Antidepressant-Like Effects of -Opioid Receptor Antagonists in the Forced Swim Test in Rats

We showed previously that cAMP response element-binding protein (CREB) within the nucleus accumbens (NAc) of rats regulates immobility in the forced swim test (FST), an assay used to study depression. Because CREB regulates expression of dynorphin (which acts at -opioid receptors) in NAc neurons, these findings raised the possibility that -receptors mediate immobility behaviors in the FST. Here, we report that i.c.v. administration of the -antagonist nor-binaltorphimine dose dependently decreased immobility in the FST, suggesting that it has antidepressant-like effects. Implicating a specific effect at -receptors, similar antidepressant-like effects were seen after treatment with either of two novel, structurally dissimilar -antagonists: 5 -guanidinonaltrindole, which was effective after i.c.v. but not systemic treatment, and 5 -acetamidinoethylnaltrindole (ANTI), which was potent and effective after systemic treatment. The behavioral effects of the -antagonists resembled those of tricyclic antidepressants (desipramine) and selective serotonin reuptake inhibitors (fluoxetine and citalopram). Conversely, systemic administration of the -agonist [5 ,7 ,8 ]-Nmethyl-N-[7-[1-pyrrolidinyl]-1-oxaspiro[4.5]dec8-yl]-benzenacetamide (U-69593) dose dependently increased immobility in the FST, consistent with prodepressant-like effects. The effects of the -ligands in the FST were not correlated with nonspecific effects on locomotor activity. Furthermore, the most potent and effective -antagonist (ANTI) did not affect the rewarding impact of lateral hypothalamic brain stimulation at a dose with strong antidepressant-like effects. These findings are consistent with the hypothesis that CREB-mediated induction of dynorphin in the NAc “triggers” immobility behavior in the FST. Furthermore, they raise the possibility that -antagonists may have efficacy as antidepressants, but lack stimulant or rewardrelated effects. The neurobiology of depression is not understood. Because most antidepressants with clinical efficacy act upon monoamines [primarily norepinephrine (NE) and serotonin (5HT)], much research on depression has focused upon interactions between these neurotransmitters and their reuptake transporters and receptor proteins. However, recent research has become progressively focused upon the intracellular mechanisms of depression and antidepressant treatments (Manji et al., 2001; Duman, 2002; Nestler et al., 2002), with the goal of developing novel therapeutics that act faster, are more efficacious, and have fewer side effects. This approach has led to the study of brain circuits typically associated with reward-related processes, including the mesolimbic dopamine (DA) system (Pliakas et al., 2001; Newton et al., 2002). The mesolimbic DA system projects from the ventral tegmental area of the midbrain to the nucleus accumbens (NAc) of the basal forebrain, and is modulated directly and indirectly by noradrenergic and serotonergic inputs (Pasquier et al., 1977). This circuitry contributes importantly to the hedonic (rewarding) effects of food, sexual behavior, and addictive drugs (Carlezon and Wise, 1996b; Kreek and Koob, 1998; Wise, 1998). It has been proposed that disruption of DA function within the NAc causes anhedonia (reduced ability to experience reward) (Wise, 1982), a hallmark symptom of depression. Consistent with this notion, withdrawal from chronic amphetamine in rats causes decreases in extracellular concentrations of DA within the NAc that are accompaThis study was funded by Grant MH63266 from the National Institute of Mental Health (to W.C.), Grant DA01533 from the National Institute on Drug Abuse (to P.S.P.), an unrestricted gift from Johnson and Johnson (to W.C.), and donations by John A. Kaneb (to W.C.). Article, publication date, and citation information can be found at http://jpet.aspetjournals.org. DOI: 10.1124/jpet.102.046433. ABBREVIATIONS: NE, norepinephrine; 5HT, 5-hydroxytryptamine; DA, dopamine; NAc, nucleus accumbens; CREB, cAMP response elementbinding protein; FST, forced swim test; norBNI, nor-binaltorphimine; GNTI, 5 -guanidinonaltrindole; ANTI, 5 -acetamidinoethylnaltrindole; U-69593, (5 ,7 ,8 )-N-methyl-N-(7-[1-pyrrolidinyl]-1-oxaspiro[4.5]dec8-yl)-benzenacetamide; ICSS, intracranial self-stimulation; DMI, desipramine; FLX, fluoxetine; COC, cocaine; CIT, citalopram; SSRI, selective serotonin reuptake inhibitor; ANOVA, analysis of variance; LSD, least significant difference. 