Addiction Neuroscience

RationalePrenatal alcohol exposure (PAE) can result in Fetal Alcohol Spectrum Disorder (FASD), which consists of a group of diagnosable medical conditions that can include an increased risk for anxiety disorders and/or alcohol misuse, and sensory issues, such as increased mechanical sensitivity. ObjectiveThis study investigated how a single moderate PAE on gestational day 12 (G12) alters anxiety-like behavior, ethanol (EtOH) intake, and mechanical sensitivity across the lifespan of Sprague Dawley rats. MethodsPregnant dams were exposed to vaporized EtOH or room air (control) for 6 hours (BECs [~]108 mg/dL). Testing in male and female offspring began at three different ages: juveniles ([~]postnatal day (P) 25), adolescents ([~]P45) and adults ([~]P80). ResultsThe greatest PAE effects were observed in adolescent animals, with alterations in anxiety-like behaviors demonstrated in the light-dark box and elevated plus maze. Additionally, adolescent female animals consumed more sweetened EtOH compared to males. However, PAE adolescent animals consuming less sweetened EtOH compared to their counterparts, which was also observed in adult PAE females. Interestingly, this effect is reversed in juvenile and adolescent males when tested with unsweetened EtOH, with juvenile females consuming more EtOH also. Finally, PAE and air animals exhibited increased mechanical sensitivity following post-natal EtOH consumption across all ages. ConclusionThese data demonstrate that there are age- and sex-specific effects of PAE on anxiety-like behaviors, EtOH intake, and mechanical sensitivity that are more distinct in adolescent animals.

BackgroundDisturbance of circadian rhythms is a hallmark of substance use disorders, with depressant drugs often causing soporific effects such as reduced sleep latency. The suprachiasmatic nucleus (SCN) of the hypothalamus is the central circadian pacemaker in mammals, regulating daily rhythms in physiology and behavior. However, the cellular mechanisms through which depressants alter SCN function remain poorly defined. MethodsWe used whole-cell patch clamp electrophysiology in acute brain slices to examine how alcohol and opioids modulate excitatory glutamatergic transmission onto SCN neurons. Ethanol effects were examined both acutely and following chronic exposure paradigms. Optogenetic stimulation was used to activate either RHT input or -opioid receptor-expressing (MOR) terminals, and MOR agonists were used to assess opioid-mediated effects on synaptic transmission. ResultsWe show that acute application of ethanol paradoxically enhances SCN firing rates. In contrast, chronic alcohol exposure reduces glutamatergic drive. We also found that activating MOR+ terminals produced bidirectional modulation of SCN firing and that MOR+ inputs formed functional glutamatergic synapses onto SCN neurons. Notably, this transmission could be suppressed by the MOR agonists DAMGO and fentanyl. ConclusionsTogether, these findings reveal that both alcohol and opioids modulate glutamatergic input to the SCN. This work establishes the SCN as a novel target of depressant substances and highlights glutamatergic transmission as a key point of vulnerability in circadian dysregulation associated with substance use.

