Binge-Like, Naloxone-Sensitive, Voluntary Ethanol Intake at Adolescence Is Greater Than at Adulthood, but Does Not Exacerbate Subsequent Two-Bottle Choice Drinking

The present study assessed the effects of ethanol exposure during adolescence or adulthood. We exposed Wistar rats, males or females, to self-administered 8–10% (v/v) ethanol (BINGE group) during the first 2 h of the dark cycle, three times a week (Monday, Wednesday, and Friday) during postnatal days (PDs) 32–54 or 72–94 (adolescent and adults, respectively). During this period, controls were only handled, and a third (IP) condition was given ethanol intraperitoneal administrations, three times a week (Monday, Wednesday, and Friday), at doses that matched those self-administered by the BINGE group. The rats were tested for ethanol intake and preference in a two-bottle (24 h long) choice test, shortly before (PD 30 or 70) and shortly after (PD 56 or 96) exposure to the binge or intraperitoneal protocol; and then tested for free-choice drinking during late adulthood (PDs 120–139) in intermittent two-bottle intake tests. Binge drinking was significantly greater in adolescents vs. adults, and was blocked by naloxone (5.0 mg/kg) administered immediately before the binge session. Mean blood ethanol levels (mg/dl) at termination of binge session 3 were 60.82 ± 22.39. Ethanol exposure at adolescence, but not at adulthood, significantly reduced exploration of an open field-like chamber and significantly increased shelter-seeking behavior in the multivariate concentric square field. The rats that had been initially exposed to ethanol at adolescence drank, during the intake tests conducted at adulthood, significantly more than those that had their first experience with ethanol at adulthood, an effect that was similar among BINGE, IP and control groups. The study indicates that binge ethanol drinking is greater in adolescent that in adults and is associated with heightened ethanol intake at adulthood. Preventing alcohol access to adolescents should reduce the likelihood of problematic alcohol use or alcohol-related consequences.SECYT Consolidar 2018ANPyCTUniversidad de GranadaPICT 2015-032

Caloric Restriction in Group-Housed Mice: Littermate and Sex Influence on Behavioral and Hormonal Data

This study was partially funded by the Master's program in Basic and Applied Neuroscience and Pain, University of Granada, the Junta de Andalucia (grants CTS109 and B-CTS-422-UGR18) and the Spanish Ministry of Health (National Drug Plan grant 2020I049). AV-A was recipient of a predoctoral fellowship (Grant/Award number: FPU18/05012) of the Ministerio de Universidades, Spain. LR-L has a research contract under the grant B-CTS-422-UGR18 (Programa Operativo FEDER Andalucia 2014-2020).We acknowledge the technical assistance of Ms. Ana Fernández Ibáñez, who helped with the ACTH hormone analysis, and of Mr. Daniel González Fernández, who edited the final English version of this document.Much of the research done on aging, oxidative stress, anxiety, and cognitive and social behavior in rodents has focused on caloric restriction (CR). This often involves several days of single housing, which can cause numerous logistical problems, as well as cognitive and social dysfunctions. Previous results in our laboratory showed the viability of long-term CR in grouped rats. Our research has studied the possibility of CR in grouped female and male littermates and unrelated CB6F1/J (C57BL/6J x BALBc/J hybrid strain) mice, measuring: (i) possible differences in body mass proportions between mice in ad libitum and CR conditions (at 70% of ad libitum), (ii) aggressive behavior, using the number of pushes and chasing behavior time as an indicator and social behavior using the time under the feeder as indicator, and (iii) difference in serum adrenocorticotropic hormone (ACTH) concentrations (stress biomarker), under ad libitum and CR conditions. Results showed the impossibility of implementing CR in unrelated male mice. In all other groups, CR was possible, with a less aggressive behavior (measured only with the number of pushes) observed in the unrelated female mice under CR conditions. In that sense, the ACTH levels measured on the last day of CR showed no difference in stress levels. These results indicate that implementantion of long-term CR in mice can be optimized technically and also related to their well-being by grouping animals, in particular, related mice.Master's program in Basic and Applied Neuroscience and Pain, University of GranadaJunta de Andalucia CTS109 B-CTS-422-UGR18Spanish Ministry of Health (National Drug Plan grant) 2020I049Ministerio de Universidades, Spain FPU18/05012Programa Operativo FEDER Andalucia B-CTS-422-UGR1

From binge eating to binge drinking: A new and robust paradigm for assessing binge ethanol self-administration in male rats

