Male Long-Evans Rats Research Articles

Effects of dopamine receptor antagonism and amphetamine-induced psychomotor sensitization on sign- and goal-tracking after extended training

The dopamine system is important for incentive salience attribution, where motivational value is assigned to conditioned cues that predict appetitive reinforcers. However, the role of dopamine in this process may change with extended training. We tested the effects of dopamine D1-like and D2-like receptor antagonism on the expression of sign-tracking and goal-tracking conditioned responses following extended Pavlovian conditioned approach (PCA) training. We also tested if amphetamine-induced psychomotor sensitization accelerates the enhanced acquisition of sign-tracking that is observed with extended training. In experiment 1, 24 male Long-Evans rats received 20 PCA sessions in which one lever (CS+, 10 s) predicted 0.2 ml sucrose (10 %, w/v) delivery and the other lever (CS–) did not. SCH-23390 (D1-like antagonist) or eticlopride (D2-like antagonist) were administered before non-reinforced behavioural tests at doses of 0, 0.01, and 0.1 mg/kg (s.c.). In experiment 2, rats received vehicle or 2 mg/kg amphetamine (i.p.) for 7 days (n = 12/group). Ten days later, they received 16 PCA training sessions. Both doses of SCH-23390 reduced sign- and goal-tracking, but also reduced locomotor behaviour. A low dose of eticlopride (0.01 mg/kg) selectively reduced goal-tracking, without affecting sign-tracking or locomotor behaviour. Amphetamine produced psychomotor sensitization, and this did not affect the acquisition of sign- or goal-tracking. Following extended PCA training, dopamine D2-like receptor activity is required for the expression of goal-tracking but not sign-tracking. Psychomotor sensitization to amphetamine did not impact incentive salience attribution; however, more selective manipulations of the dopamine system may be needed.

Chronic ethanol vapor exposure potentiates cardiovascular responses to acute stress in male but not in female rats

BackgroundEthanol use is related to a wide variety of negative health outcomes, including cardiovascular diseases. Stress is also involved in numerous pathologies, such as cardiovascular diseases and psychiatric disorders. Sexual dimorphism is an important factor affecting cardiovascular response and has been proposed as a potential risk factor for sex-specific health problems in humans. Here, we evaluated the effect of prolonged ethanol vapor inhalation on arterial pressure, heart rate, and tail skin temperature responses to acute restraint stress, investigating differences between male and female rats.MethodsWe exposed male and female Long-Evans rats to ethanol vapor for 14 h, followed by ethanol withdrawal for 10 h, for 30 consecutive days, or to room air (control groups). The animals underwent surgical implantation of a cannula into the femoral artery for assessment of arterial pressure and heart rate values. The tail skin temperature was measured as an indirect measurement of sympathetic vasomotor response.ResultsChronic ethanol vapor inhalation reduced basal heart rate in both female and male rats. Sex-related difference was observed in the decrease of tail cutaneous temperature evoked by stress, but not in the pressor and tachycardiac responses. Furthermore, prolonged ethanol inhalation enhanced the blood pressure and heart rate increase caused by acute restraint stress in male, but not in female rats. However, no effect of chronic ethanol vapor was observed in the tail cutaneous temperature response to restraint in either sex.ConclusionChronic ethanol vapor exposure increased the cardiovascular reactivity to stress in male, but not in female rats.

Effects of 5-HT2A receptor agonist 2,5-dimethoxy-4-iodoamphetamine on alcohol consumption in Long-Evans rats.

The objectives of this study were to determine alcohol consumption after administration of (R)(-)-2,5-dimethoxy-4-iodoamphetamine (DOI) or naltrexone in Long-Evans rats, and to assess the effectiveness of these treatments based on individual differences in alcohol consumption. Adult male Long-Evans rats (N = 16) were given opportunities to orally self-administer a 20% (v/v) ethanol (EtOH) solution using an intermittent access, two-bottle (vs. tap water) choice procedure in their home cages. EtOH consumption and preference, total fluid consumption and food intake were measured. Last, we assessed the effects of naltrexone (1 mg/kg; subcutaneous) and (R)(-)-DOI (0.1-1 mg/kg; subcutaneous) on EtOH intake and preference using a quartile analysis. Rats showed stable EtOH (20%) intake and preference after 15 EtOH access sessions. Naltrexone produced a transient decrease in EtOH intake, but an inconsistent effect on EtOH preference, whereas DOI dose-dependently reduced EtOH intake and preference for at least 24 h. Subsequent quartile analyses revealed that rats with the highest EtOH intake during the first 60 min of access to EtOH showed greater reductions in EtOH intake and preference after DOI treatment. This is the first report to show that DOI-elicited reductions in EtOH intake and preference in rats depend on baseline EtOH intake, perhaps supporting a 'baseline dependency' hypothesis of effectiveness with phenethylamine psychedelics on EtOH consumption. If so, individuals with greater potential to develop severe AUDs may be particularly responsive to the positive motivational changes produced by treatment with psychedelics that target the 5-HT2 receptor family.