BackgroundAlcohol use disorder is a chronic relapsing condition characterized by excessive drinking and withdrawal symptoms. Alcohol dependence disrupts function across multiple brain regions, and recent evidence implicates the cortical amygdala (CoA) as a critical node in alcohol-related circuits. However, how CoA activity influences alcohol intake and brain-wide network function during withdrawal remains unclear. MethodsAlcohol dependence was induced using chronic intermittent ethanol vapor (CIE). In one cohort, electrophysiological activity of CoA neurons was assessed during withdrawal. In a second cohort, mice underwent CIE paired with two-bottle choice drinking, and inhibitory DREADDs (hM4Di) were used to suppress CoA activity during drinking and withdrawal while behavioral outcomes were measured. Brains were then collected for Fos immunolabeling and iDISCO+ based whole-brain activity mapping to determine how CoA inhibition during withdrawal altered network organization. ResultsRepeated CIE increased alcohol sensitivity in CoA neurons during withdrawal. Chemogenetic inhibition of the CoA reduced alcohol intake in dependent mice without affecting withdrawal-related behaviors. Whole-brain Fos mapping showed that CoA inhibition reduced activity within the CoA while enhancing functional connectivity across multiple brain regions, particularly in the isocortex, thalamus, and anterior hypothalamic nucleus. During withdrawal without CoA inhibition, thalamic regions exhibited negative connectivity, consistent with disrupted network function; CoA inhibition reversed this pattern, producing strongly positive thalamic and medial prefrontal cortex connectivity. ConclusionsThese findings demonstrate that alcohol dependence alters CoA sensitivity, alcohol dependence-induced drinking and brain-wide network organization during withdrawal. The CoA appears to selectively regulate withdrawal-associated alcohol drinking, and its inhibition may reduce intake by restoring thalamic and cortical connectivity. HighlightsO_LIThis study identifies the cortical amygdala as a previously underexplored brain region involved in alcohol-related behaviors. C_LIO_LIBy integrating chemogenetic inhibition with brain-wide network analysis, the study reveals candidate circuit connections through which the CoA may regulate alcohol dependence-related brain activity. C_LIO_LIThis study establishes the CoA as a potential driver of excessive alcohol drinking and alcohol-related network dysfunction. C_LI

RationaleThe progression of psychostimulant abuse is associated with a shift from recreational to habitual use (R2H-shift). Because this R2H-shift can be modeled using behavioral economics, we developed a novel Behavioral Economic model for the Analysis of Self-administration Time-curve (BEAST) to obtain R2H-shift variable(s). The relationship(s) between R2H-shift variables and drug intake (under normal and/or punishment conditions) is/are unknown. Our goal was to determine if the R2H-shift variable and intake variables obtained during the initial self-administration training phase were related to 1) drug intake at that time, and subsequent drug intake under 2) normal, 3) punishment, 4) post-punishment, and 5) price-constrained conditions. MethodLong Evans rats self-administered methamphetamine (METH, males n = 16, females n = 14), sucrose (males n = 22, females n = 22) and/or saline (males n = 3, females n = 10) under FR1 for 6 h per day for 20 days to obtain 1) followed by the assessment of subsequent drug intake under different conditions (2-5 above). We obtained all variables referenced above. We determined the relationships between all variables (multivariate analysis). ResultsThere were no sex differences detected in the METH and sucrose studies. For METH and sucrose, prior drug intake levels could predict drug intake under normal/punishment but not under price-constrained conditions. The R2H-shift variable could predict drug intake under a consumption-price curve but could not predict intake under normal/punishment conditions. ConclusionsWhile related to economic demand, the recreational-to-habitual shift rate was unrelated to drug intake levels (under normal and punishment conditions).

Excessive alcohol consumption remains a major public health challenge with limited therapeutic options. Both glucagon-like peptide-1 (GLP-1) and fibroblast growth factor-21 (FGF21) independently regulate alcohol intake through complementary metabolic and reward pathways, but their combined potential has not been explored. Here, we report that a long-acting dual agonist, GLP1-ELP-FGF21 modulates behavioural, neurophysiological, and cognitive components of alcohol seeking in mice. A single GLP1-ELP-FGF21 dose reversibly reduces voluntary alcohol intake for at least 72 hours in male mice, has sustained effects in female mice, and markedly blunts nucleus accumbens dopamine transients aligned to the initiation and termination of lick bouts during alcohol consumption. To assess its effects on decision-making, we used a novel two-choice (alcohol versus food) decision task modelled with evidence-accumulation frameworks. Alcohol choice behaviour conformed to evidence accumulation decision models: Linear Ballistic Accumulator (LBM) and Racing diffusion models (RDM). Critically, GLP1-ELP-FGF21 selectively reduces choices for alcohol and slows the latent accumulation rate for alcohol options, without affecting food-directed choice or non-decision processes. Sensory-specific satiety devaluation confirms that reductions in reward value are explained by reductions in accumulation rates. Together, these results highlight GLP1-ELP-FGF21 as a therapeutic strategy for alcohol use disorder via modulation of central reward pathways and decision-making when confronted with alcohol rewards.

