Exploring ketamine's reinforcement, cue-induced reinstatement, and nucleus accumbens cFos activation in male and female long evans rats.

Ketamine (KET), a non-competitive N-methyl-d-aspartate (NMDA) receptor antagonist, has rapid onset of antidepressant effects in Treatment-Resistant Depression patients and repeated infusions are required to sustain its antidepressant properties. However, KET is an addictive drug, and so more preclinical and clinical research is needed to assess the safety of recurring treatments in both sexes. Thus, the aim of this study was to investigate the reinforcing properties of various doses of KET (0-, 0.125-, 0.25-, 0.5 mg/kg/infusion) and assess KET's cue-induced reinstatement and neuronal activation in both sexes of Long Evans rats. Neuronal activation was assessed using the protein expression of the immediate early gene cFos in the nucleus accumbens (Nac), an important brain area implicated in reward, reinforcement and reinstatement to most drug-related cues. Our findings show that KET has reinforcing effects in both male and female rats, albeit exclusively at the highest two doses (0.25 and 0.5 mg/kg/infusion). Furthermore, we noted sex differences, particularly at the highest dose of ketamine, with female rats displaying a higher rate of self-administration. Interestingly, all groups that self-administered KET reinstated to drug-cues. Following drug cue-induced reinstatement test in rats exposed to KET (0.25 mg/kg/infusion) or saline, there was higher cFos protein expression in KET-treated animals compared to saline controls, and higher cFos expression in the core compared to the shell subregions of the Nac. As for reinstatement, there were no notable sex differences reported for cFos expression in the Nac. These findings reveal some sex and dose dependent effects in KET's reinforcing properties and that KET at all doses induced similar reinstatement in both sexes. This study also demonstrated that cues associated with ketamine induce comparable neuronal activation in the Nac of both male and female rats. This work warrants further research into the potential addictive properties of KET, especially when administered at lower doses which are now being used in the clinic for treating various psychopathologies.

Transcriptomic analysis of paternal behaviors in prairie voles.

BACKGROUND: The importance of fathers' engagement in care and its critical role in the offspring's cognitive and emotional development is now well established. Yet, little is known on the underlying neurobiology due to the lack of appropriate animal models. In the socially monogamous and bi-parental prairie vole (Microtus ochrogaster), while 60-80% of virgin males show spontaneous paternal behaviors (Paternal), others display pup-directed aggression (Attackers). Here we took advantage of this phenotypic dichotomy and used RNA-sequencing in three important brain areas to characterize gene expression associated with paternal behaviors of Paternal males and compare it to experienced Fathers and Mothers. RESULTS: While Paternal males displayed the same range and extent of paternal behaviors as experienced Fathers, we observed structure-specific transcriptomic differences between parental behaviors phenotypes. Using differential expression, gene set expression, as well as co-expression network analyses, we found that phenotypic differences between Paternal males and Attackers were mainly reflected by the lateral septum (LS), and to a lower extent, the nucleus accumbens (NAc), transcriptomes. In the medial preoptic area (MPOA), the profiles of gene expression mainly reflected differences between females and males regardless of their parental behaviors phenotype. Functional enrichment analyses of those gene sets associated with Paternal males or Attackers in the LS and the NAc revealed the involvement of processes related to the mitochondria, RNA translation, protein degradation processes, as well as epigenetic regulation of gene expression. CONCLUSIONS: By leveraging the natural phenotypic differences in parental behaviors in virgin male prairie voles alongside fathers and mothers, we identified a marked structure- and phenotype-specific pattern of gene expression associated with spontaneous paternal behaviors independently from fatherhood and pair-bonding. The LS transcriptome related to the mitochondria, RNA translation, and protein degradation processes was thus highlighted as a primary candidate associated with the spontaneous display of paternal behaviors. Altogether, our observations further characterize the behavioral and transcriptomic signature of parental behaviors in the socially monogamous prairie vole and lay the groundwork to further our understanding of the molecular underpinnings of paternal behavior.

Sex- and estrous-cycle dependent dorsal hippocampal phosphoproteomic changes induced by low-dose ketamine.