Prefrontal NMDA-receptor antagonism disrupts encoding or consolidation but not retrieval of incidental context learning

The Context Preexposure Facilitation Effect (CPFE) is a variant of contextual fear conditioning in which learning about the context, acquiring a context-shock association, and retrieval of this association occur separately across three phases (context preexposure, immediate-shock training, and retention). We have shown that prefrontal inactivation or muscarinic-receptor antagonism prior to any phase disrupts retention test freezing during the CPFE in adolescent rats (Heroux et al., 2017; Robinson-Drummer et al., 2017). Furthermore, the medial prefrontal cortex (mPFC) is the only region in which robust learning-related expression of the immediate early genes c-Fos, Arc, Egr-1 and Npas4 is observed during immediate-shock training in the CPFE (Asok et al., 2013; Heroux et al., 2018; Schreiber et al., 2014). However, the role of prefrontal NMDA-receptor plasticity in supporting preexposure- and training-day processes of the CPFE is not known. Therefore, the current study examined the effects of intra-mPFC infusion of the NMDA-receptor antagonist MK-801 or saline vehicle prior to context preexposure (Experiment 1) or immediate-shock training (Experiment 2) in adolescent Long-Evans male and female rats. This infusion given prior to context preexposure but not training abolished retention test freezing, with no difference between MK-801-infused rats and non-associative controls preexposed to an alternative context (pooled across drug). These results demonstrate a role of prefrontal NMDA-receptor plasticity in the acquisition and/or consolidation of incidental context learning (i.e., encoded in the absence of reinforcement). In contrast, this plasticity is not required for context retrieval, or acquisition, expression, or consolidation of a context-shock association during immediate-shock training in the CPFE. These experiments add to a growing body of work implicating the mPFC in Pavlovian contextual fear conditioning processes in rodents.

Caffeine increases alcohol self-administration, an effect that is independent of dopamine D2 receptor function.

The rising popularity of alcohol mixed with energy drinks (AmEDs) has become a significant public health concern, with AmED users reporting higher levels of alcohol intake than non-AmED users. One mechanism proposed to explain this heightened level of alcohol intake in AmED users is that the high levels of caffeine found in energy drinks may increase the positive reinforcing properties of alcohol, an effect that may be dependent on interactions between adenosine receptor signaling pathways and the dopamine D2 receptor. Therefore, the purpose of the current study was to confirm whether caffeine does increase the positive reinforcing effects of alcohol using both fixed ratio (FR) and progressive ratio (PR) designs, and to investigate a potential role of the dopamine D2 receptor to caffeine-induced increases in alcohol self-administration. Male Long-Evans rats were trained to self-administer a sweetened alcohol solution (10% v/v alcohol+2% w/v sucrose) on an FR2 schedule of reinforcement, and the effects of caffeine (0, 5, 10, and 20mg/kg, i. p. [intraperitoneally]) on the maintenance of alcohol self-administration and alcohol break point were examined. Parallel experiments in rats trained to self-administer sucrose (0.8% w/v) were conducted to determine whether caffeine's reinforcement-enhancing effects extended to a non-drug reinforcer. Caffeine pretreatment (5-10mg/kg) significantly increased sweetened alcohol self-administration and motivation for a sweetened alcohol reinforcer. However, similar increases in self-administration of a non-drug reinforcer were not observed. Contrary to our hypothesis, the D2 receptor antagonist eticlopride did not block a caffeine-induced increase in sweetened alcohol self-administration, nor did it alter caffeine-induced increases in motivation for a sweetened alcohol reinforcer. Taken together, these results support the hypothesis that caffeine increases the positive reinforcing effects of alcohol, which may explain caffeine-induced increases in alcohol intake. However, the reinforcement-enhancing effects of caffeine appear to be independent of D2 receptor function.