Background and Aims: Synthetic opioids cause tens of thousands of deaths each year in North America, and there are indications that synthetic opioids are also becoming increasingly prevalent in the European drug market. This study aimed to examine high-risk substance use in the German drug-using community with a particular focus on the synthetic opioids fentanyl and nitazenes and related awareness, concerns, overdose experiences, and harm-reduction behavior. Design: Cross-sectional, observational online survey. Setting: Open drug-use scenes, addiction clinics, and substitution practices at numerous geographic locations throughout Germany, August to September 2025. Participants: 235 individuals aged 14+ from the drug using community (mean age 43.4 years; 57.9% male), 79.6% recruited by peers in open drug-use scenes. Measurements: The primary outcome was substances used within the past 12 months. In addition, sources, forms, routes of administration, and perceived changes in availability and price of (synthetic) opioids were assessed as well as risk perceptions, fears, harm-reduction behavior, and overdose-related experiences. Findings: 227 respondents reported substance use with an average of 6.2 substances, and 73.1% (95% confidence interval [CI] = 67.0-78.5%) had used at least one opioid in the past year. Synthetic opioids were consumed in many parts of Germany and across all age and gender groups. Among participants who experienced a shortage of their primary opioid in the past year, 25% (95% CI = 15.8-37.2%) reported having used fentanyl instead. 56.5% (95% CI = 36.8-74.3%) of individuals using synthetic opioids reported having experienced an overdose in the past twelve months. Most of the respondents perceived synthetic opioids as posing a high risk, and a substantial proportion expressed fear that they could be mixed into their own substances. However, only 9.9% (95% CI = 6.6-14.7%) use drug checking, although the vast majority stated they would use it if it were available to them. Conclusions: Synthetic opioids, including fentanyl and nitazenes, have entered the German drug scene, with users reporting high rates of overdose and limited access to harm reduction measures. Germany may be in an early phase of a synthetic opioid transition, warranting urgent expansion of surveillance, naloxone distribution, and drug checking services.

The addictive properties of opioids are due in part to these drugs ability to alter ventral tegmental area (VTA) activity via activation of mu opioid receptors (MORs) on local and distal inputs. Prior studies have identified numerous opioid-modulated afferents to the VTA, some of which show differing levels of functional modulation by opioids, but the degree to which this parallels differences in receptor expression is not known. Hence, we used retrograde labeling combined with RNAscope to examine oprm1 mRNA expression in VTA-projecting afferents arising from a variety of distal brain regions. Because opioids are thought to be particularly influential on GABAergic afferents to the VTA, we also examined colocalization of oprm1 with GABAergic markers in VTA-projecting neurons. Interestingly, we found that oprm1 mRNA is present in both GABAergic and non-GABAergic VTA-projecting neurons. However, many (though not all) GABAergic afferents expressed higher levels of oprm1 compared to most non-GABAergic afferents (especially those arising from the cortex). These results complement previous anatomical studies that had examined oprm1 expression in these regions but in a non-quantitative way and without regard to their efferent targets. Our findings encourage future work to examine the functional implications of MOR sensitivity within these afferent pathways.

