Addiction Neuroscience

BackgroundLoss of control over drinking is a hallmark feature of alcohol use disorder (AUD) that is modeled preclinically through escalation of ethanol consumption and aversion-resistant drinking. Prior work with other reinforcers suggests that within-session unpredictable, intermittent access (uIntA) promotes loss of control over intake. However, the effect of uIntA on voluntary ethanol consumption is unknown. MethodsMale and female Long-Evans rats (n=9-10/group) underwent seven weeks of daily voluntary ethanol (20% v/v) drinking sessions under either a continuous access (ContA) or uIntA schedule. Following four weeks of baseline, rats were rendered dependent using a two-week chronic intermittent ethanol vapor exposure procedure. Daily testing was maintained through one week into withdrawal from vapor exposure. On the final day of testing, ethanol was adulterated with quinine (30 mg/L) to assess aversion-resistant drinking. ResultsRats drinking under ContA and uIntA exhibited similar levels of average daily ethanol consumption at baseline. However, uIntA elicited a more robust dependence-induced escalation of ethanol consumption compared to ContA, with uIntA sustaining escalation through early protracted withdrawal. Additionally, while rats with ContA to ethanol remained sensitive to quinine even after chronic ethanol vapor exposure, uIntA promoted aversion-resistant drinking in ethanol dependent rats. ConclusionsThese results demonstrate that, compared to ContA, uIntA maintains ethanol drinking and exacerbates AUD-related symptomatology while also providing researchers with the ability to capture additional measures of motivation and drinking patterns without increasing experimental burden. This work positions uIntA as a powerful tool to assess psychological and neurobiological factors underlying loss of control over drinking.

Alcohol use disorder (AUD) is a chronic relapsing condition with limited pharmacological treatments. Ethanol modulates neuronal excitability in part through activation of G-protein-gated inwardly rectifying potassium (GIRK/Kir3) channels, which dampen neuronal activity in reward- and stress-related circuits implicated in AUD pathophysiology. In this study, we investigated the therapeutic potential of targeting activation of GIRK channels in mouse models of ethanol intoxication. GiGA1 (G protein-independent GIRK activator type 1) is a selective activator of GIRK1/GIRK2 channels and has good brain bioavailability. Systemic GiGA1 administration prevented acquisition of ethanol-induced conditioned place preference (CPP) in both male and female mice. GiGA1 also significantly reduced voluntary ethanol intake and decreased blood alcohol concentrations, when administered to mice after they developed high preference and consumption of ethanol. Similarly, Baclofen, a GABAB receptor agonist that leads to activation of GIRK channels also decreased ethanol consumption. However, systemic Baclofen did not prevent acquisition of ethanol-dependent CPP, suggesting a broader efficacy of direct GIRK1/GIRK2 activation by GiGA1. Whole-brain c-Fos mapping as a proxy for neuronal activity revealed that GiGA1 blunted ethanol-induced neuronal activation in several AUD-relevant brain regions, including the central amygdala, paraventricular thalamus, paraventricular hypothalamus, and Edinger-Westphal nucleus. These findings demonstrate that pharmacological activation of GIRK channels modulates key neural circuits involved in ethanol reward and intake, supporting GiGA1 as a promising lead compound for targeted AUD therapy.

Background: Alcohol use disorder is associated with dysregulated glutamatergic signaling within mesocorticolimbic circuits that govern reinforcement and excessive ethanol intake. Group II metabotropic glutamate receptors (mGlu2/3) act primarily as presynaptic autoreceptors that regulate glutamate release. However, how voluntary alcohol intake alters mGlu2/3 expression within reward circuitry remains unclear. Methods and Results: We examined the effects of operant alcohol self-administration on mGlu2/3 protein expression and assessed the functional impact of group II receptor modulation on binge-like ethanol intake. Male C57BL/6J mice self-administered sweetened ethanol or sucrose under behaviorally matched conditions for 35 days. Immediately after the final session, tissue punches from the nucleus accumbens (NAc), amygdala, and prefrontal cortex were collected for Western blot analysis. Operant ethanol self-administration selectively reduced mGlu2/3 protein expression in the NAc, with no changes detected in the amygdala or prefrontal cortex. Both monomeric and dimeric mGlu2/3 protein levels were reduced, and a composite index revealed coordinated downregulation of receptor expression. In separate cohorts, systemic administration of the mGlu2/3 agonist LY379268 dose-dependently reduced binge-like ethanol intake in a limited-access home-cage drinking model, whereas positive allosteric modulation of mGlu2 receptors with LY487379 was ineffective. Conclusions: These results show that low-dose operant ethanol self-administration produces an ethanol- and region-specific reduction of mGlu2/3 protein expression in the NAc and that pharmacological activation of group II receptors, potentially involving mGlu3-specific receptors, is sufficient to suppress binge-like ethanol consumption. These data identify presynaptic mGlu2/3 dysregulation as a mechanism contributing to ethanol-related behaviors and support group II metabotropic glutamate receptors as therapeutic targets for alcohol use disorder.