Noninvasive Brain Stimulation Rescues Cocaine-Induced Prefrontal Hypoactivity and Restores Flexible Behavior

BackgroundTo obtain desirable goals, individuals must predict the outcome of specific choices, use that information to direct appropriate actions, and adjust behavior accordingly in changing environments (behavioral flexibility). Substance use disorders are marked by impairments in behavioral flexibility along with decreased prefrontal cortical function that limits the efficacy of treatment strategies. Restoring prefrontal hypoactivity, ideally in a noninvasive manner, is an intriguing target for improving flexible behavior and treatment outcomes. MethodsA behavioral flexibility task was used in Long-Evans male rats (n = 97) in conjunction with electrophysiology, optogenetics, and a novel rat model of transcranial alternating current stimulation (tACS) to examine the prelimbic cortex (PrL) to nucleus accumbens (NAc) core circuit in behavioral flexibility and determine whether tACS can restore cocaine-induced neural and cognitive dysfunction. ResultsOptogenetic inactivation revealed that the PrL-NAc core circuit is necessary for the ability to learn strategies to flexibly shift behavior. Cocaine self-administration history caused aberrant PrL-NAc core neural encoding and deficits in flexibility. Optogenetics that selectively activated the PrL-NAc core pathway prior to learning rescued cocaine-induced cognitive flexibility deficits. Remarkably, tACS prior to learning the task reestablished adaptive signaling in the PrL-NAc circuit and restored flexible behavior in a relatively noninvasive and frequency-specific manner. ConclusionsWe establish a role of NAc core–projecting PrL neurons in behavioral flexibility and provide a novel noninvasive brain stimulation method in rats to rescue cocaine-induced frontal hypofunction and restore flexible behavior, supporting a role of tACS as a therapeutic to treat cognitive deficits in substance use disorders.

Spirulina ameliorates arsenic induced reproductive toxicity in male rats.

Spirulina (Spirulina platensis), has numerous health benefits including antioxidant, immunomodulatory, and anti-inflammatory activities, works against heavy metal toxicity, and is often used as a food supplement in human, animals, birds and fishes. This study aimed to evaluate the protective ability of the dietary spirulina against the toxic effects of inorganic arsenic (iAs) on male reproductive parameters in rats. Seventy-two mature Long-Evans male rats, dividing into six groups (T0, T1, T2, T3, T4 and T5) (12 rats/group) were included in this study. The T3, T4 and T5 group rats were treated with three consecutive doses (1.0 g, 1.5 g and 2.0 g/kg feed) of spirulina in feed along with 3.0 mg NaAsO2/kg body weight (BW) in drinking water (DW) daily for 90 days. Each rat of group T1 received NaAsO2 (3.0 mg/kg BW) in DW, and those of T2 group were fed with spirulina (2.0 g/kg feed) daily for 90 days. The rats of group T0 served as the control with normal feed and water. Total arsenic (tAs) contents, reproductive parameters (testicular weight, sperm motility and morphology), and histological changes in the testicles were evaluated in these rats. Arsenic dosing significantly (p=0.003, Kruskal-Wallis test) increased the tAs contents in the testicles, decreased testes weight, sperm morphology and motility compared to the controls. The effect of arsenic dosing was also evidenced by the histological changes like decreased germinal layers in the seminiferous tubules of the treated rats. Moreover, dietary spirulina (2.0 g/kg feed) supplementation significantly (p=0.011, Kruskal-Wallis test) lowered tAs contents in testicles and increases testes weights, sperm motility and morphology. Therefore, spirulina can be used as an effective dietary supplement to ameliorate the adverse effects of arsenic induced reproductive toxicities. However, further study is required to elucidate the underlying molecular mechanisms of reduction of arsenic induced reproductive toxicity by spirulina.

Effects of repeated adolescent exposure to cannabis smoke on cognitive outcomes in adulthood.

