The Basolateral Amygdala to Ventral Hippocampus Circuit Controls Anxiety-Like Behaviors Induced by Morphine Withdrawal.

Anxiety is one of the most common comorbid conditions reported in people with opioid dependence. The basolateral amygdala (BLA) and ventral hippocampus (vHip) are critical brain regions for fear and anxiety. The kappa opioid receptor (KOR) is present in the mesolimbic regions involved in emotions and addiction. However, the precise circuits and molecular basis underlying anxiety associated with chronic opioid use are poorly understood. Using a mouse model, we demonstrated that anxiety-like behaviors appeared in the first 2 weeks after morphine withdrawal. Furthermore, the BLA and vHip were activated in mice experiencing anxiety after morphine withdrawal (Mor-A). KORs in the BLA to vHip projections were significantly increased in the Mor-A group. Optogenetic/chemogenetic inhibition of BLA inputs ameliorated anxiety-like behaviors and facilitated conditioned place preference (CPP) extinction in Mor-A mice. Knockdown of the BLA to vHip circuit KOR alleviated the anxiety-like behaviors but did not affect CPP extinction or reinstatement. Furthermore, combined treatment of inhibition of the BLA to vHip circuit and KOR antagonists mitigated anxiety-like behaviors and prevented stress-induced CPP reinstatement after morphine withdrawal. These results revealed a previously unknown circuit associated with the emotional component of opioid withdrawal and indicated that restoration of synaptic deficits with KOR antagonists might be effective in the treatment of anxiety associated with morphine withdrawal.

Several nAChRs gene variants are associated with phenotypes of heroin addiction in Chinese Han population.

Heroin addiction is a chronic and complex brain disease. Nicotinic acetylcholine receptors (nAChRs) have been shown as major control points in many of the neurological and physiological disorders involved in heroin addiction. In the present study, thirty-three SNPs across nine nAChR genes were selected and probed for their associations with heroin addiction phenotypes in 801 unrelated northwestern Chinese Han patients. We found that rs2565055 in CHRNA2 gene was associated with daily dose of methadone treatment, and rs2672215, rs2672216 and rs2741865 in CHRNA10 gene were associated with the duration of the transition from first use to dependence (DTFUD). Cox multivariable regression analysis revealed that rs3743075, rs6495309 in CHRNA3, rs2304297 in CHRNA6, and rs1948 in CHRNB4 were associated with sexual desire in patients with heroin addiction. These findings were further supported by the identification of a haplotype block spanning CHRNA5, CHRNA3, and CHRNB4 that is correlated with changes in sexual desire after long-term heroin use. Our findings highlight associations between polymorphisms in nAChRs genes and the phenotypes of heroin addiction in the Chinese Han population. We suggest several nAChRs subunits as potential novel targets for the treatment of heroin addiction.

Modafinil rescues repeated morphine-induced synaptic and behavioural impairments via activation of D1R-ERK-CREB pathway in medial prefrontal cortex.

Long-term opioid abuse causes a variety of long-lasting cognitive impairments such as attention, impulsivity and working memory. These cognitive impairments undermine behavioural treatment for drug abuse and lead to poor treatment retention and outcomes. Modafinil is a wake-promoting drug that shows potential in improving attention and memory in humans and animals. However, modafinil's effect on opioid-induced cognitive impairments remains unclear, and the underlying mechanism is poorly understood. This study showed that repeated morphine administration significantly impairs attention, increases impulsivity and reduces motivation to natural rewards in mice. Systemic modafinil treatment at low dose efficiently ameliorates morphine-induced attention dysfunction and improves motivation and working memory in mice. High dose of modafinil has adverse effects on impulsive action and attention. Local infusion of D1R antagonist SCH-23390 reverses the morphine-induced synaptic abnormalities and activation of the D1R-ERK-CREB pathway in medial prefrontal cortex (mPFC). This study demonstrated a protective effect of modafinil in mPFC neurons and offered a therapeutic potential for cognitive deficits in opioid abuse.

Differential perturbations of gut microbial profiles and co-occurrence networks among phases of methamphetamine-induced conditioned place preference.