Neuroimmune signaling is increased in postmortem brain tissue from individuals with alcohol use disorder (AUD), and growing evidence suggests that it contributes to persistent alcohol-related neuroadaptations. Interferon regulatory factor 7 (IRF7), a transcription factor downstream of endosomal Toll-like receptor signaling, is induced in alcohol-relevant brain regions and may contribute to escalated drinking. Here, we tested whether chronic intermittent ethanol (CIE) vapor exposure engages IRF7 signaling during subsequent alcohol self-administration and whether this is associated with altered molecular E/I balance in the aIC and altered functional E/I balance in aICnucleus accumbens projection neurons. Female Wistar rats (n=30) were trained to self-administer alcohol (15% v/v; FR2 vs inactive lever) during 30-minute sessions. After establishing baseline drinking, rats underwent 1-3 cycles of CIE, which increased alcohol self-administration at the 72 h post vapor test. This increase positively correlated with IRF7 levels in the anterior insular cortex (aIC) and nucleus accumbens, while molecular, and immunofluorescence showed that CIE shifted aIC excitatory/inhibitory (E/I) balance toward reduced excitation. Electrophysiological recordings further showed reduced functional E/I balance in aIC neurons projecting to the nucleus accumbens. Knockdown of IRF7 in the aIC attenuated CIE induced escalation of alcohol self-administration, supporting a role for insular IRF7 signaling in alcohol related neuroadaptations that promote escalated drinking.

BackgroundRates of binge drinking have converged significantly between the sexes over recent decades, driven by increased rates of alcohol misuse in women. However, understanding of fundamental circuitry and neurobiology driving alcohol use in females, or how this may differ from male subjects remains underexplored. MethodsWe quantified c-Fos expression across 40 brain regions in alcohol naive, alcohol anticipating and binge drinking male and female mice. In vivo fiber photometry examined sex differences in basolateral amygdala (BLA) activity changes to alcohol intake. Chemogenetic BLA inhibition investigated a functional role in binge drinking. We then assessed sex differences in BLA efferent projection activation following binge drinking. Finally, we functionally interrogated the BLA to nucleus accumbens core (AcbC) projection in binge drinking. ResultsBinge drinking reduced network modularity (number of communities with similar activation patterns) in both sexes relative to alcohol naive and anticipating same-sex counterparts. Female binge drinking mice had increased BLA c-Fos expression compared to female naive and male binge drinking counterparts. In vivo fiber photometry revealed greater and more prolonged BLA responsivity at the onset of alcohol intake in females. Global BLA inhibition reduced reward intake in both sexes. However, the BLA to AcbC projection was preferentially activated in female binge drinking mice, and inhibition of this pathway reduced binge alcohol intake exclusively in females. ConclusionsWe identified sex differences in the neural circuits engaged in binge drinking, highlighting the BLA to AcbC projection may in part underpin sex differences in alcohol misuse. This provides further evidence of distinct neurobiological drivers of alcohol-related behaviors between the sexes.

Opioid use disorder (OUD), which frequently co-occurs with HIV infection, causes long-term neurological disease, yet the epigenetic and transcriptomic effects of OUD and HIV on specific cell types and regions of the brain are poorly understood. To assess the cell-type specific impacts of OUD and HIV across the human brain, we measured single cell transcriptomes and epigenomes of 580,353 cells in the prefrontal cortex, amygdala and cerebellum of 44 donors. We cataloged over 750k candidate cis-regulatory elements (cCREs) and identified gene regulatory networks (GRNs) of transcription factor activity across 35 neuronal and non-neuronal cell types. We identified specific neuronal and glial populations whose cCREs were significantly enriched for genetic risk of addiction-related traits. In OUD donors, we found evidence for reduced metabolic function in neurons in the PFC and cerebellum as well as increased gene expression related to voltage-gated calcium channel activity in the cerebellum. Using a cerebellar organoid model, fentanyl treatment reduced metabolic activity while increasing neuronal activity. Across brain regions, HIV activated immune-related pathways in glial populations, while comorbid OUD and HIV exacerbated metabolic changes in cortical glial cells. Cerebellum-specific Bergmann glia, in addition to forebrain microglia and astrocytes, showed expansion of reactive state identity in HIV. These results highlight shared and specific changes to immune, synaptic, and metabolic processes in OUD and HIV across brain regions and reveal that cerebellar cell types are distinctly affected by opioid abuse.