Cannabis (marijuana) is the most widely used illicit drug in the USA, and consumption among adolescents is rising. Some animal studies show that adolescent exposure to delta 9-tetrahydrocannabinol or synthetic cannabinoid receptor 1 agonists causes alterations in affect and cognition that can persist into adulthood. It is less clear, however, whether similar alterations result from exposure to cannabis via smoke inhalation, which remains the most frequent route of administration in humans. To begin to address these questions, a rat model was used to determine how cannabis smoke exposure during adolescence affects behavioral and cognitive outcomes in adulthood. Adolescent male Long-Evans rats were assigned to clean air, placebo smoke, or cannabis smoke groups. Clean air or smoke exposure sessions were conducted daily during adolescence (from P29-P49 days of age ) for a total of 21 days, and behavioral testing began on P70. Compared to clean air and placebo smoke conditions, cannabis smoke significantly attenuated the normal developmental increase in body weight, but had no effects on several measures of either affect/motivation (open field activity, elevated plus maze, instrumental responding under a progressive ratio schedule of reinforcement) or cognition (set shifting, reversal learning, intertemporal choice). Surprisingly, however, in comparison to clean air controls rats exposed to either cannabis or placebo smoke in adolescence exhibited enhanced performance on a delayed response working memory task. These findings are consistent with a growing body of evidence for limited long-term adverse cognitive and affective consequences of adolescent exposure to relatively low levels of cannabinoids.

Cholinergic neurotransmission in the basolateral amygdala during cued fear extinction

Cholinergic neuromodulation plays an important role in numerous cognitive functions including regulating arousal and attention, as well as associative learning and extinction processes. Further, studies demonstrate that cholinergic inputs from the basal forebrain cholinergic system influence physiological responses in the basolateral amygdala (BLA) as well as fear extinction processes. Since rodent models display individual differences in conditioned fear and extinction responses, this study investigated if cholinergic transmission in the BLA during fear extinction could contribute to differences between extinction resistant and extinction competent phenotypes in outbred Long-Evans male rats. Experiment 1 used in vivo microdialysis to test the hypothesis that acetylcholine (ACH) efflux in the BLA would increase with presentation of an auditory conditioned stimulus (CS+) during extinction learning. Acetylcholine efflux was compared in rats exposed to the CS+, a CS- (the tone never paired with a footshock), or to a context shift alone (without CS+ tone presentation). Consistent with acetylcholine's role in attention and arousal, ACH efflux in the BLA was increased in all three groups (CS+, CS-, Shift Alone) by the initial context shift into the extinction learning chamber, but returned more rapidly to baseline levels in the Shift Alone group (no CS+). In contrast, in the group exposed to the CS+, ACH efflux in the BLA remained elevated during continued presentation of conditioned cues and returned to baseline more slowly, leading to an overall increase in ACH efflux compared with the Shift Alone group. Based on the very dense staining in the BLA for acetylcholinesterase (ACHE), Experiment 2 examined if individual differences in fear extinction were associated with differences in cholinesterase enzyme activity (CHE) in the BLA and/or plasma with a separate cohort of animals. Cholinesterase activity (post-testing) in both the BLA and plasma was higher in extinction competent rats versus rats resistant to extinction learning. There was also a significant negative correlation between BLA CHE activity and freezing during extinction learning. Taken together, our results support a role for ACH efflux in the BLA during cued fear extinction that may be modulated by individual differences in ACHE activity, and are associated with behavioral responses during fear extinction. These findings implicate individual differences in cholinergic regulation in the susceptibility to disorders with dysregulation of extinction learning, such post-traumatic stress disorder (PTSD) in humans.

Dorsal hippocampus plays a causal role in context-induced reinstatement of alcohol-seeking in rats

Drug addiction is a chronic mental disorder characterized by frequent relapses. Contextual cues associated with drug use to play a critical causal role in drug-seeking behavior. The hippocampus has been implicated in encoding drug associative memories. Here we examine whether the dorsal hippocampus mediates context-induced reinstatement of alcohol-seeking. Male Long-Evans rats were trained to self-administer alcohol in Context A. Alcohol self-administration was extinguished in a distinct context (Context B). On the test day, animals were re-exposed to the alcohol Context A or the extinction Context B. Next, to assess a causal role for the dorsal hippocampus in context-induced alcohol-seeking, on the test day, we injected cobalt chloride (CoCl2; a nonselective synapse inhibitor) or vehicle into the dorsal hippocampus, and 15 min later, rats were tested by re-exposing them to the drug-associated context. The re-exposure to the alcohol-associated Context A reinstated alcohol seeking and increased Fos-positive cells in the dorsal hippocampus neurons (CA1, CA3, and Dentate Gyrus). Pharmacological inactivation with cobalt chloride of the dorsal hippocampus attenuated the reinstatement of alcohol-seeking. Our data suggest that the dorsal hippocampus may be involved in context-induced alcohol-seeking behavior.

