Escalated Alcohol Self-Administration and Sensitivity to Yohimbine-Induced Reinstatement in Alcohol Preferring Rats: Potential Role of Neurokinin-1 Receptors in the Amygdala.

Genetic factors significantly contribute to the risk for developing alcoholism. To study these factors and other associated phenotypes, rodent lines have been developed using selective breeding for high alcohol preference. One of these models, the alcohol preferring (P) rat, has been used in hundreds of preclinical studies over the last few decades. However, very few studies have examined relapse-like behavior in this rat strain. In this study, we used operant self-administration and yohimbine-induced reinstatement models to examine relapse-like behavior in P rats. Our previous work has demonstrated that P rats show increased expression of the neurokinin-1 receptor (NK1R) in the central nucleus of the amygdala (CeA), and this functionally contributes to escalated alcohol consumption in this strain. We hypothesized that P rats would show increased sensitivity to yohimbine-induced reinstatement that is also mediated by NK1R in the CeA. Using Fos staining, site-specific infusion of NK1R antagonist, and viral vector overexpression, we examined the influence of NK1R on the sensitivity to yohimbine-induced reinstatement of alcohol seeking. We found that P rats displayed increased sensitivity to yohimbine-induced reinstatement as well as increased neuronal activation in the CeA after yohimbine injection compared to the control Wistar strain. Intra-CeA infusion of NK1R antagonist attenuates yohimbine-induced reinstatement in P rats. Conversely, upregulation of NK1R within the CeA of Wistar rats increases alcohol consumption and sensitivity to yohimbine-induced reinstatement. These findings suggest that NK1R upregulation in the CeA contributes to multiple alcohol-related phenotypes in the P rat, including alcohol consumption and sensitivity to relapse.

The neurokinin-1 receptor mediates escalated alcohol intake induced by multiple drinking models.

We have previously demonstrated that the neurokinin-1 receptor (NK1R) is upregulated in the central nucleus of the amygdala of alcohol preferring (P) rats and that this receptor mediates escalated alcohol consumption in this strain. However, it is unclear if non-genetic models of escalated consumption are also mediated by NK1R signaling, and if so, what brain regions govern this effect. In the experiments presented here, we use two methods of inducing escalated alcohol intake in outbred Wistar rats: yohimbine pretreatment and intermittent alcohol access (Monday, Wednesday, and Friday availability; 20% alcohol). We found that escalated alcohol consumption induced by both yohimbine injection and intermittent access is attenuated by systemic administration of the NK1R antagonist L822429. Also, when compared to continuous alcohol access or access to water alone, NK1R expression was increased in the nucleus accumbens (NAC) and dorsal striatum, but not the amygdala. Escalated consumption induced by intermittent access was attenuated when the NK1R antagonist L822429 was infused directly into the dorsal striatum, but not when infused into the NAC. Taken together, these results suggest that NK1R upregulation contributes to escalated alcohol consumption that is induced by genetic selection, yohimbine injection, and intermittent access. However there is a dissociation between the regions involved in these behaviors with amygdalar upregulation contributing to genetic predisposition to escalated consumption and striatal upregulation driving escalation that is induced by environmental exposures.

Bidirectional relationship between alcohol intake and sensitivity to social defeat: association with Tacr1 and Avp expression.

While epidemiological studies show that alcohol abuse is often co-morbid with affective disorders, the causal direction of this association is unclear. We examined this relationship using mouse models including social defeat stress (SDS), social interaction (SI) and voluntary alcohol consumption. C57BL6/J mice exposed to SDS segregate into two subpopulations, those that express depressive-like phenotypes ('susceptible') and those that do not ('resilient'). First, we stratified SDS-exposed mice and measured their voluntary alcohol consumption. Next, we determined whether SI behavior in alcohol-naïve mice could predict alcohol intake. Finally, we assessed the effect of binge-like alcohol exposure on sensitivity to SDS. We quantified Tacr1 (neurokinin-1 receptor gene) and Avp (vasopressin peptide gene) mRNA in brain regions involved in depression, addiction and social behavior. We found that susceptible mice consumed more alcohol compared with resilient mice, suggesting that depression-like phenotypes associate with increased alcohol intake. Interestingly, we observed a negative correlation between SI and alcohol intake in stress- and alcohol-naïve mice, suggesting that individual differences in SI associate with alcohol preference. Finally, alcohol pre-treatment increased sensitivity to SDS, indicating that alcohol exposure alters sensitivity to social stress. Quantification of mRNA revealed that increased expression of Tacr1 and Avp generally associated with decreased SI and increased alcohol intake. C57BL6/J mice are an inbred strain; thus, it is likely that individual differences in behavior and gene expression are driven by epigenetic factors. Collectively, these results support a bidirectional relationship between alcohol exposure and susceptibility to stress that is associated with variations in neuropeptide expression.