Alcohol use disorder is a major public health concern worldwide and there is a high comorbidity with psychiatric disorders. The basolateral amygdala (BLA) has been implicated in both mood and alcohol use disorders; however, the mechanisms contributing to the shared pathophysiology remain unknown. Extensive evidence indicates that ethanol modulates GABAergic signaling in the BLA, including actions on neurosteroid-sensitive, extrasynaptic {delta} subunit-containing GABAA receptors (GABAARs), which has been suggested to mediate many of the behavioral effects. In fact, several studies have suggested that 5-reduced neurosteroids, such as allopregnanolone, may mediate some of the behavioral effects of alcohol. Here we demonstrate that chronic intermittent ethanol (CIE) exposure impairs endogenous neurosteroidogenesis via downregulation of key neurosteroidogenic enzymes, 5-reductase type 1 and type 2. To examine the impact of impaired endogenous neurosteroidogenesis of the behavioral consequences of chronic alcohol exposure, including withdrawal-induced anxiety and increased alcohol consumption, we used CRISPR/Cas9 mediated knockdown of 5-reductase in the BLA. Reduced expression of 5-reductase in the BLA did not impact post-CIE alcohol intake or anxiety-like behaviors during withdrawal, perhaps because endogenous neurosteroidogenesis is already impaired following CIE. Therefore, we examined the impact of enhancing neurosteroid levels, treating mice post-CIE with SGE-516, a synthetic GABAAR positive allosteric modulator, which increased voluntary alcohol intake. These findings implicate endogenous neurosteroidogenesis in behavioral outcomes associated with withdrawal from chronic alcohol exposure. Further, this study suggests that targeting endogenous neurosteroidogenesis may be a novel and useful therapeutic target.

Alternative polyadenylation (APA) is a common posttranscriptional mechanism to regulate gene expression. APA generates mRNAs with varying lengths of 3' UTRs or transcripts that encode distinct protein carboxy-terminal ends. APA is especially important in neurons, where different mRNA variants are often asymmetrically localized to dendrites and axons, and can be locally translated into proteins. Local protein synthesis is crucial for axon guidance, synaptic plasticity, and learning and memory, key processes associated with the development of alcohol use disorder (AUD). We investigated the role of APA in AUD using a mouse model of alcohol dependence characterized by increased voluntary drinking after chronic intermittent ethanol (CIE) exposure. We examined APA during protracted withdrawal from alcohol in three brain regions of male and female mice. Our analyses revealed hundreds of genes undergoing APA in males, but substantially fewer in females, suggesting sex-specific effects of CIE on APA. Notably, male and female mice displayed distinct APA signatures. APA genes were different from differentially expressed genes (DEGs), suggesting that these molecular processes are regulated independently. We also determined that the expression of APA genes was associated with neurons, while DEGs were associated with non-neuronal cells. Many of the APA genes were involved in synaptic integrity, neuroplasticity, and neuronal maintenance, which was consistent with their enrichment in neurons. Our study suggests that APA is a crucial sex- and cell type-specific mechanism in AUD with the potential to influence localized neuronal protein expression during protracted withdrawal and to modify alcohol consumption behavior. HIGHLIGHTSO_LIChronic ethanol exposure in mice results in profound changes of APA genes in brain. C_LIO_LICommonly regulated cleavage and polyadenylation sites and genes were identified in male but not in female mice. C_LIO_LIThere was a minimal overlap between APA and differentially expressed genes (DEGs). C_LIO_LIAPA genes were primarily associated with neurons, whereas DEGs were associated with non-neuronal cells. C_LI

