Regulation of Cortico-Thalamic JNK1/2 and ERK1/2 MAPKs and Apoptosis-Related Signaling Pathways in PDYN Gene-Deficient Mice Following Acute and Chronic Mild Stress.

The crosstalk between the opioidergic system and mitogen-activated protein kinases (MAPKs) has a critical role in mediating stress-induced behaviors related to the pathophysiology of anxiety. The present study evaluated the basal status and stress-induced alterations of cortico-thalamic MAPKs and other cell fate-related signaling pathways potentially underlying the anxiogenic endophenotype of PDYN gene-deficient mice. Compared to littermates, PDYN knockout (KO) mice had lower cortical and or thalamic amounts of the phospho-activated MAPKs c-Jun N-terminal kinase (JNK1/2) and extracellular signal-regulated kinase (ERK1/2). Similarly, PDYN-KO animals displayed reduced cortico-thalamic densities of total and phosphorylated (at Ser191) species of the cell fate regulator Fas-associated protein with death domain (FADD) without alterations in the Fas receptor. Exposure to acute restraint and chronic mild stress stimuli induced the robust stimulation of JNK1/2 and ERK1/2 MAPKs, FADD, and Akt-mTOR pathways, without apparent increases in apoptotic rates. Interestingly, PDYN deficiency prevented stress-induced JNK1/2 and FADD but not ERK1/2 or Akt-mTOR hyperactivations. These findings suggest that cortico-thalamic MAPK- and FADD-dependent neuroplasticity might be altered in PDYN-KO mice. In addition, the results also indicate that the PDYN gene (and hence dynorphin release) may be required to stimulate JNK1/2 and FADD (but not ERK1/2 or Akt/mTOR) pathways under environmental stress conditions.

Role of Cannabinoid CB2 Receptor in Alcohol Use Disorders: From Animal to Human Studies.

Cumulative evidence has pointed out cannabinoid CB2 receptors (CB2r) as a potential therapeutic key target for treating alcohol use disorder (AUD). This review provides the most relevant results obtained from rodent and human studies, including an integrative section focused on the involvement of CB2r in the neurobiology of alcohol addiction. A literature search was conducted using the electronic databases Medline and Scopus for articles. The search strategy was as follows: "Receptor, Cannabinoid, CB2" AND "Alcohol-Related Disorders" AND "human/or patients"; "Receptor, Cannabinoid, CB2" AND "Alcohol" OR "Ethanol" AND "rodents/or mice/or rats". Pharmacological approaches demonstrated that the activation or blockade of CB2r modulated different alcohol-addictive behaviors. Rodent models of alcoholism revealed significant alterations of CB2r in brain areas of the reward system. In addition, mice lacking CB2r (CB2KO) show increased alcohol consumption, motivation, and relapse alterations. It has been stressed that the potential neurobiological mechanisms underlying their behavioral effects involve critical elements of the alcohol reward system. Interestingly, recent postmortem studies showed CNR2 alterations in brain areas of alcoholic patients. Moreover, although the number of studies is limited, the results revealed an association between some genetic alterations of the CNR2 and an increased risk for developing AUD. This review provides evidence that CB2r may play a role in alcohol addiction. Clinical studies are necessary to figure out whether CB2r ligands may prove useful for the treatment of AUD in humans.

Immunomodulatory Role of CB2 Receptors in Emotional and Cognitive Disorders.

Emotional behavior, memory, and learning have been associated with alterations in the immune system in neuropsychiatric and neurodegenerative diseases. In recent years, several studies pointed out the involvement of the cannabinoid receptor 2 (CB2r) in the immune system and the regulation of inflammation. This receptor is widely distributed in different tissues and organs with higher expression in spleen and immune system cells. However, CB2r has also been detected in several brain areas and different brain cell types, such as neurons and glia. These findings suggest that CB2r may closely relate the immune system and the brain circuits regulating inflammation, mood, and cognitive functions. Therefore, we review the studies that may help elucidate the molecular bases of CB2r in regulating inflammation in different brain cells and its role in the pathophysiology of psychiatric and neurodegenerative disorders.