Animal models of alcohol (ethanol) self-administration are crucial to dissect the neurobiological mechanisms underlying alcohol dependence, yet only a few of these induce pharmacologically relevant levels of alcohol consumption and rarely the alcohol self-administration co-occurs with other addictive behaviours. The present study aims to validate a novel model of voluntary ethanol consumption in male Wistar rats, in which ethanol access follows a binge eating experience. Over 10 sessions, Wistar rats were exposed to binge or control eating (i.e., the ingestion of 11.66 and 0.97 kcal/3 min, respectively, derived from a highly palatable food), immediately followed by two-bottle choice intake tests (2%, 6%, 10% or 14% w/w ethanol vs. water). Rats exposed to binge eating drank significantly more 6% or 10% (w/w) ethanol than control peers, reaching up to 6.3 gEtOH/kg. Rats stimulated with 2%, 6%, 10% or 14% ethanol after binge eating, but not those given those ethanol concentrations after control eating, exhibited significant within-group increases in ethanol drinking. This ethanol consumption was not altered by quinine adulteration (up to 0.1 g/L), and it was blocked by naltrexone (10 mg/kg), administered immediately before binge eating. Blood ethanol levels significantly correlated with ethanol consumption; and the more ethanol consumed, the greater the distance travelled in an open field test conducted after the two-bottle choice test. Altogether, this self-administration model seems a valid and robust alternative with remarkable potential for research on different stages of the alcohol addiction and, particularly, to assess interactions between alcohol consumption and others addictive-like behaviours.Junta de Andalucía, Grant/Award Numbers: CTS109, B-CTS-422-UGR18Ministerio de Universidades, Spain, Grant/Award Number: FPU18/05012Ministry of Science and Innovation, Grant/Award Number: MICIUPID2020- 114269GB-I00Spanish Ministry of Health (Government Delegation for the National Plan on Drugs), Grant/Award Number: PNSD 2020-04

Binge drinking models in rats: assessing the influence of binge eating and sigma-1 receptor

La marcada prevalencia en el consumo de alcohol en la sociedad actual y sus consecuencias en la salud (Griswold et al., 2018) ponen en evidencia la necesidad de profundizar en el conocimiento de los mecanismos que subyacen al consumo de alcohol. En este escenario, son particularmente importantes aquellos modelos animales que demuestren validez a la hora de emular el consumo humano, sin embargo, estos modelos son escasos, especialmente aquellos que modelan comportamientos complejos como pueden ser la relación entre BE y BD. De igual forma, los tratamientos farmacológicos para el AUD son también escasos y de relativa eficacia, o mostrando eficacia solamente en pacientes altamente motivados. Por lo tanto, se hace necesaria la investigación para el descubrimiento de nuevas estrategias farmacológicas que muestren mayor efectividad. Ante estos antecedentes la presente tesis doctoral se plantea con los siguientes objetivos: Teniendo en cuenta la falta de modelos animales de ingesta voluntaria de etanol tipo BD que imiten el comportamiento humano, en la presente Tesis Doctoral el primer objetivo propuesto es llenar este vacío, al menos parcialmente, proponiendo un nuevo modelo de autoadministración de etanol tipo BD en ratas. El procedimiento se basa en la exposición repetida y breve a una gran cantidad de pellets de precisión azucarados de gran palatabilidad que conducirán a su consumo como en forma de BE, lo que dará lugar a una alta autoadministración voluntaria de etanol de forma BD. Con el fin de evaluar la validez del modelo, el segundo objetivo fue evaluar si la ingesta de etanol ocurría a pesar de tener un sabor aversivo, lo cual indicaría que el consumo se había vuelto habitual o compulsivo (Hopf & Lesscher, 2014; Radke et al., 2020). Para ello, se realizó un experimento de adulteración de la solución de etanol con quinina buscando comprobar si la autoadministración de solución de etanol por parte de las ratas se volvió resistente a la quinina al final del procedimiento. El tercer objetivo consistió en corroborar si el consumo voluntario en el presente modelo esta mediado, al menos en parte, por el sistema opioide y por lo tanto el tratamiento con naltrexona resultaba efectivo para reducir la autoadministración (Guardia Serecigni, 2015). De cumplirse este objetivo de pondría de manifiesto la validez predictiva (es decir, el valor traslacional) del modelo. Varios estudios muestran la bidireccionalidad sobre el consumo de alcohol de la modulación farmacológica de los S1-R (Quadir et al., 2019), por lo tanto, el cuarto objetivo se planteó evaluar si el tratamiento con agentes farmacológicos cuya diana es el receptor S1 tenía un efecto sobre el consumo de etanol en nuestro modelo. Finalmente, para continuar con la evaluación de la efectividad de la modulación farmacológica de los S1-R, el quinto objetivo se propuso comprobar si el antagonismo S1-R era efectivo en la reducción del consumo de etanol en un modelo de consumo en ratas adolescentes.Tesis Univ. Granada.Junta de AndalucíaSantander Investigación (2020)Junta de Andalucía (grupo CTS-109)Programa Operativo FEDER de Andalucía (proyecto B-CTS-422-UGR18)Plan Nacional Sobre Drogas 2020 (proyecto PND-2020-049