Procognitive profiling of a serotonin 5-HT6 receptor antagonist in a complex model system in rats: A novel translational approach for clinical prediction

IntroductionThe serial clinical failures of novel cognitive enhancer candidates point out the lack of predictive power in the preceding animal experimentation. For a more predictive profiling of putative procognitive drugs in rodents, we recently elaborated a methodical approach which consists of three fundamental steps: 1. teaching various learning tasks representing different cognitive domains to the same cohort of animals with the aim to create a population with ‘widespread knowledge’. 2. Applying a cognitive deficit-inducing intervention to transform this cohort of animals to a ‘patient population’. 3. Testing putative procognitive drugs with a ‘clinical trial-like’ design on the wide spectrum of cognitive (dys)functions in the actual ‘patient population’. The present study has been the first trial to test the feasibility and utility of the proposed system. MethodsThe population with ‘widespread knowledge’ consisted of 2 year old male Long-Evans rats with a learning history in five-choice serial reaction time task (5-CSRTT, attentional paradigm), Morris water maze (MWM, spatial learning), a cooperative task carried out in pairs (social learning), and a skill-learning task, „pot-jumping”. For inducing cognitive deficit, thus creating a ‘patient population’ we increased the difficulty of the tasks. For the cognitive enhancer mechanism to test in the system we chose a serotonin 5-HT6 receptor antagonist compound, RO4368554. Animals were randomly assigned to vehicle- and drug treated groups based on their baseline learning performance and their response in a pilot test of increase in task difficulty. During the 13-day long treatment with 3 mg/kg ip. RO4368554 all the learning paradigms were repeatedly run with increased difficulty supplemented with a novel object recognition test (NOR, episodic memory). ResultsIn the 5-CSRTT, reducing the stimulus duration from 1 s to 0.25 s caused a significant decrease in the percentage of correct responses (from 52 % to 31 % in the control group) which was not affected by the 5-HT6 receptor antagonist treatment (correct responses decreased from 58 % to 31 %). In the MWM, replacing the escape platform to a new location did not mean a hard challenge for the rats. Members of both groups could find it within a relatively short time: mean escape latencies were 83 s and 65 s at the first replacement trial and 58 s and 74 s at the second one in the control and drug-treated groups, respectively. In the cooperation paradigm, where the rats had to perform simultaneous nose-pokes to get a reward, task difficulty was increased by requiring two consecutive simultaneous nose-poking from the animals. This caused a fall in the percentage of successful trials in both groups (from 48 % to 12 % and from 50 % to 20 % in the saline – and drug-treated group, respectively), however, by the end of the treatment RO4368554-treated animals showed significantly higher performance (29 %) than saline treated rats (2%). The NOR test, carried out with a 5 -h delay, revealed poor recognition memory in both groups (discrimination index (DI) values were 0.13 and 0.06 for saline and RO4368554, respectively). Performance in the pot jumping test was also not improved by the drug-treatment. ConclusionsThe applied study design allowed parallel measurements of the action of the test compound on several cognitive functions and to follow its time course. RO4368554 did not show notable effects on impaired attention and visual recognition; nor did it affect spatial and procedural learning, but it exerted beneficial effect on cooperative behaviour. The revealed activity pattern highlight the cognitive domain most sensitive to the particular drug effect and may give hints for further target validating and clinical studies.

Sex and age differences in locomotor and anxiety-like behaviors in rats: From adolescence to adulthood.