Background: Alcohol use disorder (AUD) is associated with altered gene expression across diverse cell types in reward-related brain regions, including the ventral tegmental area (VTA), which is rich in dopaminergic neurons. The VTA plays a central role in reward processing, learning, and memory; however, cell type-specific gene expression changes within the VTA remain uncharacterized. Methods: We applied single-nucleus RNA sequencing (snRNA-seq) to profile transcriptomic alterations associated with AUD in the VTA. Postmortem VTA samples from four individuals of European ancestry [two with AUD (one male, one female) and two matched controls (one male, one female)] were analyzed using the 10X Genomics Chromium Fixed RNA Profiling protocol. Differentially expressed genes (DEGs) were identified using Seurat, and enriched KEGG pathways was assessed by gene set enrichment analysis. Results: Nuclei were classified into six major cell types: astrocytes, endothelial cells, mature neurons, microglia, oligodendrocytes, and oligodendrocyte precursor cells (OPCs). At thresholds of P < 0.05 and |fold change| > 2.0, we identified 547 DEGs in astrocytes, 727 DEGs in endothelial cells, 715 DEGs in mature neurons, 421 DEGs in microglia, 263 DEGs in oligodendrocytes, and 432 DEGs in OPCs. DEGs across VTA cell types were enriched for pathways related to mitochondrial function, neurodegeneration, and synaptic signaling. Notably, DEGs in mature neurons were enriched for addiction-related pathways. Further subdivision of mature neurons into dopaminergic, GABAergic, glutamatergic, and unclassified subtypes revealed 526, 930, 896, and 569 DEGs, respectively. Neuronal DEGs indicate a convergence on mitochondrial/oxidative phosphorylation and neurodegeneration-related pathways across subtypes, whereas addiction- and synapse-related pathways show dopaminergic neuron-specific enrichment. Conclusions: This study provides the first cell type-resolved transcriptomic profiling of the human VTA, revealing AUD-associated gene expression alterations across neuronal, glial, and endothelial cells. The observed cell type-specific changes in synaptic plasticity and addiction-related genes offer new insights into molecular mechanisms underlying AUD pathophysiology.

Post-traumatic stress disorder (PTSD) and alcohol use disorder (AUD) are chronic psychiatric disorders that have overlapping symptomology and risk factors, including altered motivation and impulsive behavior. Inescapable exposure to a predator odor stressor (2,3,5-Trimethyl-3-Thiazoline (TMT)) produces PTSD-like symptomology in rats. Individual differences in stress-coping behaviors such as freezing and defensive digging during TMT exposure can predict long-term differences in alcohol-related behaviors and altered neurobiology. Here, we sought to evaluate the relationship between stress coping behavior during TMT exposure and different aspects of decision making. In Experiment 1, male and female rats were trained on an adjusting-amounts delay discounting task, and delay discounting curves were established before and >2 weeks after TMT exposure. In Experiment 2, female rats were trained to self-administer alcohol and sucrose in a concurrent choice procedure. Lever responses and preference for alcohol over sucrose were evaluated before and >2 weeks after TMT exposure, and then motivation for competing reinforcers was evaluated using progressive ratios. Active coping (digging) during TMT exposure was correlated with increased post-TMT impulsive choice (Experiment 1), reduced sucrose lever responses both before and after TMT exposure (Experiment 2), and reduced sucrose lever breakpoint (Experiment 2). Additionally, TMT-exposed rats had increased motivation for both alcohol and sucrose self-administration when available concurrently (Experiment 2). Overall, these findings suggest that behavior prior to and during a stressful experience can predict susceptibility to negative effects on decision making, which may help future studies identify the neurobiology underlying risk for aberrant reward-related behaviors after a traumatic event.

Problematic Sexual Behaviour (PSB) is defined as difficult to control recurrent sexual behaviours that continue despite adverse consequences, leading to social and functional impairment. There is debate whether PSB is a disorder of compulsion or addiction; PSB often co-occurs with neuropsychiatric disorders, but further elucidation regarding underlying biology is required. A deficiency in reward neurotransmitter systems (reward deficiency syndrome: RDS) may underlie a shared vulnerability to addiction. We conducted the first case-control genome wide association study (GWAS) of PSB in patients (n=448), and comparison participants with (n=196) and without PSB (n=1488). We used polygenic risk scores (PRS) to test genetic overlap with related psychiatric, behavioural and personality phenotypes. Three models were used: 1) All-PSB (patient + comparison) vs. controls, 2) Patient-PSB vs controls, and 3) RDS (yes/no). Results suggested genetic overlap of PSB with psychiatric conditions, with PRS for major depression, substance use, and others predicting PSB status. PRS for related behavioural phenotypes (e.g., externalizing, age at first sex, number of lifetime sexual partners) and personality traits also predicted PSB. The patient model showed stronger associations than the All-PSB model, and RDS had both shared and distinct genetics with PSB. As expected with the sample size, only suggestive hits were observed with single variant and gene-based tests. PSB may share genetic mechanisms with various conditions. Further research in larger cohorts is needed to better understand the underlying genetics and environmental factors involved, and to improve diagnostic classification, intervention and treatment prospects.