Biomarkers of the Endocannabinoid System in Substance Use Disorders.

Despite substance use disorders (SUD) being one of the leading causes of disability and mortality globally, available therapeutic approaches remain ineffective. The difficulty in accurately characterizing the neurobiological mechanisms involved with a purely qualitative diagnosis is an obstacle to improving the classification and treatment of SUD. In this regard, identifying central and peripheral biomarkers is essential to diagnosing the severity of drug dependence, monitoring therapeutic efficacy, predicting treatment response, and enhancing the development of safer and more effective pharmacological tools. In recent years, the crucial role that the endocannabinoid system (ECS) plays in regulating the reinforcing and motivational properties of drugs of abuse has been described. This has led to studies characterizing ECS alterations after exposure to various substances to identify biomarkers with potential diagnostic, prognostic, or therapeutic utility. This review aims to compile the primary evidence available from rodent and clinical studies on how the ECS components are modified in the context of different substance-related disorders, gathering data from genetic, molecular, functional, and neuroimaging experimental approaches. Finally, this report concludes that additional translational research is needed to further characterize the modifications of the ECS in the context of SUD, and their potential usefulness in the necessary search for biomarkers.

Inflammatory Biomarkers in Addictive Disorders.

Substance use disorders are a group of diseases that are associated with social, professional, and family impairment and that represent a high socio-economic impact on the health systems of countries around the world. These disorders present a very complex diagnosis and treatment regimen due to the lack of suitable biomarkers supporting the correct diagnosis and classification and the difficulty of selecting effective therapies. Over the last few years, several studies have pointed out that these addictive disorders are associated with systemic and central nervous system inflammation, which could play a relevant role in the onset and progression of these diseases. Therefore, identifying different immune system components as biomarkers of such addictive disorders could be a crucial step to promote appropriate diagnosis and treatment. Thus, this work aims to provide an overview of the immune system alterations that may be biomarkers of various addictive disorders.

Cannabis Use in Pregnant and Breastfeeding Women: Behavioral and Neurobiological Consequences.

Nowadays, cannabis is the most consumed illicit drug. The global prevalence of the use of cannabis in 2017 was estimated in 188 million of people, 3.8% of worldwide population. Importantly, the legalization of cannabis in different countries, together with the increase in the apparent safety perception, may result in a great variety of health problems. Indeed, an important concern is the increase in cannabis use among pregnant and breastfeeding women, especially since the content of delta9-tetrahidrocannabinol (THC) is currently around 2-fold higher than it was 15-20 years ago. The purpose of this study was to review cannabis use during pregnancy and breastfeeding including epidemiological aspects, therapeutic or preventive strategies, and experimental considerations and results from animal models of perinatal cannabis exposure to analyze the underlying neurobiological mechanisms and to identify new therapeutic approaches. A recent report revealed that among pregnant women aged 15-44, last month cannabis use prevalence was over 4.9%, raising to 8.5% in the 18-25-year-old age range. Pre- and post-natal exposure to cannabis may be associated with critical alterations in the newborn infants that are prolonged throughout childhood and adolescence. Briefly, several reports revealed that perinatal cannabis exposure was associated with low birth weight, reduction in the head circumference, cognitive deficits (attention, learning, and memory), disturbances in emotional response leading to aggressiveness, high impulsivity, or affective disorders, and higher risk to develop a substance use disorder. Furthermore, important neurobiological alterations in different neuromodulatory and neurotransmission systems have been associated with cannabis consumption during pregnancy and lactation. In spite of the evidences pointing out the negative behavioral and neurobiological consequences of cannabis use in pregnant and breastfeeding women, there are still limitations to identify biomarkers that could help to establish preventive or therapeutic approaches. It is difficult to define the direct association specifically with cannabis, avoiding other confusing factors, co-occurrence of other drugs consumption (mainly nicotine and alcohol), lifestyle, or socioeconomic factors. Therefore, it is necessary to progress in the characterization of short- and long-term cannabis exposure-related disturbances.

Association of cannabinoid receptor genes (CNR1 and CNR2) polymorphisms and panic disorder.

