LONG-TERM EFFECTS OF CHRONIC COCAINE EXPOSURE THROUGHOUT ADOLESCENCE ON ANXIETY AND STRESS RESPONSIVITY IN A WISTAR RAT MODEL C. J. ALVES, a,b A. MAGALHA ˜ ES, a,d P. MELO, a,c L. DE SOUSA, d M. A. TAVARES, e P. R. R. MONTEIRO c  AND T. SUMMAVIELLE a,c  * a IBMC – Instituto de Biologia Molecular e Celular, Universidade do Porto, Rua do Campo Alegre, 823, 4150-180 Porto, Portugal b INEB – Instituto de Engenharia Biome ´dica, Universidade do Porto, Rua do Campo Alegre, 823, 4150-180 Porto, Portugal c ESTSP – Escola Superior de Tecnologia da Sau ´de do Porto, Instituto Polite ´cnico do Porto, Rua Valente Perfeito, 322, 4400-330 Vila Nova de Gaia, Portugal d ICBAS – Instituto de Cie ˆncias Biome ´dicas Abel Salazar, Universidade de Porto, Rua de Jorge Viterbo Ferreira, 228, 4050-313 Porto, Portugal e FMUP – Faculdade de Medicina, Universidade do Porto, Alameda Professor Herna ˆni Monteiro, 4200-319 Porto, Portugal Abstract—Adolescents display increased vulnerability to engage in drug experimentation. This is often considered a risk factor for later drug abuse. In this scenario, the perma- nent effects of cocaine exposure during adolescence on anxiety levels and stress responsivity, which may result in behavioral phenotypes prone to addiction, are now starting to be unveiled. Thus, the purpose of the present study was to evaluate the long-lasting effects of chronic cocaine administration during adolescence, on anxiety-like behavior and on stress response. Adolescent male Wistar rats were daily administered 45-mg cocaine/kg of body weight in three equal intraperitoneal doses with 1-h interval, from postnatal day (PND) 35 to 50. The effects of cocaine administration on anxiety levels, assessed in the Elevated Plus Maze (EPM), and on social stress response, assessed in the resident– intruder paradigm (R/I), were evaluated 10 days after withdrawal, when rats were reaching the adulthood. The underlying dopaminergic activity, and the corticosterone and testosterone levels were determined. Our results showed that cocaine induced long-lasting alterations in the hypothalamus–pituitary–adrenals (HPA) axis function and in testosterone levels. Such alterations resulted in significant and enduring changes in behavioral responses to environmental challenges, such as the EPM and R/I, including the evaluation of potential threats that may lead to high-risk behavior and low-benefit choices. This was further supported by an altered dopaminergic function in the amygdala and hippocampus. The present findings provide new insights into how the use of cocaine during adolescent development may modulate emotional behavior later in life. Compromised ability to recognize and deal with potential threats is an important risk factor to perpetuate compulsive drug seeking and relapse susceptibility. Ó 2014 IBRO. Published by Elsevier Ltd. All rights reserved. Key words: adolescence, anxiety, cocaine, corticosterone, dopaminergic system, social stress. INTRODUCTION The complex and everlasting features of addiction determine its chronic nature, where the risk of relapse is present even after long withdrawal periods. In drug addiction, several risk factors were identified as promoters of relapse during withdrawal periods, and among these, high anxiety levels and stress are commonly reported (Shaham et al., 2000; Brown et al., 2012; Buffalari et al., 2012). As relapse is prevalent in recovery periods, a better understanding of the involved risk factors will promote the development of more effec- tive treatment strategies in drug addiction. The psychostimulant effects of cocaine are mainly due to its action on the mesocorticolimbic monoaminergic system (Ungless et al., 2001; Nogueira et al., 2006; Gu et al., 2010; Zhang et al., 2013), a neural pathway involved in the processing of affective information and emotional regulation (Kelley and Berridge, 2002). Expo- sure to cocaine induces a neuroadaptational process that will lead to altered emotional behavior (Young et al., 2011), such as increased anxiety levels and impaired stress responsivity. In cocaine abusers, high levels of anxiety were reported after cessation of cocaine use (Gawin and Kleber, 1986; Satel et al., 1991; Coffey et al., 2000). In animal studies, elevated anxiety levels were shown in withdrawal periods, either in initial stages (Harris and Aston-Jones, 1993; Sarnyai et al., 1995) or http://dx.doi.org/10.1016/j.neuroscience.2014.07.008 0306-4522/Ó 2014 IBRO. Published by Elsevier Ltd. All rights reserved. * Correspondence to: T. Summavielle, Addiction Biology Group, Instituto de Biologia Molecular e Celular (IBMC), Rua do Campo Alegre, 823, 4150-180 Porto, Portugal. Tel: +351-226074900; fax: +351-226099157. E-mail address: tsummavi@ibmc.up.pt (T. Summavielle).   These authors contributed equally to the current manuscript. Abbreviations: ANOVA, analysis of variance; DA, dopamine; DAT, dopamine transporter; DOPAC, 3,4-dihydroxyphenylacetic acid; EDTA, ethylenediaminetetraacetic acid; EPM, Elevated Plus Maze; GAPDH, glyceraldahyde-3-phosphate-dehydrogenase; HPA, hypothalamus– pituitary–adrenals; HVA, homovanillic acid/4-hydroxy-3- methoxyphenylacetic acid; Nac, nucleus accumbens; PND, postnatal day; qRT-PCR, quantitative real-time polymerase chain reaction; R/I, Resident–intruder paradigm; SDS–PAGE, sodium dodecyl sulfate polyacrylamide gel electrophoresis; TH, tyrosine hydroxylase; VTA/ SN, ventral mesencephalon (comprising the ventral tegmental area and substantia nigra). Neuroscience 277 (2014) 343–355 343