Opioid withdrawal is associated with heightened pain sensitivity, including allodynia. Although opioid-induced allodynia is well-documented in humans and animal models, the relationship between the severity of opioid withdrawal-induced allodynia and individual addiction-like behaviors remains poorly understood. To address this gap, Heterogeneous Stock rats underwent long access (12 hours/day) intravenous oxycodone self-administration, followed by measurement of mechanical sensitivity at six timepoints across three weeks of abstinence. Rats were stratified by an Addiction Index derived from individual differences in the escalation of oxycodone intake, motivation to consume oxycodone, tolerance to oxycodones analgesic effects, and acute withdrawal-induced mechanical pain sensitivity. Here, we show that oxycodone withdrawal induces significant and prolonged allodynia for up to three weeks, with High Addiction Index rats exhibiting greater intensity and longer duration of pain sensitivity than Low Addiction Index rats. Results remained consistent even when excluding allodynia from the Addiction Index, highlighting the robustness of the association between addiction-like severity and protracted allodynia. Linear regression associations revealed that self-administration behaviors, particularly oxycodone intake escalation and motivation to seek oxycodone, predicted subsequent withdrawal-induced allodynia severity. These findings demonstrate that greater addiction-like severity is associated with more intense and prolonged withdrawal-induced pain, supporting mechanical allodynia as a marker of addiction severity. These results motivate future work to define the mechanisms linking addiction severity to protracted opioid withdrawal-induced pain, with the goal of informing targeted clinical interventions for individuals most susceptible to severe abstinence-related allodynia.

The endogenous peptide dynorphin (Dyn) and its target the kappa opioid receptor (KOR) play a crucial role in regulating factors related to stress and reward. The KOR is expressed in multiple cell types in the nucleus accumbens (NAc), including presynaptic dopamine (DA) terminals, where it inhibits DA release modulates the function of the DA transporter (DAT). The Dyn/KOR system is upregulated by exposure to drugs of abuse including the DAT inhibitor, cocaine, and their activity is integrally involved in negative affective states associated with withdrawal from substance abuse. We aimed to better understand the impact of the Dyn/KOR system on presynaptic DA terminals and potential effects on DAT interactions with cocaine by measuring the impact of the KOR agonist U50,488 on electrically-evoked DA release and subsequent reuptake in NAc slices from C57BL6/J mice. We showed that superfusion of U50,488 inhibited DA release and markedly reduced cocaine-induced inhibition of DA reuptake, indicating tolerance to cocaine effects. We replicated this finding in the NAc of rhesus macaques using the DAT/NET inhibitor nomifensine, demonstrating that these mechanisms are conserved across DAT inhibitors and in non-human primates. KOR activation results in phosphorylation of the Threonine-53 site on the DAT, a process thought to mediate its impact on DAT function. We tested whether this phosphorylation site is required for the KOR-mediated reduction cocaine effects. To tackle this question, we employed a knock-in mouse line with an Alanine-53 on the DAT (DAT-T53A), rendering that residue insensitive to phosphorylation. We show that DAT-T53A mice have enhanced DA release and uptake, and U50,488 has a reduced inhibitory effect on peak DA release. Remarkably, U50,488 no longer modified the effect of cocaine on uptake in these mice, demonstrating the dependence of this effect on phosphorylated Threonine-53 and highlighting a potential mechanism underlying cocaine tolerance.