The gut-brain axis provides a pathway for the interaction between gut microbiota and methamphetamine (METH) addiction. However, the gut microbial signatures during different phases of METH use remain unclear. In the present study, we established models of acquisition, extinction, and reinstatement of METH-induced conditioned place preference (CPP) in male mice and detected the gut microbiome profiles of the fecal samples at the three phases by 16S rRNA gene sequencing. Our results revealed that the richness of the gut microbiome increased following repeated METH administration, and it decreased after 4 weeks of abstinence. The microbial richness remained at a low level after one METH challenge at the reinstatement phase. The abundance of several genera including Prevotella, Bacteroides, and Lactobacillus differentially altered among phases of METH-induced CPP. The co-occurrence networks of the gut microbiome became weaker and more unstable during the development of METH-induced CPP at the extinction and reinstatement phases. Notably, the predicted gene functions of short-chain fatty acid metabolism, which were correlated with the abundance of Prevotella, Bacteroides, and Lactobacillus, were found differentially enriched among phases of METH-induced CPP. Our findings highlight a potential association between perturbations of the gut microbiome and different phases of METH use.

The role of SIRT1 in the basolateral amygdala in depression-like behaviors in mice.

Previous investigations have implicated the basolateral amygdala (BLA) epigenetic mechanisms in the pathophysiology of depression. SIRT1 is a NAD+-dependent class III histone deacetylase, widely expresses in BLA. However, epigenetic mechanisms in the BLA under the regulation of SIRT1 in the depression are largely uncharacterized. Under the chronic unpredictable chronic mild stress (CUMS) mouse model, we used adeno-associated viral vectors (AAV) that encoded SIRT1-shRNA or SIRT1 to specifically knockdown or overexpress SIRT1 in BLA neurons, respectively. CUMS procedure induced significant depression symptoms including the decreased sucrose preference, the less bodyweight gained, the decreased immobile latency and the increased immobile time both in forced swim test (FST) and tail suspension test (TST). Knockdown of SIRT1 in BLA glutamatergic neurons reversed these depression-like behaviors and restored the synaptic abnormalities. Overexpression of SIRT1 in BLA glutamatergic neurons induced depression-like behaviors in non-stressed control mice. The result of protein expressions and ultrastructural changes were consistent with the behavioral results. Our study suggested that downregulation of SIRT1 in BLA has certain beneficial effect on CUMS-induced depression-like behaviors such as anorexia, anhedonia, hopelessness and despair. In addition, the increased expression of SIRT1 may be the immediate cause of depressive-like symptoms. The abnormal expression of SIRT1 may affect the transcriptional regulation mechanism and signaling protein acetylation, affecting neuroplasticity and ultimately contribute to MDD. In the stress-susceptible mice, these two mechanisms may co-exist, but the specific mechanism needs further research.

Association study of Catechol-o-methyltransferase and Alpha-1-adrenergic receptor gene polymorphisms with multiple phenotypes of heroin use disorder.

Heroin use disorder is a chronic relapsing brain disease containing multiple phenotypes. These phenotypes vary among heroin users and might be influenced by genetic factors. Single-nucleotide polymorphisms (SNPs) of catechol-O-methyltransferase (COMT) and alpha-1-adrenergic receptor (ADRA1A) genes are associated with heroin use disorder. However, it has not been clarified which phenotypes of heroin use disorder are related to these genes. To address this question, we recruited 801 unrelated heroin users and divided them into different subgroups according to four important phenotypes of heroin use disorder. Then 7 SNPs in the functional region of these genes were systematically screened and genotyped using a SNaPshot assay. We found that the A allele of ADRA1A rs1048101 was associated with a shorter duration of transition from first use to addiction. Subjects with the C allele of ADRA1A rs3808585 were more susceptible to memory impairment after heroin use disorder. Subjects with the G allele of COMT rs769224 were more likely to take a higher dose of heroin every day. Our study confirmed the association between polymorphisms of COMT and ADRA1A with those specific phenotypes of heroin use disorder, which will be instructive for the precise treatment of the disease.

Differential alteration in gut microbiome profiles during acquisition, extinction and reinstatement of morphine-induced CPP.