Binge eating promotes ethanol self-administration in female rats with a history of intermittent ethanol exposure at adolescence

Background: Ethanol drinking begins during adolescence and, particularly when occurs in a binge-like pattern, exerts lingering adverse consequences. Pre-clinical studies indicate that intermittent ethanol exposure (IEA, a model of repeated ethanol intoxication), or binge eating (BE) can increase subsequent ethanol consumption. It is unknown if the promoting effects of BE upon ethanol drinking are found in female rats and are modulated by IEA at adolescence. This study assessed interactive effects between IEA and BE, upon ethanol drinking. Methods: Female Wistar rats were given 4.0 g/kg ethanol, every other day from postnatal day 25-45. At adulthood, they were exposed to sessions in which a brief offering of a sizeable portion of highly palatable sugary pills was followed by a 120-min exposure to an ethanol bottle. Results: Exploratory activity and recognition memory was not affected by the IEA. Glutathione peroxidase and catalase activity, and lipid peroxidation (measured in blood and brain at the end of the procedure) were not significantly affected by IEA or BE exposure. BE alone had a mild promoting effect on ethanol ingestion. Those rats that underwent IEA and BE, however, exhibited heightened and sustained ethanol self-administration (average of 2.12 g/kg/120 min, vs 1.15 g/kg/120 min of the other groups), that persisted throughout the BE sessions. IEA and a history of BE also promoted ethanol intake or preference in a two-bottle endpoint test. Conclusion: The study suggests that exposure to IEA exerts, when followed by BE at adulthood, promoting effects upon ethanol intake, particularly at concentrations ≥ 6%.his work was supported by PICT 2019-00180 and PICT 2017-0874 of Agencia Nacional de Promoción Científica y Tecnológica (FONCyT), to RMP and MBV, respectively; and by the Spanish Ministry of Health (Government Delegation for the National Plan on Drugs, PNSD 2020-049) to CMC and IMH. LRL was a recipient of a fellowship of the Programa Becas Santander Iberoamérica Investigación - UGR 2020/2021. The funders had no further role in study design; in the collection, analysis, and interpretation of data; in the writing of the report; and in the decision to submit the paper for publication

Binge ethanol self-administration at adolescence, in Wistar rats, promotes ethanol drinking at adulthood in a S1RA sensitive manne

Abstract Background Binge drinking at adolescence is a risk factor for problematic alcohol (ethanol) consumption later in life, yet the murine studies that modelled this phenomenon via ethanol self-administration have provided mixed findings. Antagonism of the sigma-1 receptor (S1-R) system at adolescence modulates ethanol’s motivational effects and intake. It is still unknown, however, whether this antagonism would protect against enhanced ethanol intake at adulthood after adolescent binge ethanol exposure. Methods Exp. 1 and 2 tested adults male or female Wistar rats -exposed or not to ethanol self-administration at adolescence (postnatal days 31–49; nine 2-hour sessions of access to 8–10% ethanol)- for ethanol intake using 24-h two-bottle choice test (Exp. 1) or time restricted, single-bottle, tests (Exp. 2). Experiments 2–5 evaluated, in adolescent or adult rats, the effects of the S1-R antagonist S1RA on ethanol intake and on ethanol-induced conditioned taste or place aversion. Ancillary tests (e.g., novel object recognition, ethanol-induced locomotor activity) were also conducted. Results Adolescent ethanol exposure promoted ethanol consumption at both the restricted, single-bottle, and at the two-bottle choice tests conducted at adulthood. S1RA administration reduced ethanol intake at adulthood and facilitated the development of ethanol-induced taste (but not place) aversion. Conclusions S1RA holds promise for lessening ethanol intake after chronic and substantial ethanol exposure in adolescence that results in heightened ethanol exposure at adulthood. This putative protective effect of S1-R antagonism may relate to S1RA exacerbating the aversive effects of this drug.This work was supported by PICT-2021-I-A-00035 and PICT-2019–00180 of Agencia Nacional de Promoción Científica y Tecnológica (FONCyT), to RMP, the Spanish Ministry of Health (Government Delegation for the National Plan on Drugs, PNSD 2020–049) to CMC and IMH and the grant C-CTS-307-UGR23 (Programa Operativo FEDER Andalucía 2021–2027) to RMP, CMC and IM