Opioid use disorder (OUD) is a chronic, relapsing disease driven by the reinforcing properties of opioids and perpetuated by avoidance of the negative affective states associated with the absence of the drug. Most available OUD treatments directly engage the {micro}-opioid receptor and may induce side effects that can compromise their therapeutic efficacy, thus underscoring the need for novel therapeutic alternatives. Calcitonin gene-related peptide (CGRP) is produced by a small population of neurons in the parabrachial nucleus (PBN) that has been shown to modulate itch, pain, as well as appetitive behaviors. Using a cell-specific nuclear labeling approach coupled with RNA-sequencing, we generated a baseline transcriptome of CGRPPBN neurons and confirmed expression of multiple genes associated with behavioral responses to appetitive stimuli, as well as enrichment of the {micro}-opioid receptor, suggesting that CGRPPBN neuron function may be sensitive to the presence of opioids. Indeed, cFos immunostaining showed that CGRPPBN neuron activity increases during early morphine abstinence and reduces gradually over 48 hours. Given the inhibitory effects of opioids on CGRPPBN neuron activity, we next tested whether these neurons could regulate opioid reinforcement. Using a mouse model of morphine intravenous self-administration, we found that chemogenetic inhibition of CGRPPBN neurons significantly reduced the number of morphine rewards earned in both single-dose and dose-response tests but did not affect context-induced morphine seeking after 21 days of abstinence. These results suggest that CGRPPBN neurons are sensitive to opioid administration and can regulate appetitive behaviors such as morphine-taking. Considering that CGRP signaling is regulated by opioid administration, molecular targets that regulate CGRP neurotransmission without direct -opioid receptor engagement may therefore serve as novel therapeutic avenues for the treatment of OUD. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=195 SRC="FIGDIR/small/712659v1_ufig1.gif" ALT="Figure 1"> View larger version (56K): org.highwire.dtl.DTLVardef@1fb9c9borg.highwire.dtl.DTLVardef@1e6ba79org.highwire.dtl.DTLVardef@dc60f5org.highwire.dtl.DTLVardef@61adaf_HPS_FORMAT_FIGEXP M_FIG C_FIG

The misuse of opioid medications is a significant health issue in the United States. Very few studies have investigated the effect of opioids on perineuronal nets (PNNs), scaffold-like structures that surround neurons and are involved in the regulation of plasticity-dependent mechanisms such as development, learning and memory, and acquisition of addiction-like phenotypes. Regulation of PNNs in the orbitofrontal cortex (OFC) during periods of drug intoxication or withdrawal is widely unknown. In this study, male Wistar rats were injected with fentanyl (0.125 mg/kg, s.c.) or 0.9% saline twice daily for 7 days and once on day 8 (7continuous days following by 3 days of abstinence) or twice daily for 15 days (5 continuous days followed by 2 days of abstinence for more than 3 weeks) and twice on day 16. Antinociception was evaluated using the tail immersion test immediately before and 30 minutes after injections. Whole-brain coronal slices were collected, and immunohistochemistry was used to identify Wisteria Floribunda Agglutinin (WFA)-positive PNNs and parvalbumin (PV)-expressing cells. Results confirmed that repeated fentanyl injections induced tolerance to the antinociceptive effects, which normalized following acute abstinence periods. WFA intensity decreased following 8 days of injections. Analyses confirmed significant correlations between PV+ density and tail withdrawal latency following 8 days of fentanyl injections. These data confirm that repeated fentanyl injections modulate both WFA+ and PV+ expression in the rodent brain and antinociceptive tolerance in a duration-dependent manner. Overall, these data suggest that perineuronal nets may mediate opioid-induced behavioral effects, such as antinociceptive tolerance, following repeated administration and abstinence in rats.

"Dry Hitting" is a unique phenomenon of e-cigarette use that has been shown to produce toxic chemical degradants and byproducts. Although it is widely understood that nicotine exposure during adolescence impacts neurobiological and behavioral function, little is known about how dry hitting may impact users. We hypothesized that subjects repeatedly exposed to nicotine dry hit vapor would exhibit distinct behavioral responses compared with saturated nicotine vapor and would differentially alter the expression of perineuronal nets (PNNs) in the rodent brain. Using a customized system of e-cigarette vapor inhalation, adolescent male Wistar rats (PND 31-40) received vaporized nicotine (30 or 60 mg/mL; [~]2.5-3 mL/cage), nicotine with dry hits (60 mg/mL; 1.75-2 mL/cage), or propylene glycol (PG) vehicle for 30 minutes over 7 daily sessions. Locomotor activity, antinociception, and elevated plus maze testing were used to assess behavioral response to drug intoxication and tolerance. Immunohistochemistry was used to identify Wisteria Floribunda Agglutinin (WFA)-positive PNN structures in the amygdala and insular cortex. Rats exposed to dry hits exhibited behavioral responses (locomotor sensitization, antinociception) similar to those of rats exposed to saturated nicotine vapor, but spent more time in the open arms of the elevated plus maze. Immunohistochemical analyses confirmed significantly greater WFA intensity in the central nucleus of the amygdala, but not the basolateral amygdala or insular cortex, of rats exposed to dry hits. Overall, these data confirm the impact of dry hit vapor on behavioral responses and perineuronal net expression in rats during adolescence.

