed to investigate the effects of serotonin/norepinephrine reuptake In this test, animals were individually placed on a hot-plate inhibitors (amitriptyline, venlafaxine) and 5-HT1A-5-HT1B/1D (May AHP 0603 Analgesic Hot-plate; Commat, Turkey) receptor agonist (dihydroergotamine) on morphine analgesia with the temperature adjusted to 55 ± 1 °C. The latency to and tolerance in rats. the first sign of paw licking or jump response to avoid the heat was taken as an index of the pain threshold; the cut-off time was 30 s in order to avoid damage to the paw. The Materials and methods antinociceptive response on the hot-plate is considered to result from a combination of central and peripheral Animals mechanisms [24]. Experiments were performed using male adult Wistar Albino Tail-flick test rats (weighing 175–190 g). Animals were maintained under standard conditions: 12-h light–dark schedule (lights on at We used a standardized tail flick apparatus (May TF 0703 0800 hours) with ad libitum food and water and constant Tail-flick Unit; Commat, Turkey) to evaluate thermal noci- temperature (21 ± 1 °C). All experiments were carried out ception. The radiant heat source was focused on the distal 123 J Physiol Sci (2012) 62:317–323 319 portion of the tail at 3 cm after administration of the vehicle and study drugs. Following vehicle or compound adminis- tration, tail-flick latencies (TFL) were obtained. The infrared intensity was adjusted so that basal TFL occurred at 2.8 ± 0.4 s. Animals with a baseline TFL below 2.4 or above 3.2 s were excluded from further testing. The cutoff latency was set at 15 s to avoid tissue damage. Any animal not responding after 15 s was excluded from the study. The hyperalgesic response in the tail-withdrawal test is generally attributed to central mechanisms [24, 25]. Experimental protocols The analgesic effects of amitriptyline, venlafaxine, dihy- droergotamine, and morphine were considered at 30-min intervals (0, 15, 30, 60, 90, and 120 min) by tail-flick and hot-plate test in rats (n = 6–7). In the morphine-treated rats after induction of morphine tolerance,