Background and objectives: Panic disorder (PD) is an anxiety disorder characterized by recurrent and unexpected panic attacks along with sudden onset of apprehension, fear or terror. The endocannabinoid system (ECS) has a role in stress recovery, regulating anxiety. The aim of this study was to analyze potential genetic alterations in key ECS targets in patients suffering from panic disorders.Design and methods: We analyzed single nucleotide polymorphisms (SNPs) of the cannabinoid receptors (CNR1; CNR2) and the endocannabinoid hydrolytic enzyme fatty acid amide hydrolase (FAAH) genes in 164 Spanish PD patients and 320 matched controls.Results: No significant differences were observed in the SNPs of the CNR2 and FAAH genes tested. However, when analyzing genotype-by-sex interaction at A592G (rs2501431) and C315T (rs2501432) in the CNR2 gene, the presence of the G-allele in males was associated with a protective haplotype. Genotyping analysis revealed that variants in CNR1 confer vulnerability to PD, with a significantly increased risk associated with the G-allele (rs12720071) and C-allele (rs806368). This finding was consistent when analyzing genotype-by-sex interaction, where females presented a greater PD risk.Conclusions: Polymorphisms at the CNR1 gene may be a risk factor for PD contributing to sex-specific dysfunction in females.

Slc7a11 (xCT) protein expression is not altered in the depressed brain and system xc- deficiency does not affect depression-associated behaviour in the corticosterone mouse model.

Objectives: The cystine/glutamate antiporter (system xc-) is believed to contribute to nonvesicular glutamate release from glial cells in various brain areas. Although recent investigations implicate system xc- in mood disorders, unambiguous evidence has not yet been established. Therefore, we evaluated the possible role of system xc- in the depressive state. Methods: We conducted a protein expression analysis of the specific subunit of system xc- (xCT) in brain regions of the corticosterone mouse model, Flinders Sensitive Line rat model and post-mortem tissue of depressed patients. We next subjected system xc- deficient mice to the corticosterone model and analysed their behaviour in several tests. Lastly, we subjected additional cohorts of xCT-deficient and wild-type mice to N-acetylcysteine treatment to unveil whether the previously reported antidepressant-like effects are dependent upon system xc-. Results: We did not detect any changes in xCT expression levels in the animal models or patients compared to proper controls. Furthermore, loss of system xc- had no effect on depression- and anxiety-like behaviour. Finally, the antidepressant-like effects of N-acetylcysteine are not mediated via system xc-. Conclusions: xCT protein expression is not altered in the depressed brain and system xc- deficiency does not affect depression-associated behaviour in the corticosterone mouse model.

Disrupted Neuroglial Metabolic Coupling after Peripheral Surgery.

Immune-related events in the periphery can remotely affect brain function, contributing to neurodegenerative processes and cognitive decline. In mice, peripheral surgery induces a systemic inflammatory response associated with changes in hippocampal synaptic plasticity and transient cognitive decline, however, the underlying mechanisms remain unknown. Here we investigated the effect of peripheral surgery on neuronal-glial function within hippocampal neuronal circuits of relevance to cognitive processing in male mice at 6, 24, and 72 h postsurgery. At 6 h we detect the proinflammatory cytokine IL-6 in the hippocampus, followed up by alterations in the mRNA and protein expression of astrocytic and neuronal proteins necessary for optimal energy supply to the brain and for the reuptake and recycling of glutamate in the synapse. Similarly, at 24 h postsurgery the mRNA expression of structural proteins (GFAP and AQP4) was compromised. At this time point, functional analysis in astrocytes revealed a decrease in resting calcium signaling. Examination of neuronal activity by whole-cell patch-clamp shows elevated levels of glutamatergic transmission and changes in AMPA receptor subunit composition at 72 h postsurgery. Finally, lactate, an essential energy substrate produced by astrocytes and critical for memory formation, decreases at 6 and 72 h after surgery. Based on temporal parallels with our previous studies, we propose that the previously reported cognitive decline observed at 72 h postsurgery in mice might be the consequence of temporal hippocampal metabolic, structural, and functional changes in astrocytes that lead to a disruption of the neuroglial metabolic coupling and consequently to a neuronal dysfunction.SIGNIFICANCE STATEMENT A growing body of evidence suggests that surgical trauma launches a systemic inflammatory response that reaches the brain and associates with immune activation and cognitive decline. Understanding the mechanisms by which immune-related events in the periphery can influence brain processes is essential for the development of therapies to prevent or treat postoperative cognitive dysfunction and other forms of cognitive decline related to immune-to-brain communication, such as Alzheimer's and Parkinson's diseases. Here we describe the temporal orchestration of a series of metabolic, structural, and functional changes after aseptic trauma in mice related to astrocytes and later in neurons that emphasize the role of astrocytes as key intermediaries between peripheral immune events, neuronal processing, and potentially cognition.