Numerous emotional and cognitive processes mediated by the hippocampus present differences between sexes and can be markedly influenced by hormonal status in males and females of several species. In rodents, the dorsal hippocampus (dHPC) is known to contribute to the rapid antidepressant actions of the NMDA receptor antagonist ketamine. We and others have demonstrated a greater sensitivity to the fast-acting antidepressant ketamine in female versus male rats that is estrogen- and progesterone-dependent. However, the underlying mechanisms remain unclear. Using an acute low dose (2.5 mg/kg) of ketamine that is behaviorally effective in female but not male rats, a label-free phosphoproteomics approach was employed to identify ketamine-induced changes in signaling pathway activation and phosphoprotein abundance within the dHPC of intact adult male rats and female rats in either diestrus or proestrus. At baseline, males and females showed striking dissimilarities in the dHPC proteome and phosphoproteome related to synaptic signaling and mitochondrial function-differences also strongly influenced by cycle stage in female rats. Notably, phosphoproteins enriched in PKA signaling emerged as being both significantly sex-dependent at baseline and also the primary target of ketamine-induced protein phosphorylation selectively in female rats, regardless of cycle stage. Reduced phosphoprotein abundance within this pathway was observed in males, suggesting bi-directional effects of low-dose ketamine between sexes. These findings present biological sex and hormonal milieu as critical modulators of ketamine's rapid actions within this brain region and provide greater insight into potential translational and post-translational processes underlying sex- and hormone-dependent modulation of ketamine's therapeutic effects.

Sex- and dose-dependent abuse liability of repeated subanesthetic ketamine in rats.

RATIONALE: Subanesthetic ketamine (KET) elicits rapid, robust, but transient antidepressant effects. KET's antidepressant actions can be augmented and maintained for a longer duration when repeatedly delivered. However, KET is recreationally abused, raising long-term treatment safety concerns. Women are more likely than men to seek treatment for depression, escalate from casual to compulsive drug use, and are more sensitive to antidepressants. Similarly, female rodents are more sensitive than males to KET's rapid antidepressant-like behavioral effects; dose-response thresholds in these assays equal 2.5 and 5.0mg/kg (i.p.), respectively. This suggests the utility of preclinical rodent models in optimizing sex-differential KET therapy protocols and minimizing adverse drug reactions. OBJECTIVES: Here, we assessed behavioral and biochemical correlates of abuse liability following six serial KET treatments on alternating days at three subanesthetic, antidepressant-like doses (2.5, 5.0, or 10mg/kg, i.p.) in adult male and female rats. A potential role for ΔFosB-mediated transcription in the nucleus accumbens is outlined in the context of KET-mediated locomotor sensitization. RESULTS: Antidepressant-like threshold doses (2.5, 5.0mg/kg KET) failed to evoke a conditioned place preference in all animals, but only males positively responded to a higher dose (10mg/kg). Behavioral sensitization to 5.0 or 10mg/kg KET's locomotor-activating effects was established in both sexes, and females' sensitized response to 5.0mg/kg was greater than males'. KET-induced hyperlocomotion positively correlated with ΔFosB protein expression in the nucleus accumbens. rAAV-ΔJunD inhibition of ΔFosB-mediated transcription in the accumbens failed to block locomotor sensitization to 10mg/kg KET. CONCLUSIONS: These data suggest that in rats, six alternating-day treatments with 2.5mg/kg KET do not induce apparent behavioral signatures of abuse liability despite accumulation of ΔFosB protein in the accumbens. Additionally, females are more sensitive than males to KET's locomotor-stimulant properties, both acutely and after repeated treatments. More studies are needed to determine brain regions and neural mechanisms responsible for KET-induced behavioral adaptations and to extrapolate these data to inform sex-dependent strategies for long-term KET therapy protocols for depression.

Sex Differences in the Pharmacokinetics of Low-dose Ketamine in Plasma and Brain of Male and Female Rats.

Recent work from our group and others has revealed a higher sensitivity of female rodents to the antidepressant-like effects of the N-methyl d-aspartate receptor antagonist ketamine strongly influenced by circulating estrogen and progesterone levels. However, in the absence of any preclinical studies of pharmacokinetic sex differences using low-dose ketamine in rats, it is unclear whether the effects of sex and hormonal milieu on ketamine's behavioral actions are influenced by differences in ketamine metabolism between male and female rats. Therefore, this work examined whether sex and hormonal status affect ketamine metabolism and distribution in male and female rats using a low antidepressant-like dose selectively effective in females. Intact male rats and female rats in either diestrus (low estrogen, progesterone) or proestrus (high estrogen, progesterone) were administered low-dose ketamine, and their plasma and brains were collected to analyze levels of ketamine and its metabolites norketamine (NK) and dehydronorketamine. Females exhibited greater concentrations of ketamine and NK over the first 30 min following treatment in both brain and plasma, largely accounted for by slower clearance rates and longer half-lives. Interestingly, despite the impact of ovarian hormones on behavioral sensitivity to ketamine, no appreciable differences in pharmacokinetic parameters existed between proestrus and diestrus female rats. This work is the first to demonstrate sex differences in ketamine pharmacokinetics in rats, and suggests that while sex differences in metabolism may influence the amount of ketamine and NK reaching target areas in the brain, the impact of circulating hormone levels here is negligible.