Alcohol use disorder (AUD) affects over 28 million people in the U.S and is associated with neurobiological alterations, including in the basal ganglia. Within the basal ganglia, the caudate nucleus (caudate) and putamen are implicated in AUD due to their roles in ethanol reinforcement, with the caudate receiving inputs from cortico-associative areas and the putamen receiving inputs from somatosensory areas, supporting goal-directed and habitual behaviors respectively. These distinct behavioral roles are supported by dopamine signaling, including phasic dopamine, involved in assessing action-outcome associations, and tonic dopamine, which reflects ongoing dopaminergic tone that biases action initiation. Intrastriatal dopamine release is modulated by cholinergic interneurons via nicotinic acetylcholine receptors. Dysregulation of these mechanisms can contribute to the transition from occasional to habitual ethanol drinking. Here, we used in-vitro fast-scan cyclic voltammetry to measure dopamine signaling in male (n=6) and female (n=6) rhesus macaques following six months of ethanol self-administration. In putamen, ethanol increased tonic dopamine in both sexes, with females exhibiting greater release and faster dopamine uptake rates than males. In the caudate, ethanol self-administration enhanced dopamine uptake rates only in males. Phasic dopamine release was enhanced in caudate of both sexes but only putamen in males. nAChR blockade revealed that phasic dopamine release in males, but not females, was dependent on cholinergic modulation. These results demonstrate basal and sex-specific dopamine release and uptake are uniquely altered in rhesus macaque caudate and putamen in conjunction with chronic ethanol drinking.

Glucagon-like peptide-1 receptor agonists (GLP-1 RAs) are approved for treating type 2 diabetes and obesity and are under investigation as potential treatments for substance use disorders (SUD). GLP-1 RA-induced weight loss is thought to arise from both peripheral effects on gastrointestinal function and central modulation of appetite and reward circuits, though the exact mechanisms are unclear. Functional magnetic resonance imaging (fMRI) studies examining brain responses to reward-related cues can help clarify the central mechanisms through which GLP-1 RAs influence reward-seeking behavior. We systematically reviewed the fMRI literature examining how GLP-1 RAs affect brain responses to reward-related cues. We identified 1,209 records through a comprehensive literature search. After screening, only 11 studies met eligibility criteria. The vast majority assessed reactivity to food-related cues, with only one examining drug-related cues (alcohol), leaving neural mechanisms relevant to SUD largely unexplored. None included non-food emotional stimuli as control conditions. Several methodological limitations emerged. Most studies enrolled 20 or fewer participants per group, limiting statistical power. Treatment protocols varied substantially, with some assessing cue responses after single-dose administration and others after chronic treatment. Heterogeneity in medications used further confounds interpretation. The limited evidence tentatively suggests that acute GLP-1 RA administration may reduce brain reactivity to food cues in appetite and reward regions. However, effects appear inconsistent and may attenuate over time. Future studies should recruit larger samples, standardize agents and dosing, and assess responses to diverse motivationally relevant stimuli.

Astrocytes are the most abundant glial cells in the brain and an integrative component of the neural network. Studies have shown that ethanol altered expression of an astrocyte marker, i.e., glial fibrillary acidic protein (GFAP), in two key corticolimbic regions, the medial prefrontal cortex (mPFC) and nucleus accumbens (NAc). These regions comprise anatomically and functionally different subregions, i.e., the prelimbic (PL) and infralimbic (IL) cortex of the mPFC, the shell and core subregions of the NAc. However, ethanol effects on GFAP expression within these subregions remain largely unknown. In addition, effects of pharmacological manipulation of astrocytes on alcohol drinking have been understudied. Western blot was conducted to determine GFAP expression in subregions of the mPFC and NAc after chronic ethanol drinking. Fluorocitrate, an astrocyte-specific metabolic inhibitor, was administered to inhibit astrocytes and was tested on ethanol drinking. Ethanol drinking enhanced GFAP protein expression in the PL cortex and NAc core, but not in the IL cortex or NAc shell. Intra-ventricular administration of fluorocitrate reduced ethanol intake and preference, but increased water consumption during choice ethanol drinking. In addition, fluorocitrate did not affect total fluid consumption or basal locomotor activity. These results indicate that chronic ethanol drinking induced GFAP elevation in a subregion-specific manner within the mPFC and NAc, and that metabolic inhibition of astrocytes selectively attenuated ethanol drinking without non-specific effects on water drinking or general activity. Together, these results suggest that astrocytes may play an important role in ethanol drinking. HighlightsO_LIEthanol drinking enhanced GFAP levels in the PL cortex and NAc core. C_LIO_LIFluorocitrate inhibition of astrocytes reduced intermittent ethanol drinking. C_LIO_LIFluorocitrate did not alter total fluid consumption or basal locomotor activity. C_LI