Running Opposes the Effects of Social Isolation on Synaptic Plasticity and Transmission in a Rat Model of Depression.

Stress, such as social isolation, is a well-known risk factor for depression, most probably in combination with predisposing genetic factors. Physical exercise on the other hand, is depicted as a wonder-treatment that makes you healthier, happier and live longer. However, the published results on the effects of exercise are ambiguous, especially when it comes to neuropsychiatric disorders. Here we combine a paradigm of social isolation with a genetic rat model of depression, the Flinders Sensitive Line (FSL), already known to have glutamatergic synaptic alterations. Compared to group-housed FSL rats, we found that social isolation further affects synaptic plasticity and increases basal synaptic transmission in hippocampal CA1 pyramidal neurons. These functional synaptic alterations co-exist with changes in hippocampal protein expression levels: social isolation in FSL rats reduce expression of the glial glutamate transporter GLT-1, and increase expression of the GluA2 AMPA-receptor subunit. We further show that physical exercise in form of voluntary running prevents the stress-induced synaptic effects but do not restore the endogenous mechanisms of depression already present in the FSL rat.

Hippocampal-Dependent Antidepressant Action of the H3 Receptor Antagonist Clobenpropit in a Rat Model of Depression.

BACKGROUND: Histamine is a modulatory neurotransmitter regulating neuronal activity. Antidepressant drugs target modulatory neurotransmitters, thus ultimately regulating glutamatergic transmission and plasticity. Histamine H3 receptor (H3R) antagonists have both pro-cognitive and antidepressant effects; however, the mechanism by which they modulate glutamate transmission is not clear. We measured the effects of the H3R antagonist clobenpropit in the Flinders Sensitive Line (FSL), a rat model of depression with impaired memory and altered glutamatergic transmission. METHODS: Behavioral tests included the forced swim test, memory tasks (passive avoidance, novel object recognition tests), and anxiety-related paradigms (novelty suppressed feeding, social interaction, light/dark box tests). Hippocampal protein levels were detected by Western blot. Hippocampal plasticity was studied by in slice field recording of CA3-CA1 long-term synaptic potentiation (LTP), and glutamatergic transmission by whole-cell patch clamp recording of excitatory postsynaptic currents (EPSCs) in CA1 pyramidal neurons. RESULTS: Clobenpropit, administered systemically or directly into the hippocampus, decreased immobility during the forced swim test; systemic injections reversed memory deficits and increased hippocampal GluN2A protein levels. FSL rats displayed anxiety-related behaviors not affected by clobenpropit treatment. Clobenpropit enhanced hippocampal plasticity, but did not affect EPSCs. H1R and H2R antagonists prevented the clobenpropit-induced increase in LTP and, injected locally into the hippocampus, blocked clobenpropit's effect in the forced swim test. CONCLUSIONS: Clobenpropit's antidepressant effects and the enhanced synaptic plasticity require hippocampal H1R and H2R activation, suggesting that clobenpropit acts through disinhibition of histamine release. Clobenpropit reverses memory deficits and increases hippocampal GluN2A expression without modifying anxiety-related phenotypes or EPSCs in CA1 pyramidal neurons.

Skeletal muscle PGC-1α1 modulates kynurenine metabolism and mediates resilience to stress-induced depression.