Sigma-1 antagonism inhibits binge ethanol drinking at adolescence

Ethanol use during adolescence is a significant health problem, yet the pharmacological treatments to reduce adolescent binge drinking are scarce. The present study assessed, in male and female adolescent Wistar rats, if the sigma-1 receptor (S1-R) antagonists S1RA or BD-1063 disrupted ethanol drinking. Methods: Three times a week, for two weeks, the rats received the S1-R antagonists. Thirty min later they were exposed, for 2 h, to a bottle of 8% or 10 % v/v ethanol. A 24 h, two-bottle, ethanol intake test was conducted after termination of these procedures. A subset of these rats was tested for recognition memory via the novel object recognition test. Results: The rats given 64 mg/kg S1RA drank, in each binge session, significantly less than vehicle counterparts. Male rats given 4 or 16 mg/kg S1RA drank significantly less than those given 0 mg/kg in session 3 or in session 1 and 2, respectively; whereas female rats given 4 or 16 mg/kg drank significantly less than females given 0 mg/kg in session 2–5 or in sessions 2–6, respectively. Administration of 32 mg/kg, but not of 2 or 8 mg/kg, BD-1063 suppressed, across sessions, ethanol drinking. S1-R antagonism reduced absolute ethanol drinking at the two-bottle choice post-test. Recognition memory was not affected by the ethanol exposure. Conclusions: The results indicate that S1-R antagonists may be promising targets to prevent increases in ethanol intake at adolescence. The persistent effect of S1-R antagonism in free-choice drinking suggests that modulation of the S1-R is altering plastic effects associated with ethanol exposure.Fil: Ruiz Leyva, Leandro. Universidad de Granada; España. Instituto de Investigación Biosanitaria (IBS); EspañaFil: Salguero, Juan Agustín. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Instituto de Investigación Médica Mercedes y Martín Ferreyra. Universidad Nacional de Córdoba. Instituto de Investigación Médica Mercedes y Martín Ferreyra; ArgentinaFil: Morón, Ignacio. Universidad de Granada; España. Instituto de Investigación Biosanitaria (IBS); EspañaFil: Portillo-Salido, Enrique. Parc Científic de Barcelona; EspañaFil: Cendán, Cruz Miguel. Instituto de Investigación Biosanitaria (IBS); España. Universidad de Granada; EspañaFil: Pautassi, Ricardo Marcos. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Instituto de Investigación Médica Mercedes y Martín Ferreyra. Universidad Nacional de Córdoba. Instituto de Investigación Médica Mercedes y Martín Ferreyra; Argentin

Binging from Food to Alcohol: A Sequential Interaction Between Binging Behaviors in Male Wistar Rats

The development of excessive alcohol (ethanol) and/or highly palatable food self-administration is an essential task to elucidate the neurobiological mechanisms that underlie these behaviors. Previous work has highlighted that ethanol self-administration is modulated by both the induction of aversive states (i.e., stress or frustration) and by the concurrent availability of appetitive stimuli (e.g., food). In our protocol, rats are food deprived for three days until they reach 82%–85% of their ad libitum weight. After that, rats are exposed daily for 10 days to a brief binge or control eating experience with highly sugary and palatable food (i.e., the ingestion of 11.66 and 0.97 kcal/3 min, respectively), which is followed by a two-bottle-choice test (ethanol vs. water) in their home cages for 90 min. This model induces robust binge eating, which is followed by a selective increase in ethanol self-administration. Therefore, this protocol allows to study: a) behavioral and neurobiological factors related to binge eating, b) different stages of alcohol use, and c) interactions between the latter and other addictive-like behaviors, like binge eating.Spanish Ministry of Health (Government Delegation for the National Plan on Drugs: PNSD 2020-049)Junta de Andalucía (grants CTS109 and HUM784)University of Granada (PP2022.PP-16)PICT-2019- 00180 and PICT-2018-00597 of FONCyT-Argentin