Lack of social interaction results in various behavioral abnormalities in rodents, including increased anxiety levels, altered sociability, and impaired cognitive ability. Epigenetic factors regulate gene expression, however, how they contribute to juvenile social isolation (jSI)-induced behavioral alterations remains largely unknown. Here, we focused on the nucleus accumbens (NAc), a critical brain region of the reward system that regulates motivation-related behaviors. We first performed RNA-seq on neuronal nuclei and found alterations in genes related to neuronal function, as well as in transcriptional and epigenetic regulation. Protein-protein interaction (PPI) analysis of differentially expressed genes (DEGs) showed that top key nodes among down-regulated genes include membrane receptors (Ntrk2, Grin3a, and Grik1) and an apoptosis regulator (Bcl2). To further investigate whether jSI-induced gene expression alterations are mediated by histone modifications, we next performed CUT&Tag for four histone modifications (H3K4me1, H3K4me3, H3K27ac, and H3K27me3), and the results implied that epigenetic alterations may also play a role in neuronal function as well as transcriptional regulation. Reanalysis of previously published RNA-seq data on the manipulation of histone modification-associated factors (including Kdm6b, Brd4, and Setd1a) suggested that these enzymes were probably involved in jSI-induced gene expression alterations. Taken together, our comprehensive analysis implies the involvement of histone modification regulation in jSI-related alterations of gene expression in NAc.

Relapse to alcohol use is frequently triggered by re-exposure to alcohol-associated cues, and extinction-based interventions reduce relapse vulnerability. However, the cellular mechanisms through which extinction suppresses alcohol seeking remain unclear. Using a mouse model of operant alcohol self-administration, we examined how extinction training alters activity of defined striatal direct-pathway medium spiny neuron (dMSN) populations in the dorsomedial striatum (DMS). Extinction significantly reduced cue-induced reinstatement and decreased reactivation of acquisition-recruited dMSN ensembles during relapse. Chemogenetic activation of acquisition-recruited dMSN ensembles during extinction impaired extinction learning and enhanced subsequent reinstatement, even in the absence of manipulation at test, indicating that suppression of these ensembles is required for effective extinction. In contrast, selective activation of striosomal dMSNs during extinction accelerated extinction learning and further reduced reinstatement without affecting locomotor activity. These findings demonstrate that extinction suppresses alcohol seeking through coordinated modulation of distinct striatal dMSN populations, involving both reduced engagement of acquisition-related ensembles and recruitment of striosomal circuits. Together, these findings provide mechanistic insight into how extinction reshapes striatal circuitry to suppress relapse-related behavior and highlight defined striatal dMSN populations as potential substrates for enhancing extinction-based interventions.

PurposeOpioid overdose deaths disproportionately affect racial and ethnic minority populations in the United States, yet claims-based evidence characterizing the multi-dimensional structure of these disparities across incidence, treatment access, costs, and insurance coverage remains limited. MethodsWe conducted a retrospective cross-sectional and longitudinal cohort analysis using the HealthVerity Launch Sample, a large administrative claims database. The study population comprised 3,675,823 patients across 5 racial groups enrolled between 2020 and 2024. Eight primary analyses were conducted, including age-sex standardized overdose rates, temporal disparity trends, medication-assisted treatment (MAT) receipt, naloxone access, pharmacy costs, insurance payer type, care setting, and multivariable logistic regression for overdose risk. ResultsBlack patients had the highest age-sex standardized overdose rate (363.4 per 100,000; rate ratio [RR] = 1.27 vs. White), and those with opioid use disorder (OUD) received MAT at a rate 35% lower than White patients (19.8% vs. 30.7%; RR = 0.645), driven primarily by a buprenorphine access deficit. AIAN patients demonstrated consistent multi-dimensional disadvantage across naloxone access, MAT engagement, and pharmacy costs. After adjustment for payer type, age, and sex, all non-White groups showed lower adjusted odds of overdose than White patients (Black OR = 0.87; AIAN OR = 0.25), with Medicaid enrollment carrying 7.06 times the overdose odds of commercial insurance. ConclusionInsurance type is the dominant predictor of overdose risk, and the disproportionate Medicaid enrollment of Black patients is both a consequence of structural disadvantage and access disparities. Targeted interventions such as buprenorphine expansion in Medicaid and enhanced naloxone distribution are recommended.