Excessive alcohol drinking is a leading cause of preventable death in the United States. High alcohol consumption and persistent drinking despite adverse events, also known as compulsive drinking, are key criteria that contribute to the development and progression of alcohol use disorder (AUD). There is a clear need to better understand the mechanisms that support these related but distinct behaviors. The serotonin (5-HT) system has been associated with alcohol consumption and risk of alcohol dependence, however given the complexity of this system, there remains much to discover regarding specific alcohol related phenotypes. The current study uses a combination of volitional home-cage drinking and operant conditioning to phenotype mice based on ethanol intake and persistence of alcohol drinking following quinine adulteration, a model to study compulsive drinking. Brain tissue of 10 regions known to be implicated in regulating executive function, reward, and stress was collected, and gene expression of serotonergic receptors, transporters, and enzymes was quantified. Three opioid receptors were included given their well-established roles in alcohol-related behaviors and interactions with the 5HT system. Region-specific gene expression patterns emerged, with serotonergic and opioid receptor expression differentially associated with alcohol drinking phenotype. 5-HT and opioid receptors displayed opposing directionality across regions, consistent with functional heterogeneity within the system. These findings identify region-specific molecular alterations following chronic alcohol that may contribute to individual differences in alcohol drinking phenotypes, highlighting candidate targets for biomarkers of increased alcohol use disorder susceptibility or as interventions aimed at preventing the progression of AUD.

BackgroundFatty foods and alcohol (i.e., ethanol) produce strong reinforcing effects, in part by altering cholinergic interneuron (CIN) activity and tonic dopamine (DA) release within the nucleus accumbens (NAc). Ethanol and fatty foods also both stimulate hepatic and possibly local brain bile acid (BA) synthesis, which raises the possibility that BAs may act as a common upstream regulator of these substances shared mesolimbic effects. MethodsThe current study investigated whether BAs can directly alter mesolimbic activity. Electrophysiological data from acute mouse brain slices was collected to assess BA effects on NAc CIN firing, as well as on excitatory and inhibitory postsynaptic CIN inputs. Bile effects on NAc DA release and clearance rates were measured through voltammetry. ResultsWe found that low concentrations of a 1:1 mixture of BAs cholic acid (CA) and deoxycholic acid (DCA; 1-10 M) increased CIN firing rate, whereas high BA concentrations (1-10 mM) decreased CIN firing. We further demonstrated that BA-induced excitatory effects on CIN firing are independently mediated by at least two mechanisms: Takeda G-protein-coupled receptor 5 (TGR5) activation and suppression of inhibitory CIN currents. Additionally, our results indicate that BAs modulate inhibitory input in a complex manner, reducing frequency at low concentrations, but increasing at high concentrations, and increasing amplitude at low concentrations current amplitude, and the distribution of postsynaptic current amplitude sizes across concentrations. Finally, our voltammetry data indicate that while low BA concentrations enhance NAc DA release without affecting DA uptake, high BA concentrations robustly inhibit accumbal DA release. ConclusionOur findings provide evidence that BAs exert direct modulatory effects on neural activity in the striatum.