Risk-taking behaviors are a primary contributor to elevated adolescent injury and mortality. Locomotor and anxiety-like behaviors in rodents have been used to examine risk-taking. Here, we examined risk-taking behavior (i.e., changes in locomotor and anxiety-like behaviors) from early to late adolescence and adulthood in male and female rats in the open-field (OF) apparatus and the light-dark (LD) test. We also examined whether these behaviors are affected by an early adolescent immune stressor, lipopolysaccharide (LPS). Long-Evans male and female rats were injected with LPS (200μg/kg) or vehicle control in early adolescence (postnatal day [PND] 30 and 32). Anxiety-like behavior and locomotor activity were measured in early (PND 38-40), late adolescence (PND 50), and adulthood (PND 88 and 98) in the OF and in early adolescence (PND 42) and adulthood (PND 90) in the LD test. Early and late adolescent rats displayed significantly greater locomotor and anxiety-like behaviors than adult rats in the OF and LD test. Sex differences were also found, with adolescent and adult females displaying greater locomotor and anxiety-like behaviors than male rats in the OF and LD tests. LPS administered two times in early adolescence did not have a significant impact on either locomotor or anxiety-like behaviors suggesting minimal impact of the immune stressor.

Effects of pituitary adenylate cyclase-activating polypeptide isoforms in nucleus accumbens subregions on ethanol drinking.

While limited research has implicated the neuropeptide, pituitary adenylate cyclase-activating polypeptide (PACAP), in problematic alcohol use, the brain regions and isoforms involved in this effect remain to be determined. One region that has been found both to exhibit PACAP binding and, separately, to be involved in ethanol drinking is the nucleus accumbens (NAc). Thus, this study sought to characterize the effect of the PACAP isoforms in the NAc on ethanol drinking under the intermittent-access two-bottle-choice paradigm, in male and female Long-Evans rats. With microinjection into the medial NAc shell, PACAP-27 but not PACAP-38 was found to dose-dependently reduce binge-like ethanol drinking. In contrast, the PACAP receptor antagonist, PACAP (6-27), but not PACAP (6-38), enhanced ethanol drinking. This effect of PACAP was substance specific, as neither isoform in the NAc shell affected binge-like sucrose drinking. It was also anatomically specific, as PACAP-38 rather than PACAP-27 suppressed ethanol drinking when injected into the NAc core, and PACAP-27 instead enhanced drinking when injected into the caudal third of the medial NAc shell. Finally, while PACAP-38 in the NAc shell affected stress-related exploratory behavior, reducing time spent in the light chamber of a light-dark box, PACAP-27 did not significantly affect behavior in a light-dark box or open field. Together, these results, showing that PACAP-27 in the NAc shell attenuates binge-like ethanol drinking without affecting select stress-related behaviors, suggest that compounds related to this PACAP isoform should be investigated as potential novel therapeutics for the treatment of alcohol use disorder.

Low-dose alcohol: Interoceptive and molecular effects and the role of dentate gyrus in rats.

Alcohol abuse and dependence are world-wide health problems. Most research on alcohol use focuses on the consequences of moderate to high levels of alcohol. However, even at low concentrations, alcohol is capable of producing effects in the brain that can ultimately affect behavior. The current studies seek to understand the effects of low-dose alcohol (blood alcohol levels of ≤10mM). To do so, these experiments utilize a combination of behavioral and molecular techniques to (1) assess the ability of the interoceptive effects of a low dose of alcohol to gain control over goal-tracking behavior in a Pavlovian discrimination task, (2) determine brain regional differences in cellular activity via expression of immediate early genes (IEGs), and (3) assess the role of the dentate gyrus in modulating sensitivity to the interoceptive effects of a low dose of alcohol. Here, we show that intragastric administration of a dose of 0.8 g/kg alcohol produces blood alcohol levels ≤10mM in both male and female Long-Evans rats and can readily be trained as a Pavlovian interoceptive drug cue. In rats trained on this procedure, this dose of alcohol also modulates expression of the IEGs c-Fos and Arc in brain regions known to modulate expression of alcohol interoceptive effects. Finally, pharmacological inactivation of the dentate gyrus with GABA agonists baclofen and muscimol disrupted the ability of a low dose of alcohol to serve as an interoceptive cue. Together, these findings demonstrate behavioral and molecular consequences of low-dose alcohol.

Neonatal exposure to genistein affects reproductive physiology and behavior in female and male Long-Evans rats.

