Developmental and behavioral effects in neonatal and adult mice following prenatal activation of endocannabinoid receptors by capsaicin.

Despite the apparent abundance of ligand-gated transient receptor potential vanilloid type 1 (TRPV1) and possible cross talk between the endocannabinoid and endovanilloid systems in the central nervous system (CNS), it is unclear what role TRPV1 receptor activation in CNS plays in neurobehavioral development. We previously reported that capsaicin or WIN55212-2 induces risk aversion in the plus-maze test, which was dependent on the gender and mouse strain used. In this study, pregnant BALBc mice were administered capsaicin (1.0 or 4.0 mg/kg, i.p.) during the second week of gestation. Developmental effects of prenatal exposure to capsaicin were assessed in neonates, and behavioral effects were assessed in adult offspring. Gender- and dose-specific variations in ultrasonic vocalizations, weight gain, righting reflex, and general activity of the pups were observed. Prenatal exposure to capsaicin altered plus-maze performance, especially with further exogenous capsaicin challenge. Furthermore, dose- and gender-specific effects were evident in the conditioned place preference/aversion paradigm following conditioning with capsaicin in adult animals. The capsaicin-induced aversion in the plus-maze test was enhanced by WIN55212-2 and blocked by pretreatment with vanilloid antagonist capsazepine or the CB1 receptor antagonist rimonabant, demonstrating an interaction between the endocannabinoid and endovanilloid systems in CNS. Taken together, the interaction between the endocannabinoid and endovanilloid signaling systems can be exploited for therapeutic applications in health and disease.

Behavioral effects of psychostimulants in mutant mice with cell-type specific deletion of CB2 cannabinoid receptors in dopamine neurons.

Activation of the endocannabinoid system modulate dopaminergic pathways that are involved in the effects of psychostimulants including amphetamine, cocaine, nicotine and other drugs of abuse. Genetic deletion or pharmacological activation of CB2 cannabinoid receptor is involved in the modulation of the effects of psychostimulants and their rewarding properties. Here we report on the behavioral effects of psychostimulants in DAT-Cnr2 conditional knockout (cKO) mice with selective deletion of type 2 cannabinoid receptors in dopamine neurons. There was enhanced psychostimulant induced hyperactivity in DAT-Cnr2 cKO mice, but the psychostimulant-induced sensitization was absent in DAT-Cnr2 cKO compared to the WT mice. Intriguingly, lower doses of amphetamine reduced locomotor activity of the DAT-Cnr2 cKO mice. While cocaine, amphetamine and methamphetamine produced robust conditioned place preference (CPP) in both DAT-Cnr2 cKO and WT mice, nicotine at the dose used induced CPP only in the WT but not in the DAT-Cn2 cKO mice. However, pre-treatment with the CB2R selective agonist JWH133, blocked cocaine and nicotine induced CPP in the WT mice. The deletion of CB2Rs in dopamine neurons modified the levels of tyrosine hydroxylase, and reduced the expression of dopamine transporter gene expression in DAT-Cnr2 cKO midbrain region. Taken together, our data suggest that CB2Rs play a role in the modulation of dopamine-related effects of psychostimulants and could be exploited as therapeutic target in psychostimulant addiction and other psychiatric disorders associated with dopamine dysregulation.

Behavioral Evaluation of Seeking and Preference of Alcohol in Mice Subjected to Stress.

The alcohol preference model is one of the most widely used animal models relevant to alcoholism. Stressors increase alcohol consumption. Here we present a protocol for a rapid and useful tool to test alcohol preference and stress-induced alcohol consumption in mice. In this model, animals are given two bottles, one with a diluted solution of ethanol in water, and the other with tap water. Consumption from each bottle is monitored over a 24-h period over several days to assess the animal's relative preference for the ethanol solution over water. In the second phase, animals are stressed by restraining them for an hour daily and their subsequent preference of tap water or the ethanol solution is evaluated. Preference is measured by the volume and/or weight or liquid consumed daily, which is then converted to a preference ratio. The alcohol preference model was combined with the conditioned place preference paradigm to determine alcohol conditioning and preference following the deletion of CB2 cannabinoid receptors in dopaminergic neurons in the DAT-Cnr2 Cre-recombinant conditional knockout (cKO) mice in comparison with the wild-type control mice.

Cannabinoid type 2 receptors in dopamine neurons inhibits psychomotor behaviors, alters anxiety, depression and alcohol preference.

Cannabinoid CB2 receptors (CB2Rs) are expressed in mouse brain dopamine (DA) neurons and are involved in several DA-related disorders. However, the cell type-specific mechanisms are unclear since the CB2R gene knockout mice are constitutive gene knockout. Therefore, we generated Cnr2-floxed mice that were crossed with DAT-Cre mice, in which Cre- recombinase expression is under dopamine transporter gene (DAT) promoter control to ablate Cnr2 gene in midbrain DA neurons of DAT-Cnr2 conditional knockout (cKO) mice. Using a novel sensitive RNAscope in situ hybridization, we detected CB2R mRNA expression in VTA DA neurons in wildtype and DAT-Cnr2 cKO heterozygous but not in the homozygous DAT-Cnr2 cKO mice. Here we report that the deletion of CB2Rs in dopamine neurons enhances motor activities, modulates anxiety and depression-like behaviors and reduces the rewarding properties of alcohol. Our data reveals that CB2Rs are involved in the tetrad assay induced by cannabinoids which had been associated with CB1R agonism. GWAS studies indicates that the CNR2 gene is associated with Parkinson's disease and substance use disorders. These results suggest that CB2Rs in dopaminergic neurons may play important roles in the modulation of psychomotor behaviors, anxiety, depression, and pain sensation and in the rewarding effects of alcohol and cocaine.