Endocannabinoid (eCB) signaling is a key regulator of reward-related dopaminergic signaling, particularly in response to drugs of abuse, such as cocaine. To date, our understanding of this mechanism has primarily been limited to male subjects. Prior work establishes that female cocaine users have more adverse outcomes, and female rats show greater sensitivity to cannabinoid type 1 receptor (CB1R) regulation of cocaine self-administration. Therefore, we hypothesize that female rats exhibit enhanced eCB regulation of cocaine-evoked dopamine (DA). We used in vivo fiber photometry recording of the dopamine biosensor, dLight 1.3b, in the nucleus accumbens medial shell (NAcms) in response to cocaine in male and female rats. Rats were pretreated with cannabinoid-targeting drugs to investigate the effects of CB1R inactivation or augmentation of the eCB 2-AG on cocaine-evoked DA. Our results revealed that CB1R inactivation attenuates cocaine-evoked DA in male and female rats, but females showed enhanced sensitivity for CB1R regulation of cocaine-evoked DA. Cocaine-evoked DA was enhanced by augmenting 2-AG levels, and females again showed increased sensitivity to this manipulation. Finally, females show greater cocaine-evoked DA when in a non-estrous cycle compared to estrous, reinforcing that estrous cycle is a determinant of cocaine-evoked DA. These data indicate that females show enhanced eCB regulation of cocaine-evoked DA signaling, underscoring the importance of sex as a biological variable in our understanding of endocannabinoid regulation of drug reward. HighlightsO_LICB1R inactivation attenuates cocaine-evoked DA in NAcms, preferentially in females C_LIO_LI2-AG augmentation via MAGL inhibition enhances cocaine-evoked DA, with female bias C_LIO_LIEstrous phase modulates the dopamine response to a high dose of cocaine in females C_LIO_LIMale and female rats show similar baseline DA and locomotor responses to cocaine C_LI

IntroductionNeural cue reactivity is increasingly being investigated as a biomarker of treatment response and relapse prediction in addiction disorders. Whilst aberrant brain responses to salient cues (e.g. drugs) have been widely reported in addiction, it is unclear whether these brain responses persist during longer-term abstinence, how they compare between substance use disorder and obesity, and relate to potential differences in eating behaviours. As part of the Gut Hormones in ADDiction (GHADD) neuroimaging study, we investigated how salient cue reactivity to drugs or food, craving and eating behaviours compare in three clinical populations where alterations have been previously observed: abstinent nicotine use disorder (NUD) and alcohol use disorder (AUD), and obesity. MethodsThis study compared group differences in salient cue reactivity and eating behaviours between ex-smokers (n=25, ExS), adults with alcohol dependence who are abstinent (n=26, AAD), adults with obesity who were actively dieting (n=26, OB). Participants completed a high-energy food, preferred alcohol and cigarette functional magnetic resonance imaging (fMRI) cue reactivity task, along with eating behaviour questionnaires, appetite visual analogues scales and an ad libitum test meal. ResultsExS exhibited greater blood oxygen level dependent (BOLD) signal to high-energy food pictures in several reward processing regions in both whole brain and region of interest (ROI) analyses, compared with the OB and AAD groups, with no difference in their appeal rating. Compared with the OB group, ExS exhibited greater BOLD signal to cigarette pictures in the frontal gyrus, orbitofrontal cortex, frontal pole and insula, with no difference in their appeal rating. There were no group differences in preferred alcohol cue reactivity. The AAD group rated sweet taste as more pleasant, and consumed more calories from sweet dishes in the ad libitum meal than the OB and ExS groups. ConclusionsThe presence of heightened cue reactivity to high-energy foods in ex-smokers could contribute to post-quitting weight gain after smoking cessation. Neuroimaging findings were consistent with persistence of some salient drug cue reactivity, despite absence of craving, after medium term abstinence in ExS, but not in AAD. This study also adds to the body of evidence supporting a sweet taste preference endophenotype predisposing individuals to AUD. These changes in eating behaviour in NUD and AUD may provide targets for treatments to reduce substance misuse and facilitate abstinence.