0022-3565/03/3051-323–330$7.00 THE JOURNAL OF PHARMACOLOGY AND EXPERIMENTAL THERAPEUTICS Vol. 305, No. 1 Copyright © 2003 by The American Society for Pharmacology and Experimental Therapeutics 46433/1052156 JPET 305:323–330, 2003 Printed in U.S.A. 323 at A PE T Jornals on N ovem er 9, 2017 jpet.asjournals.org D ow nladed from nied by behavioral depression (Paulson et al., 1991). Similarly, cocaine withdrawal decreases glucose metabolism (Hammer et al., 1993) and reduces sodium currents within this region (Zhang et al., 1998). Chronic stress, a putative precipitator of depression, dramatically alters DA transmission within the NAc (Di Chiara et al., 1999). Together, these studies suggest that the NAc has an important role in the neurobiology of depression. Recently, experience-dependent molecular adaptations that affect DA function within the NAc have been linked to the expression of depression-like signs in rats. Stress elevates the activity of the transcription factor cAMP response element-binding protein (CREB) within the NAc shell (Pliakas et al., 2001). Elevated expression of CREB within the NAc shell increases immobility in the forced swim test (FST) (Pliakas et al., 2001), a rodent model often used to study depression (Porsolt et al., 1977). The effects of CREB within the NAc seem related to its ability to regulate transcription of dynorphin, an endogenous -opioid receptor ligand (Carlezon et al., 1998). Indeed, the -antagonist nor-binaltorphimine (norBNI) attenuates the behavioral effects of elevated CREB expression within the NAc (Carlezon et al., 1998; Pliakas et al., 2001), most likely by blocking -opioid receptors that normally inhibit neurotransmitter release from mesolimbic DA neurons (Shippenberg and Rea, 1997; Di Chiara and Imperato, 1988; Svingos et al., 1999). Recent evidence suggests that the actions of norBNI within the NAc itself are sufficient to cause an antidepressant-like effect in the learned helplessness paradigm (Newton et al., 2002). Taken together, these data raise the possibility that CREB-mediated transcription of dynorphin within the NAc decreases DA function, which triggers signs of depression. Because of the possible connection between dynorphin and symptoms of depression, the present study was designed to use pharmacological tools to determine whether -opioid receptors regulate depression-like signs in the FST. One advantage of the FST is that it identifies in rats treatments with antidepressant effects in humans (Porsolt et al., 1977; Detke et al., 1995). We reported previously (Pliakas et al., 2001) that high doses of norBNI, a well characterized -antagonist with long-lasting effects (Jones and Holtzman, 1992; Spanagel and Shippenberg, 1993), can increase the latency to become immobile in the FST, a putative indicator of antidepressant-like effects. A goal of the present studies was to extend this work by examining the effects of norBNI over a more complete range of doses, while using a more stringent and widely accepted method of scoring the FST (behavioral sampling; Detke et al., 1995). Furthermore, to examine whether the antidepressant-like effects of norBNI are associated with specific actions at -receptors, we conducted similar studies using two novel and structurally dissimilar -antagonists: 5 -guanidinonaltrindole (GNTI) (Jones and Portoghese, 2000; Jewett et al., 2001; Negus et al., 2002) and 5 -acetamidinoethylnaltrindole (ANTI) (Stevens et al., 2000). We also explored the possibility that the -agonist [5 ,7 ,8 ]N-methyl-N-[7-[1-pyrrolidinyl]-1-oxaspiro[4.5]dec8-yl]-benzenacetamide (U-69593) would have the opposite (prodepressantlike) effects on behavior in the FST. In parallel, we examined the effects of each agent on locomotor activity. Finally, because systemic ANTI had strong antidepressant-like effects in the FST, we examined whether it has reward-related effects using intracranial self-stimulation (ICSS) (Wise, 1998). Materials and Methods Rats. A total of 591 male Sprague-Dawley rats (Charles River Laboratories, Inc., Wilmington, MA) were used in this study. Rats used for forced swim testing or locomotor activity testing were housed in groups of four and weighed 325 to 375 g at the time of testing, whereas those used for ICSS testing were housed singly and weighed 350 to 400 g at the time of stereotaxic surgery. All rats were maintained on a 12-h light (7:00 AM–7:00 PM)/12-h dark cycle with free access to food and water except during testing. Experiments were conducted in accordance with the 1996 Guide for the Care and Use of Laboratory Animals (National Institutes of Health) and McLean Hospital policies. Drugs. Desipramine HCl (DMI), fluoxetine HCl (FLX), cocaine HCl (COC), norBNI, and U-69593 were obtained from Sigma-Aldrich (St. Louis, MO). Citalopram (CIT) was obtained from Forest Laboratories (New York, NY). GNTI and ANTI were synthesized at the University of Minnesota. Dosages of all drugs were based on their salt form. DMI, FLX, norBNI, and GNTI were dissolved in distilled water, and COC and CIT were dissolved in saline. U-69593 was dissolved in 0.1 N acetic acid diluted with distilled water. All drugs were administered in a volume of 1 ml/kg, except for FLX (and the FLX-associated vehicle groups), which was administered in a volume of 2 ml/kg because of poor solubility. FST. Three hundred seventy one rats were used for studies of standard psychotropic agents (DMI, FLX, CIT, and COC) and -ligands (norBNI, GNTI, ANTI, and U-69593) in the FST. The FST is a 2-day procedure in which rats swim under conditions in which escape is not possible. On the 1st day, the rats are forced to swim for 15 min. The rats initially struggle to escape from the water, but eventually they adopt a posture of immobility in which they make only the movements necessary to keep their heads above water. When the rats are retested 24 h later, immobility is increased. Treatment with standard antidepressant drugs within the 24-h period between the first exposure to forced swimming and retesting can block facilitated immobility, an effect correlated with antidepressant efficacy in humans (Porsolt et al., 1977; Detke et al., 1995). No treatment was given before the 1st day of the FST for studies involving the standard psychotropic agents, ANTI, or U-69593. For the studies involving norBNI and GNTI, each rat was anesthetized with i.p. pentobarbital (65 mg/kg) and given s.c. atropine sulfate (0.25 mg/kg) to reduce bronchial secretions. Microinjections (i.c.v., 0.3 mm posterior to bregma, 1.2 mm lateral from midsaggital suture, and 4.0 mm below dura; Paxinos and Watson, 1986) of norBNI (1.25–20 g) or GNTI (5.0–20 g) were administered in 2.0 l over 10 min using a Hamilton syringe with a 26-gauge needle (Pliakas et al., 2001). At the dosages used, a single treatment with these agents has extended efficacy: norBNI produces a -receptor-specific blockade in rats (Jones and Holtzman, 1992) for 3 weeks (Spanagel and Shippenberg, 1993). GNTI has detectable -receptor-specific effects in monkeys for 10 days (Negus et al., 2002), although its effects in rats do not seem to endure as long as than those of norBNI (Jewett et al., 2001). (ANTI was not tested in i.c.v. studies because it is a novel compound with modifications to its chemical structure that made it hypothetically more bioavailable by systemic injection than norBNI or GNTI). Rats recovered from surgery for 2 days before the