Depression is a debilitating condition with a profound impact on quality of life for millions of people worldwide. Physical exercise is used as a treatment strategy for many patients, but the mechanisms that underlie its beneficial effects remain unknown. Here, we describe a mechanism by which skeletal muscle PGC-1α1 induced by exercise training changes kynurenine metabolism and protects from stress-induced depression. Activation of the PGC-1α1-PPARα/δ pathway increases skeletal muscle expression of kynurenine aminotransferases, thus enhancing the conversion of kynurenine into kynurenic acid, a metabolite unable to cross the blood-brain barrier. Reducing plasma kynurenine protects the brain from stress-induced changes associated with depression and renders skeletal muscle-specific PGC-1α1 transgenic mice resistant to depression induced by chronic mild stress or direct kynurenine administration. This study opens therapeutic avenues for the treatment of depression by targeting the PGC-1α1-PPAR axis in skeletal muscle, without the need to cross the blood-brain barrier.

Hippocampal and prefrontal dopamine D1/5 receptor involvement in the memory-enhancing effect of reboxetine.

Dopamine modulates cognitive functions through regulation of synaptic transmission and plasticity in the hippocampus and prefrontal cortex (PFC). Thus, dopamine dysfunction in depression may be particularly relevant for the cognitive symptoms. The norepinephrine transporter inhibitor reboxetine facilitates memory processing in both healthy volunteers and in depressed patients and increases dopamine release in both the hippocampus and PFC. We investigated the potential involvement of the hippocampal and PFC dopamine D1/5 receptors in the cognitive effects of reboxetine using the object recognition test in rats. Infusion of the D1/5 antagonist SCH23390 into the dorsal hippocampus or medial PFC prior to the exploration of the objects impaired memory. Conversely, infusion of the D1/5 agonist SKF81297 into the dorsal hippocampus or medial PFC facilitated memory. Reboxetine similarly facilitated recognition memory in healthy rats and the D1/5 antagonist SCH23390 reversed this effect when infused into the dorsal PFC, but not when infused into the hippocampus. Moreover, systemic reboxetine increased the levels of the NMDA subunit GluN2A in the PFC but not in the hippocampus. Finally, we demonstrate that a single dose of reboxetine does not affect immobility in the forced swim test but improves recognition memory in the Flinders sensitive line (FSL) rat model for depression. The present data in rats are in line with effects of reboxetine on memory formation in healthy volunteers and depressed patients and indicate the involvement of PFC dopamine D1/5 receptors.

Voluntary alcohol drinking enhances proopiomelanocortin gene expression in nucleus accumbens shell and hypothalamus of Sardinian alcohol-preferring rats.

BACKGROUND: Evidence obtained in humans and rodents indicates that beta-endorphin (encoded by the proopiomelanocortin [POMC] gene) is critical in the regulation of alcohol drinking behavior. However, the alcohol effect on POMC gene expression has not been studied in rodent mesolimbic regions, such as the nucleus accumbens (NAc). METHODS: In this study, we first utilized POMC-enhanced green fluorescent protein (EGFP) transgenic mice to visualize POMC neurons and found that POMC-EGFP cells were modestly distributed throughout the NAc shell and core, in addition to the hypothalamic arcuate nucleus. POMC mRNA expression in the NAc of mice and rats was confirmed using reverse transcriptase-polymerase chain reaction and solution hybridization assays. We then investigated whether there are genetically determined differences in basal mRNA levels of POMC and mu opioid receptor (MOP-r) between selectively bred Sardinian alcohol-preferring (sP) and nonpreferring (sNP) rats, and whether these mRNA levels are altered in sP rats after alcohol drinking (10%, unlimited access) for 17 days. RESULTS: Alcohol-naïve sP rats had higher basal POMC mRNA levels than sNP rats only in hypothalamus. Alcohol drinking increased POMC mRNA levels in both the NAc shell (by 100%) and the hypothalamus (by 50%) of sP rats. Although sP rats had lower basal levels of MOP-r mRNA and GTPγS binding in NAc shell than sNP rats, voluntary alcohol consumption had no effect on MOP-r mRNA levels in the NAc shell. CONCLUSIONS: Our results define the distribution of POMC-expressing neurons in the NAc of mice and rats. Higher POMC expression at basal levels in sP rats (genetically determined), along with increases after drinking (alcohol-induced) in the NAc shell and hypothalamus, suggests that the POMC systems play a role in high alcohol preference and consumption.