Early life adverse (ELA) experiences such as child maltreatment (MALT) are associated with physical and mental illness, including substance use disorders (SUDs), but underlying neurobiological mechanisms remain unclear. This study examined long-term effects of infant MALT on adult brain serotonin (5HT) and dopamine (DA) receptors in corticolimbic regions involved in reward and emotional control, using positron emission tomography (PET) imaging, a translational infant MALT macaque model of cocaine use disorder (CUD) risk and a COC self-administration (SA) paradigm. The study focused on regional serotonin 5HT1A, 5HT2A, and dopamine D2/D3 receptor availability (BP) differences between MALT and Control animals using PET, both at baseline (pre-COC SA) and following chronic COC SA (once they reached a total of 100 mg/kg intake). We also examined whether levels of these neurochemical receptors predicted COC SA measures, including reinforcing effects and potency using fixed-ratio (FR) peak response rates and progressive-ratio (PR) peak breakpoint. Our findings showed long-term effects of infant MALT on 5HT, but not DA, receptors in corticolimbic circuits. Specifically, MALT animals showed lower 5HT1A BP in the anterior cingulate cortex (ACC), medial prefrontal cortex (mPFC), and hippocampus compared to Controls. A MALT by Sex interaction effect was detected in 5HT2A BP in the OFC, with lower levels in MALT than Control males, but not in females. In addition, upregulation of 5HT1A and 5HT2A receptors was detected following chronic COC SA in most PFC subregions, hippocampus, and NAcc, particularly in the Control group. These findings suggest long-term effects of ELA on adult 5HT, but not DA, receptors in corticolimbic regions involved in emotional and reward processes. We also found associations between PET baseline (pre-COC SA) receptor BP data and COC SA measures. In particular, a positive correlation between 5HT1A receptor BP in caudate and peak FR Response Rates, whereas amygdala 5HT1A receptor levels were positively correlated with peak PR breakpoint and negatively correlated with peak FR Response Rates. Overall, these findings suggest an important role of 5HT1A and 5HT2A PFC receptors in early COC-related changes in reward circuitry and of amygdala 5HT receptors on cocaine-maintained behaviors. The dynamic change of these 5HT1A and 5HT2A receptors following chronic COC exposure was blunted in animals with ELA. It would be important to understand the biological consequences of these dynamic changes in 5HT receptors and whether they are associated with other stages of the addiction cycle, for example COC relapse, which could inform future pharmacological interventions that target 5HT receptors for treatment of CUD. Simple SummaryWe studied the long-term effects of early life adversity (ELA) on adult brain dopamine (DA) and serotonin (5HT) signaling in corticolimbic regions involved in emotional and reward regulation. We used specific PET radioligands that bind to the DA D2/D3, 5HT1A and 5HT2A receptors, finding lower levels of 5HT, but not DA, receptors binding potential (BP) in animals that experienced ELA. We also found associations between PET receptor BP measures and reinforcing effects of cocaine in i.v. self-administration paradigms using fixed- and progressive-ratio reinforcement schedules. In addition, a strong upregulation of 5HT, but not DA, receptors was identified following chronic cocaine exposure in prefrontal cortex (PFC). Our findings suggest long-term effects of ELA on adult PFC 5HT1A and 5HT2A receptors. The findings also suggest an important role of 5HT1A and 5HT2A, more so than D2/D3, receptors in early cocaine-related changes in reward circuitry. The early dynamic changes of these 5HT receptors could serve as biomarkers for cocaine use disorder (CUD) and inform future pharmacological interventions.

Accurate quantification of rodent licking behavior is essential for studies of fluid intake, including investigations of alcohol use disorder and obesity. Existing lickometry systems vary widely in sensing modality, cost, scalability, and data resolution, and many available systems either require specialized housing or store only binary lick/no lick data based on thresholding. Here we present CLiQR (Capacitive Lick Quantification in Rodents), an open-source capacitive lickometry system designed for high-throughput recording of licking behavior in home-cage environments while preserving the full capacitance time series. The system uses MPR121 capacitive sensors connected to custom metal-tipped serological pipette sippers and a centralized desktop computer to record data from up to 24 animals concurrently, with capacity for two-bottle choice experiments. Validation experiments demonstrated that the capacitive signals reliably distinguish licking from non-licking interactions. Total lick counts showed a strong positive correlation with measured fluid consumption (r = 0.827, p < 0.0001), confirming that detected events provide a meaningful proxy for intake. All information necessary to reproduce the system is shared openly in this manuscript and online. By combining scalability, full-trace data acquisition, and low cost, CLiQR provides a flexible and extensible platform for high-throughput behavioral neuroscience experiments and enables retrospective improvement of lick-detection algorithms. Significance StatementUnderstanding ingestive behavior requires measuring both total consumption and consumption pattern. Licking microstructure provides information about motivation, palatability, and behavioral strategies (i.e., binge-like front-loading); yet many existing lickometry systems are limited by high cost, low scalability, specialized housing requirements, or loss of information due to event-only data storage. We introduce CLiQR, an open-source capacitive lickometry system that enables high-throughput, home-cage recording from dozens of animals while preserving the full time series of capacitance data. By retaining raw data, CLiQR allows post hoc validation and reanalysis of licking behavior, addressing a key limitation of many current systems. This approach increases experimental flexibility, improves data transparency, and lowers barriers to large-scale studies of ingestive behavior.