Circulating Estradiol Regulates Brain-Derived Estradiol via Actions at GnRH Receptors to Impact Memory in Ovariectomized Rats.

Systemic estradiol treatment enhances hippocampus-dependent memory in ovariectomized rats. Although these enhancements are traditionally thought to be due to circulating estradiol, recent data suggest these changes are brought on by hippocampus-derived estradiol, the synthesis of which depends on gonadotropin-releasing hormone (GnRH) activity. The goal of the current work is to test the hypothesis that peripheral estradiol affects hippocampus-dependent memory through brain-derived estradiol regulated via hippocampal GnRH receptor activity. In the first experiment, intracerebroventricular infusion of letrozole, which prevents the synthesis of estradiol, blocked the ability of peripheral estradiol administration in ovariectomized rats to enhance hippocampus-dependent memory in a radial-maze task. In the second experiment, hippocampal infusion of antide, a long-lasting GnRH receptor antagonist, blocked the ability of peripheral estradiol administration in ovariectomized rats to enhance hippocampus-dependent memory. In the third experiment, hippocampal infusion of GnRH enhanced hippocampus-dependent memory, the effects of which were blocked by letrozole infusion. Results indicate that peripheral estradiol-induced enhancement of cognition is mediated by brain-derived estradiol via hippocampal GnRH receptor activity.

Antagonism of brain insulin-like growth factor-1 receptors blocks estradiol effects on memory and levels of hippocampal synaptic proteins in ovariectomized rats.

RATIONALE: Treatment with estradiol, the primary estrogen produced by the ovaries, enhances hippocampus-dependent spatial memory and increases levels of hippocampal synaptic proteins in ovariectomized rats. Increasing evidence indicates that the ability of estradiol to impact the brain and behavior is dependent upon its interaction with insulin-like growth factor-1 (IGF-1). OBJECTIVE: The goal of the current experiment was to test the hypothesis that the ability of estradiol to impact hippocampus-dependent memory and levels of hippocampal synaptic proteins is dependent on its interaction with IGF-1. METHODS: Adult rats were ovariectomized and implanted with estradiol or control capsules and trained on a radial-maze spatial memory task. After training, rats were implanted with intracerebroventricular cannulae attached to osmotic minipumps (flow rate 0.15 µl/h). Half of each hormone treatment group received continuous delivery of JB1 (300 µg/ml), an IGF-1 receptor antagonist, and half received delivery of aCSF vehicle. Rats were tested on trials in the radial-arm maze during which delays were imposed between the fourth and fifth arm choices. Hippocampal levels of synaptic proteins were measured by western blotting. RESULTS: Estradiol treatment resulted in significantly enhanced memory. JB1 blocked that enhancement. Estradiol treatment resulted in significantly increased hippocampal levels of postsynaptic density protein 95 (PSD-95), spinophilin, and synaptophysin. JB1 blocked the estradiol-induced increase of PSD-95 and spinophilin and attenuated the increase of synaptophysin. CONCLUSIONS: Results support a role for IGF-1 receptor activity in estradiol-induced enhancement of spatial memory that may be dependent on changes in synapse structure in the hippocampus brought upon by estradiol/IGF-1 interactions.

A role for hippocampal actin rearrangement in object placement memory in female rats.

Actin rearrangement, the polymerization of globular actin (G-actin) to filamentous actin, causes morphological changes in dendritic spines and is hypothesized to be a substrate of learning and memory. The ovarian hormone estradiol promotes hippocampal actin rearrangement and enhances performance on hippocampus-dependent tasks, including object placement memory. The goals of the current study were to determine a role for actin rearrangement and its regulatory pathway in object placement memory in female rats and to determine if estradiol impacts actin rearrangement in ovariectomized rats during the performance of the task. In an initial experiment, young adult Long-Evans rats were ovariectomized and implanted with capsules containing either cholesterol vehicle or estradiol. Bilateral intrahippocampal infusions of aCSF vehicle or the actin rearrangement inhibitor, latrunculin A, were administered 15 min prior to initiation of the object placement task. Latrunculin A dose-dependently impaired object placement memory. Estradiol had no impact on the ability of latrunculin A to affect performance. In a second experiment, rats were ovariectomized and received implants containing cholesterol or estradiol. Half of each hormone treatment group was exposed to the object placement memory task and half underwent control procedures. Immediately following completion of behavior, rats were euthanized and hippocampi removed. Western blotting was used to measure hippocampal levels of phosphorylated and total levels of a regulator of actin polymerization, the actin depolymerization factor cofilin. Exposure to the object placement memory task resulted in significant increases in phosphorylated levels of cofilin. Estradiol treatment had no impact on protein levels. These data support a role for hippocampal actin rearrangement and its regulatory proteins in object placement memory in female rats and suggest that chronic estradiol treatment does not impact hippocampal actin arrangement.