Dysfunctional hippocampal activity affects emotion and cognition in mood disorders.

Mood disorders, such as major depressive disorder (MDD), bipolar disorder and generalized anxiety disorder usually comprise mood related as well as cognitive symptoms and the interaction between these symptoms is still not clear. Most antidepressant drugs have a positive effect on mood but do not treat the cognitive dysfunctions or even aggravate the symptoms. In this review we will evaluate the association between mood and cognition in the context of mood disorders. In the first section we will summarize the brain circuits at the intersection between cognition and emotion, highlighting the role of the hippocampus. In the second section, we will survey the contribution of the glutamate and GABA systems in the pathophysiology of mood disorders, making an effort to understand the link between emotions and cognition and how novel therapeutic approaches deal with them. In the third section we will explore the monoamine involvement in the emotion/cognition duality in the context of mood disorders. Finally we will underline the role of synaptic plasticity and neurogenesis in depression. We consider that a broader knowledge about the integrative mechanisms involved in specific aspects of mood disorders is crucial in the development of more powerful and effective antidepressant drugs. This article is part of a Special Issue entitled: Brain Integration.

Increased ethanol intake in prodynorphin knockout mice is associated to changes in opioid receptor function and dopamine transmission.

The purpose of this study was to examine the role of the prodynorphin gene in alcohol sensitivity, preference and vulnerability to alcohol consumption. Handling-induced convulsion (HIC) associated to alcohol, alcohol-induced loss of righting reflex (LORR), hypothermic effects in response to acute ethanol challenge, blood ethanol levels (BELs), conditioned place preference, voluntary ethanol consumption and preference, tyrosine hydroxylase (TH), dopamine transporter (DAT) and proenkephalin (PENK) gene expression, and µ-, δ- and κ-opioid agonist-stimulated [S(35) ]- guanosine 5'-triphosphate-binding autoradiography were studied in prodynorphin knockout (PDYN KO) and wild-type (WT) mice. There were no differences in HIC, LORR or the decrease in body temperature in response to acute ethanol challenge between PDYN KO and WT mice. PDYN KO mice presented higher BEL, higher ethanol-conditioned place preference and more ethanol consumption and preference in a two-bottle choice paradigm than WT mice. These findings were associated with lower TH and higher DAT gene expression in the ventral tegmental area and substantia nigra, and with lower PENK gene expression in the caudate-putamen (CPu), accumbens core (AcbC) and accumbens shell (AcbSh) in PDYN KO. The functional activity of the µ-opioid receptor was lower in the CPu, AcbC, AcbSh and cingulate cortex (Cg) of PDYN KO mice. In contrast, δ- and κ-opioid receptor-binding autoradiographies were increased in the CPu and Cg (δ), and in the CPu, AcbC and Cg (κ) of PDYN KO. These results suggest that deletion of the PDYN gene increased vulnerability for ethanol consumption by altering, at least in part, PENK, TH and DAT gene expression, and µ-, δ- and κ-opioid receptor functional activity in brain areas closely related to ethanol reinforcement.

Innate difference in the endocannabinoid signaling and its modulation by alcohol consumption in alcohol-preferring sP rats.