Background: Alcohol use disorder (AUD) is marked by high relapse rates often driven by craving, yet less is known about whether in vivo, social, and place-based alcohol cues are differentially associated with craving across affective states. This study examined independent and affect-contingent associations of these cues with momentary craving in adults with AUD enrolled in an alcohol intervention study. Methods: Thirty-three adults with AUD completed up to four daily ecological momentary assessments (EMA) for 28 days. EMA prompts assessed craving, in vivo alcohol exposure, being around usual drinking partners, being in usual drinking places, and high-arousal positive affect (PA) and negative affect (NA). Multilevel mixed-effects models adjusted for demographics, intervention phase (1 = post, 0 = pre), AUD severity, and temporal and contextual covariates. Results: EMA compliance was high (median per-participant = 86.6%). Within-person elevations in in vivo alcohol exposure and being around usual drinking partners were independently associated with greater momentary craving, whereas being in usual drinking places was not. In vivo alcohol exposure was more strongly associated with craving during higher-than-usual PA ({beta} = 0.08, p = .032), whereas being in usual drinking places was more strongly associated with craving during higher-than-usual NA ({beta} = 0.06, p = .036), adjusting for intervention phase, which was associated with lower craving. Conclusions: Findings support the need for personalized just-in-time adaptive interventions tailored to high-risk, momentary cue-affect contexts in AUD, beyond low-frequency clinician-delivered feedback that may reduce average craving but not fully address real-time risk. ClinicalTrials.gov registration: NCT05135767.

Maladaptive consummatory behaviors can arise from dysregulated circuits, like the extended amygdala that governs motivation and feeding. Neurotensin (NTS) is expressed throughout the central, peripheral, and enteric nervous systems with well-established roles in energy balance and feeding. SBI-553, a {beta}-arrestin-biased allosteric modulator of NTSR1, recruits {beta}-arrestin while attenuating Gq-mediated signaling. We used SBI-553 to examine NTS modulation of extended amygdala GABAergic signaling, and probed its effects on food consumption in mice. Ex vivo, we found that NTS and SBI-553 differentially modulates GABAergic neurotransmission across extended amygdala subregions. In vivo, SBI-553 reduces palatable food consumption in both fed and food-deprived mice, with greater reductions under fasted conditions. SBI-553 alters activation across CeA subregions in a sex- and feeding-state-dependent manner: SBI-553 increases cFos immunofluorescence in the CeAL and CeAC, but not the CeAM. This work supports neurotensinergic modulation as a compelling target for further investigation into the neural substrates of consummatory behaviors. HighlightsO_LINTS enhances GABAergic transmission in the CeAL and the ovBNST C_LIO_LISBI-553 blocks NTS-induced modulation in the CeAL but not in the ovBNST C_LIO_LISBI-553 attenuates feeding of a palatable high-carbohydrate food C_LIO_LIThe effect of SBI-553 on feeding is driven by energy deficit/motivation to feed C_LI Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=156 SRC="FIGDIR/small/722083v2_ufig1.gif" ALT="Figure 1"> View larger version (46K): org.highwire.dtl.DTLVardef@198a6fborg.highwire.dtl.DTLVardef@fae407org.highwire.dtl.DTLVardef@1909d9corg.highwire.dtl.DTLVardef@15b8c57_HPS_FORMAT_FIGEXP M_FIG C_FIG