Integrative analysis of sex differences in the rapid antidepressant effects of ketamine in preclinical models for individualized clinical outcomes.

In major depressive disorder, women exhibit higher lifetime prevalence and different antidepressant response rates than men, which illustrates the importance of examining individual differences in the pathophysiology of depression and therapeutic response. In recent years, the consideration of sex in related preclinical research has thus gained interest-particularly in light of novel evidence for rapid-acting antidepressants. Notably, the literature recently revealed a higher sensitivity of females to the antidepressant effects of the N-methyl-D-aspartate receptor antagonist ketamine, in both baseline and preclinical conditions. Combined with its fast-acting and relatively sustained properties, this evidence highlights ketamine as a particularly interesting therapeutic alternative for this sensitive population, and supports the value in considering sex as a critical factor for improved individualized therapeutic strategies.

Hedonic sensitivity to low-dose ketamine is modulated by gonadal hormones in a sex-dependent manner.

We recently reported a greater sensitivity of female rats to rapid antidepressant-like effects of ketamine compared to male rats, and that ovarian-derived estradiol (E2) and progesterone (P4) are essential for this response. However, to what extent testosterone may also contribute, and whether duration of response to ketamine is modulated in a sex- and hormone-dependent manner remains unclear. To explore this, we systematically investigated the influence of testosterone, estradiol and progesterone on initiation and maintenance of hedonic response to low-dose ketamine (2.5 mg/kg) in intact and gonadectomized male and female rats. Ketamine induced a sustained increase in sucrose preference of female, but not male, rats in an E2P4-dependent manner. Whereas testosterone failed to alter male treatment response, concurrent administration of P4 alone in intact males enhanced hedonic response low-dose ketamine. Treatment responsiveness in female rats only was associated with greater hippocampal BDNF levels, but not activation of key downstream signaling effectors. We provide novel evidence supporting activational roles for ovarian-, but not testicular-, derived hormones in mediating hedonic sensitivity to low-dose ketamine in female and male rats, respectively. Organizational differences may, in part, account for the persistence of sex differences following gonadectomy and selective involvement of BDNF in treatment response.

The Anxiolytic and Antidepressant-like Effects of Testosterone and Estrogen in Gonadectomized Male Rats.

BACKGROUND: While the influence of testosterone levels on vulnerability to affective disorders is not straightforward, research suggests this hormone may confer some degree of resiliency in men. We recently demonstrated a role for the dentate gyrus in mediating testosterone's protective effects on depressive-like behavior in gonadectomized male rats. Here, testosterone may exert its effects through androgen receptor-mediated mechanisms or via local aromatization to estradiol. METHODS: Gonadectomized male rats were implanted with a placebo, testosterone, or estradiol pellet, and subsequent protective anxiolytic- and antidepressant-like effects of testosterone and its aromatized metabolite, estradiol, were then investigated in the open field and sucrose preference tests, respectively. Moreover, their influence on gene expression in the hippocampus was analyzed by genome-wide complementary DNA microarray analysis. Finally, the contribution of testosterone's aromatization within the dentate gyrus was assessed by local infusion of the aromatase inhibitor fadrozole, whose efficacy was confirmed by liquid chromatography-tandem mass spectrometry. RESULTS: Both hormones had antidepressant-like effects associated with a substantial overlap in transcriptional regulation, particularly in synaptic plasticity- and mitogen-activated protein kinase pathway-related genes. Further, chronic aromatase inhibition within the dentate gyrus blocked the protective effects of testosterone. CONCLUSIONS: Both testosterone and estradiol exhibit anxiolytic- and antidepressant-like effects in gonadectomized male rats, while similarly regulating critical mediators of these behaviors, suggesting common underlying mechanisms. Accordingly, we demonstrated that testosterone's protective effects are mediated, in part, by its aromatization in the dentate gyrus. These findings thus provide further insight into a role for estradiol in mediating the protective anxiolytic- and antidepressant-like effects of testosterone.