The present study was undertaken to examine whether genetically predetermined differences in components of the endocannabinoid system were present in the brain of Sardinian alcohol-preferring (sP) and Sardinian alcohol-non-preferring (sNP) rats, a pair of rat lines selectively bred for opposite alcohol preference. The effects of acquisition and maintenance of alcohol drinking, alcohol withdrawal, and alcohol re-exposure on the endocannabinoid system was also assessed in the striatum of sP rats. The findings revealed significantly higher density of the CB1 receptors and levels of CB1 receptor mRNA, CB1 receptor-mediated G-protein coupling, and endocannabinoids in the cerebral cortex, hippocampus and striatum of alcohol-naive sP rats than sNP rats. A significantly lower expression of mFAAH enzyme was evident in the hippocampus of alcohol-naive sP rats. Alcohol drinking (during both acquisition and maintenance phases) in sP rats resulted in a significant reduction in striatal CB1 receptor-mediated G-protein coupling whereas alcohol withdrawal attenuated this effect. Alcohol consumption was also associated with markedly increased levels of endocannabinoids in the striatum. Co-administration of the CB1 receptor antagonist, rimonabant (SR141716A) reduced alcohol intake, and reversed alcohol-induced changes in CB1 receptor-mediated G-protein activation. These findings provided a new insight into a potential genetic basis of excessive alcohol consumption, suggesting innate differences in the endocannabinoid system might be associated with higher alcohol preference in sP rats. The data also indicate a modulation of CB1 receptor-mediated signaling following alcohol consumption, and further strengthen the potential of the endocannabinoid system as a target for the treatment of alcohol related behaviors.

Prodynorphin gene deletion increased anxiety-like behaviours, impaired the anxiolytic effect of bromazepam and altered GABAA receptor subunits gene expression in the amygdala.

This study evaluated the role of prodynorphin gene in the regulation of anxiety and associated molecular mechanisms. Emotional responses were assessed using the light-dark test, elevated plus maze and social interaction tests in prodynorphin knockout and wild-type mice. Corticotrophin releasing factor and proopiomelanocortin gene expressions in the hypothalamus were evaluated after restraint stress using in situ hybridization. The anxiolytic efficacy of bromazepam and GABA(A) receptor subunits gene expression in the amygdala were also assessed in both genotypes. The deletion of prodynorphin increased anxiety-like behaviours and proopiomelanocortin gene expression in the arcuate nucleus (two-fold). Moreover, the anxiolytic action of bromazepam was significantly attenuated in the mutant mice. Decreased GABA(A)γ(2) and increased GABA(A)ß(2) gene expression receptor subunits were found in the amygdala of prodynorphin knockout mice. These results indicate that deletion of prodynorphin gene is associated with increased anxiety-like behaviours, enhanced sensibility response to stress stimuli, reduced anxiolytic efficacy of bromazepam and altered expression of the GABA(A) receptor subunits.

CB1 receptor blockade decreases ethanol intake and associated neurochemical changes in fawn-hooded rats.

BACKGROUND: This study was undertaken to identify the neurochemical changes underlying the attenuation of voluntary ethanol intake induced by the cannabinoid CB1 receptor antagonist AM251 in fawn-hooded rats. METHODS: Rats were exposed to the 2-bottle-choice paradigm (ethanol 10% v/v or water) for 15 days. After this period, rats received AM251 (3 to 6 mg/kg, i.p.) or vehicle. RESULTS: Voluntary ethanol intake decreased (30%) with the administration of incremental dosages of AM251 (3 mg/kg, 5 days and 6 mg/kg, 5 days) in rats with acquired high preferring ethanol consumption (>3.5 g of ethanol/kg/d). Ethanol intake significantly decreased proopiomelanocortin expression in the arcuate nucleus (38.31%) and micro-opioid-DAMGO-stimulated [(35)S]-GTPgamma binding in the caudate-putamen (40%), nucleus accumbens core (AccC) (32.87%), and shell (AccS) (34.21%). Moreover, ethanol intake increased tyrosine hydroxylase (TH) gene expression in the substantia nigra (24%) and ventral tegmental area (23%) and corticotrophin-releasing gene expression in the paraventricular hypothalamic nucleus (41.6%). The reduction of ethanol intake induced by AM251 was associated with blockade or significant reduction of the changes produced by ethanol in the expression of these genes in key regions related to drug dependence. Interestingly, treatment with AM251 reduced (20%) TH gene expression in rats drinking only water. In this respect, the action of AM251 in reducing TH gene expression may not be specific. CONCLUSION: Taken together, these results revealed that blockade of cannabinoid CB1 receptors (CB1r) decreased voluntary ethanol intake in ethanol-habituated rats by normalizing the neurochemical alterations induced by ethanol.