Relapse-prevention strategies aimed at reducing relapse following abstinence, primarily focus on reducing cravings that lead to drug-seeking triggered by stress, drug-related cues, or re-exposure to the drug. Because addictive drugs form persistent associative contextual memories, we investigated how reactivation of cocaine-related hippocampal memories influences subsequent drug-seeking. Here, we tagged dorsal dentate gyrus (dDG) memory ensembles involved in encoding either a first or fourth cocaine exposure (15mg/kg, i.p) in male and female c57BL/6 mice using a TetTag approach. Mice underwent cocaine conditioned place preference (CPP), extinction, and reinstatement. We assessed whether optical reactivation of tagged cocaine-related ensembles could substitute for a cocaine priming injection to reinstate CPP, whether reactivation altered cocaine-induced reinstatement, and if these effects differed depending on stage of drug exposure. We also compared these effects to reactivation of saline-associated ensembles. Cocaine produced robust locomotor activation during conditioning, and sensitization developed across repeated drug exposures. Reactivation of a cocaine-related engram alone did not reinstate CPP. However, reactivation of the first cocaine exposure engram attenuated cocaine-induced reinstatement. In contrast, reactivation of the fourth exposure engram did not confer this protective effect. Interestingly, reactivation of saline-associated ensembles also reduced cocaine-induced reinstatement specifically in females, suggesting dDG ensemble reactivation may modulate relapse-related behavior through interference or neuromodulatory disruption of cocaine-associated representations, consistent with our prior work. These findings raise the possibility that early contextual experiences form competing or destabilizing representations that interfere with later cocaine-seeking when reactivated. Females also displayed greater sensitivity to locomotor-inducing effects of cocaine memory reactivation, although this was dissociated from CPP. Together, these findings show that cocaine memories are distinct across drug experience and selective reactivation of dDG engrams can differentially influence drug-seeking.

Individuals with Fetal Alcohol Spectrum Disorders (FASDs) show reduced subicular volume, and preclinical studies compliment this by demonstrating that third-trimester-equivalent ethanol exposure induced apoptosis in corticolimbic regions, including the subiculum. The subiculum mediates hippocampal-cortical communication critical for long-term memory consolidation. Within the distal dorsal subiculum, a population of bursting neurons uniquely express VGLUT2 and they play a key role in memory processing. We hypothesized that third-trimester-equivalent ethanol exposure would reduce neuronal and VGLUT2+ cell density in the dorsal subiculum and reduce the excitability of bursting neurons, providing a mechanism for long-term memory impairments observed in FASD. To test this, postnatal day (P)7 mice received a subcutaneous injection of ethanol and long-term effects were assessed in adolescence (P35-62). Using transgenic mice with fluorescently labeled VGLUT2+ neurons, and immunohistochemistry we observed a significant reduction in neuronal density in males and an increase in VGLUT2+ cell density in females. Using whole-cell patch clamp electrophysiology, we observed a reduction in action potentials per burst in both sexes. Additionally, females showed reduced overall excitability, and a subset of neurons exhibited a shift to regular spiking. These findings suggest that development ethanol exposure disrupts subicular output by impairing burst firing, potentially weaking hippocampal-cortical communication and contributing to the cognitive deficits associated with FASD. HighlightsO_LIThird-trimester ethanol targets VGLUT2+ neurons in the dorsal subiculum C_LIO_LIEthanol reduced neuronal density in male dorsal subiculum C_LIO_LIEthanol increases VGLUT2+ cell density in females C_LIO_LIEthanol reduces action potential per burst in both sexes C_LIO_LIFemales show reduced excitability and loss of bursting in some cells C_LI

Apuschkin et al. (2024) proposed a GPCR-based transcriptomic atlas for midbrain dopamine (DA) neuron subpopulations, including candidates such as Nmur1, Cckar, and Ffar4. To guide genetic targeting, these markers must reflect functional expression in adult DA neurons. Using in situ hybridization, Cre-dependent reporter lines, and both intracranial and systemic viral approaches, we find no evidence of adult Nmur1-mediated recombination in DA neurons, while Cckar-driven recombination is consistent with developmental expression only. Notably, Ffar4 expression overlaps extensively with Ntsr1 midbrain populations, indicating that it does not define a distinct DA neuron class. Furthermore, analysis of independent spatial transcriptomic datasets together with our MERFISH data shows that many proposed GPCR markers are not detectably expressed in adult DA neurons. These findings demonstrate that transcriptomic enrichment does not always yield reliable adult markers and highlight the need for functional validation prior to use in circuit targeting.