Efficacy of Peptide Anxiolytic Selank During Modeling of Withdrawal Syndrome in Rats with Stable Alcoholic Motivation

52 Bulletin of Experimental Biology and Medicine, Vol. 157, No. 1, May, 2014 PHARMACOLOGY AND TOXICOLOGY Effi cacy of Peptide Anxiolytic Selank during Modeling of Withdrawal Syndrome in Rats with Stable Alcoholic Motivation L. G. Kolik, A. V. Nadorova, and M. M. Kozlovskaya Translated from Byulleten’ Eksperimental’noi Biologii i Meditsiny, Vol. 1567, No. 1, pp. 61-65, January, 2014 Original article submitted January 31, 2013 We studied the effects of selank on the development of symptoms of acute 48-h alcohol withdrawal in outbred rats drinking 10% ethanol as the only source of fl uid for 24 weeks. In alcohol-preferring animals (mean daily ethanol intake >5.0 g/kg) allowed free choice between 10% ethanol and water, single intraperitoneal injection of selank in a dose of 0.3 mg/kg eliminated anxiety induced by ethanol withdrawal in tests elevated plus maze and social interaction tests and prevented the formation of mechanical allodynia without affecting ethanol consumption. The fi ndings suggest that selank is effective in eliminating of alcohol withdrawal symptoms in rats. Key Words: selank; alcohol withdrawal; anxiety; allodynia; rats Benzodiazepine anxiolytics temporary relieving man- by rapid onset of therapeutic effect and the absence of ifestation of psychopathological symptoms associ- undesirable side effects [4]. Experimental studies have ated with withdrawal syndrome play an important shown that it had a broad spectrum of psychotropic role in the complex medical therapy for alcoholism. activity and restored integrative activity of the CNS However, numerous side effects, such as respiratory impaired by neurotoxic factors of different genesis depression, excitement, potentiation of the narcogene [5]. The mechanism of the central action of selank effect of ethanol, and risk of drug dependence limit includes inhibition of enzyme degradation of enkeph- their use for pharmacological correction of withdraw- alins [1]. Signifi cant therapeutic effect of selank on al syndrome [8]. cognitive functions in combination with anxio lytic Apart from detoxifi cation procedures, schemes and stimulating effects have been demonstrated in of drug therapy for withdrawal syndrome include a clinical settings [6]. wide range of neuropsychotropic drugs (neuroleptics, Here we studied the effects of selank on behavior- antidepressants, and nootropics) that exert various side al manifestations of withdrawal syndrome in chroni- effects [3]. In light of this, the search for more specifi c cally alcoholized rats. and safe antialcoholic drugs for the relief of manifestations of alcohol withdrawal syndrome is an urgent MATERIALS AND METHODS problem. Selank, an original Russian peptide preparation, Experiments were carried out on 8-month-old male is effective in the treatment of generalized anxiety outbred rats with mean body weight 450-470 g in the disorders and asthenic conditions. It is characterized active phase of the experiment (Stolbovaya nursery). Alcohol abuse was modeled using the method for forced alcoholization of the animals providing a V. V. Zakusov Research Institute of Pharmacology, Russian Acad- emy of Medical Sciences, Moscow, Russia. Address for correspon- 10% ethanol solution as the sole source of fl uid for dence: [email protected]. L. G. Kolik 24 weeks. The animals were kept in individual ca- 0007 -4888/14/15710052 © 2014 Springer Science+Business Media New York L. G. Kolik, A. V. Nadorova, and M. M. Kozlovskaya 53 ges under standard vivarium conditions (vivarium of with previously published study protocol [9]. Anxio- V. V. Za kusov Research Institute of Pharmacology) lytic effect was assessed by lengthening the time of at 12-h light/dark cycle with free access to water and active social behavior of the “experimental” rat against chow. Changes in body weight and ethanol consump- the “resident” rat. tion (g/kg) were recorded weekly. During forced alco- Reaction threshold in response to mechanical holization, ethanol was consumed in physiologically stimulation was determined using calibrated von Frey signifi cant amount (6.5±0.2 and 5.0±0.3 g/kg at the fi laments (0.04, 0.07, 0.16, 0.40, 0.60, and 1.00 g), ap- start and end of the experiment, respectively). The ex- plied through a perforated fl oor to the rear paws of rats periments were performed on rats with stable alcoholic placed individually in Plexiglas chambers. Fast paw motivation that was evaluated by mean daily ethanol withdrawal was considered as a positive response and consumption (per 1 kg body weight per day) under stimulation was performed with a smaller fi lament or free choice between 10% ethanol and water. the same hair was used if the rat did not lick or shake After 48-h alcohol deprivation, the animals were the paw. Mechanical sensitivity threshold was deter- tested for alcohol deprivation