The present study was designed to examine the effects of neonatal genistein exposure on measures of reproductive physiology and behavior. Approximately 24 h after birth, female and male Long-Evans rat pups were injected daily with genistein (150 µg, subcutaneous; n = 29) or olive oil (n = 23) between postnatal days 1 and 5. After weaning, we examined all subjects daily until they reached puberty (i.e. vaginal opening in female rats and preputial separation in male rats). For all female subjects, we also examined vaginal cytology. After monitoring estrous cyclicity, the female subjects were given the opportunity to interact with a gonadally intact male or a sexually receptive female rat on the day of behavioral estrus to assess sexual motivation (i.e. partner-preference test with and without physical contact), which has never been evaluated before. For all male subjects, we assessed the development of copulatory behavior and sexual motivation (partner-preference test without physical contact). Consistent with previous findings, we found that neonatal exposure to genistein did not affect puberty onset in female or male rats. However, female rats exposed to genistein displayed significantly more irregular estrous cycles than controls. Neonatal genistein exposure also altered the development of male copulatory behavior, as indicated by an increase in mount frequency and intromission frequency and shorter interintromission intervals. We extended previous findings confirming that neither female nor male sexual motivation was affected by neonatal genistein. The results of the present study have important implications for the development of reproductive physiology and behavior in human neonates exposed to genistein in soy-based baby formula.

The synthetically produced predator odor 2,5-dihydro-2,4,5-trimethylthiazoline increases alcohol self-administration and alters basolateral amygdala response to alcohol in rats.

Post-traumatic stress disorder (PTSD) is a psychiatric illness that can increase the risk for developing an alcohol use disorder (AUD). While clinical data has been useful in identifying similarities in the neurobiological bases of these disorders, preclinical models are essential for understanding the mechanism(s) by which stressors increase the risk for escalated alcohol consumption. The purpose of these studies was to examine if exposure of male Long-Evans rats to the synthetically derived predator odor 2,5-dihydro-2,4,5-trimethylthiazoline (TMT; a component of fox feces) would increase sweetened alcohol self-administration, potentially by facilitating transfer of salience towards cues, and alter neuronal response to alcohol as measured by the immediate early gene c-Fos. In experiment 1, rats exposed to repeated (4×) TMT showed reductions in port entries in Pavlovian conditioned approach and increases in sweetened alcohol self-administration. In experiment 2, rats exposed to repeated TMT showed blunted basolateral amygdala c-Fos response to alcohol. In experiment 3, rats exposed to single, but not repeated TMT, showed increases in sweetened alcohol self-administration, and no change in anxiety-like behavior or hyperarousal. In experiment 4, rats continued to show a significant corticosterone response to TMT after repeated exposures. In summary, exposure of male rats to TMT can cause escalations in sweetened alcohol self-administration and reduction in BLA response to alcohol. These studies outline and utilize a novel preclinical model that can be used to further neurobiological understanding of the emergence of escalated alcohol consumption following stress exposure.

Glucagon-Like Peptide-1 Receptor Signaling in the Ventral Tegmental Area Reduces Alcohol Self-Administration in Male Rats.

The misuse and abuse of alcohol is a major public health issue. However, available treatments are limited with variable efficacy. Recently, preclinical studies show that glucagon-like-peptide-1 (GLP-1) and its analogue Exendin-4 (Ex4) potently reduce a range of alcohol intake behaviors, thus highlighting its potential as a treatment for alcohol use disorders. However, the neural mechanisms and sites of action mediating the effects of Ex4 on alcohol intake behaviors remain to be characterized. This study examined the ventral tegmental area (VTA) as a site of action for the effects of GLP-1 on alcohol intake. Male Long-Evans rats were given intermittent access to 20% alcohol and trained to nose poke for 20% alcohol. Rats received intra-VTA injections of Ex4 (vehicle, 0.01, 0.05μg), and the effects of VTA Ex4 on alcohol self-administration, motivation, and relapse were assessed. When compared to vehicle treatment, intra-VTA Ex4 (0.01, 0.05μg) delivery significantly reduced alcohol self-administration, an effect that was particularly prominent in high alcohol drinkers. However, VTA Ex4 did not reduce reacquisition of alcohol self-administration after extinction nor the motivation to obtain alcohol. Importantly, the lower dose of Ex4 (0.01μg) used had no effect on food intake or locomotor activity, suggesting that the reduction in alcohol self-administration observed was not secondary to caloric intake or motor deficits. Together, these findings provide support for the VTA as a key site of action for GLP-1 on alcohol self-administration but not the reacquisition of alcohol self-administration or motivation to work for alcohol.