Dietary docosahexaenoic acid alleviates autistic-like behaviors resulting from maternal immune activation in mice.

The prevalence of autism spectrum disorders over the last several decades has risen at an alarming rate. Factors such as broadened clinical definitions and increased parental age only partially account for this precipitous increase, suggesting that recent changes in environmental factors may also be responsible. One such factor could be the dramatic decrease in consumption of anti-inflammatory dietary omega-3 (n-3) polyunsaturated fatty acids (PUFAs) relative to the amount of pro-inflammatory omega-6 (n-6) PUFAs and saturated fats in the Western diet. Docosahexaenoic acid (DHA) is the principle n-3 PUFA found in neural tissue and is important for optimal brain development, especially during late gestation when DHA rapidly and preferentially accumulates in the brain. In this study, we tested whether supplementation of a low n-3 PUFA diet with DHA throughout development could improve measures related to autism in a mouse model of maternal immune activation. We found that dietary DHA protected offspring from the deleterious effects of gestational exposure to the viral mimetic polyriboinosinic-polyribocytidilic acid on behavioral measures of autism and subsequent adulthood immune system reactivity. These data suggest that elevated dietary levels of DHA, especially during pregnancy and nursing, may help protect normal neurodevelopment from the potentially adverse consequences of environmental insults like maternal infection.

Exaggerated responses to stress in the BTBR T+tf/J mouse: an unusual behavioral phenotype.

This study shows that the BTBR T+tf/J mouse, a model for autism spectrum disorder (ASD), has increased levels of the stress hormone corticosterone, when compared to C57BL/6J mice. In addition, we have shown that tail suspension of the BTBR produces a heightened anxiety response in the elevated plus maze. These results suggest that the BTBR mouse is stressor-reactive exhibiting hormone responses that might predispose it to ASD.

Genetic and developmental influences on infant mouse ultrasonic calling. III. Patterns of inheritance in the calls of mice 3-9 days of age.

Infant mice produce ultrasonic calls that may elicit retrieval by adult mice. Age-related differences and genetic effects, such as additivity and directional dominance, have been found for most call characteristics at 3 days of age. Significant maternal effects have been reported for calling rate. However, little is known about how the influence of these genetic effects changes with age. This study explored developmental-genetic patterns of inheritance of seven ultrasonic call characteristics at ages 3-9 days, from groups of mice derived from a complete 4 x 4 diallel cross. The results indicate that additive variance contributes significantly to all characteristics for all ages. Maternal effects have a small effect on call characteristics. Dominance effects decrease with age for rate, range, and length of calls, suggesting less selective pressure toward higher rates, greater range, and longer calls as pups become more competent thermoregulators.

Prenatal cocaine exposure alters maternal retrieval behavior in mice.

Previous studies have demonstrated alterations in maternal retrieval behavior as a result of direct cocaine exposure. To establish the influence of prenatal cocaine exposure on pup retrieval, we exposed pups of three F1 genotypes by injecting their mothers (all C57BL/10J strain) with 20 mg/kg cocaine hydrochloride or saline subcutaneously on gestation days 7 to 17. When those pups became adults, control and exposed females were mated with males of the same genotype and tested for pup retrieval on postpartum days 4 and 5. Because ultrasonic calls are known to be elicitors of maternal retrieval behavior, the rate of ultrasonic calling was measured. Prenatal cocaine exposure exerted a significant effect upon retrieval latency on day 4. No relationship was found between genotype and retrieval latency.

The effects of cold, rotation, and genotype on the production of ultrasonic calls in infant mice.

There has been a revival of interest recently in the ultrasonic calls of infant rodents as investigators are using them to assess neurobehavioral development and animal models of anxiety. We compared the rates of ultrasonic calling of infant mice of two genotypes in two situations, cold and rotation. The subjects of study were 169 mouse pups from 29 litters and of two F1 genotypes, C57BL/10J x DBA/2J and C57BL/10J x SJL/J. Half of each litter was recorded in a cool situation for 20 seconds and the other half was recorded while rotating at 10 rpm for 20 seconds. All pups were recorded on days of age 2 to 8. Rotation elicited calling at about twice the rate as cool temperature on each day of age and on average across days; situation (cold or rotation) accounted for over 50% of the variation between litters. Genotype also altered call rate, and on some days situation and genotype interacted. In studies of neurobehavioral development and the effects of pharmacological agents on infant mice, it is particularly important to understand the roles of the stimuli and genotypes employed.