Selective phosphodiesterase inhibitors improve performance on the ED/ID cognitive task in rats.

A number of selective phosphodiesterase (PDE) inhibitors have been demonstrated to improve learning in several rodent models of cognition. Given that schizophrenia is associated with impairments in frontal lobe-dependent cognitive functions (e.g., working memory and cognitive flexibility), we examined whether PDE inhibitors would attenuate cognitive deficits associated with schizophrenia. Persistent suppression of N-methyl-D-aspartate (NMDA) receptor function produces enduring structural changes in neocortical and limbic regions in a pattern similar to changes reported in schizophrenia. This similarity suggests that subchronic treatment with NMDA receptor antagonists (e.g., phencyclidine, PCP) may represent a useful preclinical model of neurobiological and related cognitive deficits associated with schizophrenia. We treated male Long-Evans rats with subchronic PCP (5 mg/kg, ip, BID, 7 d) or saline and then examined the effects of acute treatment with selected doses of PDE inhibitors that have been demonstrated to regulate both intracellular levels of cAMP and/or cGMP, and to improve cognitive function. We used an extradimensional-intradimensional (ED/ID) test of cognitive flexibility similar to those used in humans and non-human primates for assessing executive function. Subchronic treatment with PCP produced a selective impairment on ED shift (EDS) performance without significant impairment on any other discrimination problem when compared to saline-treated control animals. Selected doses of the four PDEIs evaluated (PDE2: BAY 60-7550; PDE4: rolipram; PDE5: sildenafil; PDE10A: papaverine) were able to significantly attenuate this cognitive deficit in EDS performance. This suggests that this rodent model of executive function was sensitive to pro-cognitive effects of intracellular effects resulting from PDE inhibition. Together, these data suggest that inhibition of PDE activity may represent valuable therapeutic targets to improve cognition associated with neuropsychiatric disorders that feature cognitive dysfunction as a key symptom.

Environmental enrichment ameliorates phencyclidine-induced cognitive deficits.

Recent work has suggested that environmental enrichment during development can enhance aspects of learning and memory, however its effects on executive function and cognitive flexibility have not been well studied. The goal of this research was to evaluate whether environmental enrichment (EE) that included wheel running would improve cognitive performance in young male Long Evans rats that received subchronic administration of either phencyclidine (PCP) or saline. We used a sensitive extradimensional/intradimensional (ED/ID) test of cognitive flexibility similar to that used in humans and nonhuman primates for assessing executive function. PCP-treated rats demonstrated a selective impairment on ED shift (EDS) performance without significant impairment on other discrimination problems when compared to saline treated control animals. A separate group of animals that received PCP + EE demonstrated significantly improved performance on EDS and reversal learning problems, whereas the saline + EE group demonstrated a non-selective improvement in overall performance when compared to non-enriched saline controls. The saline + EE group demonstrated greater activity levels as measured by wheel running when compared to the PCP + EE group, but no significant associations were found between wheel running and cognitive performance. Together, these data suggest that EE that features wheel running may have promoted a general cognitive enhancement while also selectively acting upon neurobiological mechanisms that subserve executive function and cognitive flexibility in impaired animals. Development of novel treatment methodologies that target mechanisms underlying the ameliorative effects of EE in this model of cognitive impairment may be a useful tool in the development of new therapeutic strategies for disorders that feature cognitive dysfunction as a key symptom.

AM 251 differentially effects food-maintained responding depending on food palatability.

Ligands functioning as antagonists and inverse agonists at the cannabinoid CB(1)-receptor (e.g., AM 251, AM 281, and rimonabant (previously identified as SR141716)) have been demonstrated to have effects on satiety, consumption of, and the motivation to work for, or obtain food. These represent behavioral effects that may also be linked to characteristics such as food palatability or motivation to obtain food. Given the recent removal of rimonabant from clinical trials, a thorough characterization of ingestive behaviors that are associated with other likely candidate drugs is warranted. In the present study, normal weight male Long Evans rats were trained to respond for grain or chocolate-flavored food pellets under progressive-ratio schedules of reinforcement. Rats received acute injections of the CB(1) receptor antagonist AM 251 (0.3-3.0 mg/kg) or vehicle prior to daily testing sessions. Administration of AM 251 produced significant dose-dependent reductions in responding for, deliveries of, and break points (BP) associated with chocolate-flavored but not grain pellets. These data add to the literature demonstrating the ability of CB(1) antagonists to selectively reduce motivation to obtain highly palatable reinforcers.