Acquisition of remifentanil self-administration: Enhanced in female rats but no effect of adolescent stress exposure

Animal models of acquisition have been vital in shaping our understanding of vulnerability factors that influence susceptibility to drugs of abuse. Decades of research substantiates a number of biological, environmental, and behavioral factors that predict vulnerability – many of which have been important in the development of early intervention efforts in humans. The goal of the present study was to examine the acquisition of a synthetic opioid derivative in 66 adult male and female Long-Evans rats following histories of stress exposure during adolescence. Stress-exposed rats were subjected to a mild stress paradigm, which included alternating exposure to synthetic fox feces and physical restraint for eight days. Following stress induction and assessment, all rats were implanted with intravenous catheters in order to self-administer remifentanil (1 μm/kg/infusion) with no prior operant training. Acquisition of remifentanil self-administration was measured over 15 days. Findings indicate that regardless of stress condition, female rats acquired remifentanil self-administration sooner and emitted more active lever presses than males. Stress exposed animals exhibited increased anxiety-like response compared to the control group following exposure to stress, operationalized as decreased exploratory behavior on an Elevated Plus Maze. However, these effects were not expressed as significant differences in self-administration by stress. Together, these findings indicate that sex differences are evident in remifentanil self-administration.

Appetitive olfactory conditioning in the neonatal male rat facilitates subsequent sexual partner preference

Pairing a neutral odor with a male rat’s initial sexual experiences to ejaculation produces a subsequent conditioned ejaculatory preference (CEP) in which males ejaculate preferentially with receptive females that bear the odor relative to unscented receptive females. In 1986, Fillion and Blass reported that neonatal male rats exposed to a neutral lemon odor (citral) painted on their mother’s ventrum while nursing ejaculated faster as adults with sexually receptive, citral-scented females compared to unscented receptive females. The present study examined whether the same odor paired with tactile reward in neonatal male rats would alter the subsequent expression of a CEP. Newborn Long-Evans male rats were separated from their mothers each day beginning on Postnatal Day 1 and placed into a Plexiglas cage that contained either unscented or citral-scented bedding (N = 8/group). During each trial, rats were stroked from head to toe with a soft, narrow paintbrush, after which they were returned to their mothers. Males were weaned at 21 days of age and housed in same-treatment pairs for an intervening 50 days. Following habituation to a large open field, males were presented with two sexually receptive Long-Evans females, one scented with citral, and the other unscented, for a 30-min test of copulation. Males in the Paired group copulated and ejaculated preferentially with the scented female whereas males in the Unpaired group showed no preference. Pairing a neutral odor with a reward state in infancy generates a preference in male rats to ejaculate with sexually receptive females bearing the same odor in adulthood.

CaMKII antagonism in the ventral tegmental area impairs acquisition of conditioned approach learning in rats

This study investigated the role of calcium2+/calmodulin-dependent protein kinase II (CaMKII), a protein in the second messenger pathway of NMDA receptors, in the ventral tegmental area (VTA) in the acquisition and performance of conditioned approach learning. Male Long-Evans rats (N = 79) were exposed to 3 (to test acquisition) or 7 (to test performance) conditioning sessions in which they received 30 paired presentations of a light stimulus (CS) and a food pellet (US) on a random time schedule. These conditioning sessions were then followed by one 30-min session without the CS or US and lastly by a CS-only test session, where only the light stimulus was presented (without food) according to the same schedule as the conditioning sessions. Bilateral intra-VTA injections of KN93 (vehicle, 3.0, 4.5 or 6.0 μg/0.5 μL), a CaMKII inhibitor, were administered prior to each conditioning session to test effects on the acquisition of conditioned approach or prior to the CS-only test session to test effects on the performance of conditioned approach. KN93, when given prior to conditioning sessions, significantly reduced the number of conditioned approach responses emitted during CS presentations in the CS-only test. When KN93 was given prior to the CS-only test it had no effect. These results suggest that CaMKII activation in the VTA is necessary for the acquisition, but not the performance, of reward-related learning.