Substance addiction is a chronic and complicated disease involving genetic and environmental factors. Coregulated by the above factors, perturbations of the gut microbiome have been shown to have an essential role in the development of many neuropsychiatric disorders, including addiction. However, shifts in the gut microbiome during different stages of morphine addiction remain uncharacterized. In the present study, we harvested fecal samples from mice at the acquisition (both the control and morphine groups), extinction and reinstatement stages of morphine-induced conditioned place preference (CPP). Gut microbiome profiles were detected with 16S ribosomal RNA gene sequencing. We observed an increase in community richness following morphine conditioning, and it decreased after 4 weeks of abstinence. The abundance of Verrucomicrobia increased and Bacteroides decreased at the acquisition of morphine-induced CPP, while a recovery trend was found at the extinction stage. Several discriminative genera were identified for the characterization of different stages of morphine CPP. Functional analysis of taxa with differential abundance between CPP stages was mainly enriched in the pathways of amino acid metabolism. Taken together, our findings will extend the association between dysbiosis of the gut microbiome and the opioid-induced rewarding or reinforcing behaviors.

Naloxone reversed cognitive impairments induced by repeated morphine under heavy perceptual load in the 5-choice serial reaction time task.

Repeated opioids abuse may produce long-lasting and complicated cognitive deficits in individuals. Naloxone is a typical mu-opioid receptor antagonist widely used in clinical treatment for opioid overdose and opioid abuse. However, it remains unclear whether naloxone affects morphine-induced cognitive deficits. Using the 5-choice serial reaction time task (5-CSRTT), the present study investigated cognitive profiles including attention, impulsivity, compulsivity, and processing speed in repeated morphine-treated mice. Repeated morphine administration (10 mg/kg, i.p.) induced complex cognitive changes including decreased attention and increased impulsivity, compulsivity, processing speed. Systemic naloxone administration (5 mg/kg, i.p.) reversed these cognitive changes under the heavy perceptual load in 5-CSRTT. Using the novel object recognition (NOR), Y-maze and open-field test (OFT), the present study investigated the memory ability and locomotor activity. Naloxone reversed the effect of morphine on recognition memory and locomotion but had no effect on working memory. In addition, repeated morphine administration decreased the expression of postsynaptic density protein 95 (PSD95) and cAMP response element binding protein (CREB) phosphorylation in the prefrontal cortex (PFC) and hippocampus (HIP), and these effects were significantly reversed by naloxone in PFC. Our study suggests that repeated exposure to morphine affects multiple cognitive aspects and impairs synaptic functions. Systemic naloxone treatment reverses the mu-opioids-induced cognitive changes, especially under the heavy perceptual load, possibly by restoring the synaptic dysfunctions.

PI3K-AKT-GSK3ß-CREB signaling pathway regulates anxiety-like behavior in rats following alcohol withdrawal.

BACKGROUND: Alcohol abuse and anxiety disorders often occur concurrently, but their underlying cellular mechanisms remain unclear. Neuroadaptation within the medial prefrontal cortex (mPFC) have been implicated in the molecular mechanisms underlying alcohol drinking behavior and withdrawal. METHODS: A chronic alcohol exposure rat model (35 consecutive days of 10% alcohol intake and 48 h of withdrawal) was established, then, wortmannin (0.5 µg/side) was injected bilaterally into the mPFC. The elevated plus maze (EPM) and open field test (OFT) were used to assess anxiety-like behavior. Western blot assays were used to assess protein levels. RESULTS: We found that anxiety-like behavior peaked approximately 6 h after alcohol withdrawal. However, wortmannin greatly decreased alcohol intake and attenuated anxiety-like behavior in the alcohol exposure rats. Moreover, the PI3K-AKT-GSK3ß signaling pathway was activated after alcohol withdrawal, and phosphorylation of the downstream cAMP response element-binding protein (CREB) was increased. Wortmannin uniformly reversed PI3K-AKT-GSK3ß-CREB pathway phosphorylation. LIMITATIONS: The downstream GSK3ß activity was not intervened and a single dose level of wortmannin was used. CONCLUSION: Our results suggest that activating the PI3K-AKT-GSK3ß-CREB pathway in the mPFC is an important contributor to the molecular mechanisms underlying alcohol withdrawal. PI3K signaling pathway inhibitors are thus potential candidates for treating alcohol abuse.