effect (ADE) to assess mined by the minimum pressure that caused paw with- the nature of formation of alcohol dependence. Ethanol drawal response (3 of 5 attempts). Signifi cant decrease intake was recorded over the fi rst 90 min under condi- in sensitivity thresholds upon fi lament application was tions of free choice between water and 10% ethanol. regarded as mechanical allodynia [11]. Anxiety-related behavior in elevated plus maze The following preparations were used in the study: (EPM) test was evaluated as described previously selank (Thr-Lys-Pro-Arg-Pro-Gly-Pro) in the anxio- [10]. The animals were placed on central platform lytic dose of 0.3 mg/kg intraperitoneally (Institute of and basic spatial and temporal parameters (time spent Molecular Genetics, Russian Academy of Sciences) on the open arms, number of entries into the open and naloxone in a dose of 1.0 mg/kg intraperitoneally arms, number of entries into the closed arms) were (Sigma). Water was injected once 30 min before test- recorded over 300 sec. Increased number of entries ing in a ratio of 0.1 ml/100 g body weight of animal. into the open arms and time spent in open arms in the The study was performed in 5 groups of animals: absence of changes in motor activity (total number of intact rats (not offered ethanol; group 1), animals with entries into the open and closed arms) were consid- stable alcoholic motivation against the background ered as a manifestation of the anxiolytic effect. The of 10% ethanol consumption (group 2), and animals time in the open arms and number of entries in open with stable alcoholic motivation 48 h after ethanol arms were compared and calculated by the following withdrawal receiving water for injections (group 3), formulas: selank (group 4), or naloxone (group 5). The data were processed using the nonparametric time in open arms=time in the open Mann–Whitney U test. arms (sec)/300 sec×100% (1), number of entries into closed arms=number RESULTS of entries into closed arms/total number of entries into open and closed arms×100% (2). Evaluation of the effects of drugs on ADE severity showed that alcohol intake was not signifi cantly af- Social interaction test consists in evaluation of fected by selank (1.08±0.12 g/kg) in contrast to non- the time of active social behavior for rats in pairs (as selective opioid receptor antagonist naloxone (0.36± a measure of anxiety) to record both anxiogenic and 0.06 g/kg vs. 0.88±0.05 g/kg in controls; p<0.001). anxiolytic properties of the drug. Testing was per- Ethanol withdrawal in animals with stable alco- formed in an open fi eld (60×60 cm area surrounded holic motivation was associated with increased anxi- with 50-cm white walls and divided into 16 squares ety, which was seen from reduced time spent in open with a black marker) for 5 minutes under dimmed arms of EPM (p<0.05) and number of entries into open light. A pair of rats (“experimental” and “resident”; arms in comparison with the corresponding parameters difference in body weight did not exceed 10 g) kept in intact rats (Fig. 1). Naloxone had no effect on ani- in individual cells prior to the experiment were placed mal anxiety in EPM. Selank increased the time spent in unfamiliar surroundings of the open fi eld. The dura- (p<0.01) and entries into open arms (p<0.05), which tion of active behavior of “experimental” rat against attests to the anxiolytic effect of this drug. “resident” one (grooming, sniffi ng, rushing, wrestling, Ethanol withdrawal in animals with stable alco- chasing) was registered. Passive contact (sitting and/ holic motivation signifi cantly reduced (p<0.05) the or lying together) was not considered. Stress response time of social interaction. Selank prolonged the active to social interaction was evaluated based on individual contact of rats in pairs (p<0.01) suggesting the anxio- behavioral data of “experimental” rat in accordance lytic effect of this drug (Fig. 2). 54 Bulletin of Experimental Biology and Medicine, Vol. 157, No. 1, May, 2014 PHARMACOLOGY AND TOXICOLOGY Fig. 1. Effects of selank on EPM behavior of rats with stable alcoholic motivation 48 h after ethanol withdrawal. Ordinate: a) time spent in open arms, b) entries into the open arms. I: group 1 (intact rats); II: group 3; III: group 4; IV: group 5. p<0.05 in comparison with *group 1 (I), +group 3 (II). Each group included 9-10 animals. Fig. 2. Effects of selank on behavior of rats with stable alcoholic motivation 48 h after ethanol withdrawal in social interaction test. I: group 1; II: group 3; III: group 4; IV: group 5. Ordinate: time of social Fig. 3. Effects of selank on tactile sensitivity thresholds with interaction for rats in the pair. Total observation time (300 sec) is non-painful mechanical stimulation 48 h after ethanol withdrawal taken as 100%. xp<0.05 in comparison with group 1 (I); *p<0.01 in (median, 25th and 75th quantiles).

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