Behavioral Neuroscience publications

Internal states including stress and satiety, affect cue-induced feeding behaviors, yet the underlying neural mechanisms remain poorly understood. The nucleus of the solitary tract (NTS), a key hindbrain hub that integrates interoceptive and viscerosensory signals and projects to reward processing nuclei, is anatomically well-positioned to modulate cue-induced feeding behaviors in response to internal state changes. Using behavioral paradigms combined with chemogenetics and fibre photometry, this study investigated the hypothesis that NTS mediates the effects of stress and satiety on cue-induced feeding behaviors via A2 neurons and modulation of ventral tegmental area (VTA) dopamine signaling. We first showed that both foot shock stress and outcome specific satiety (i.e., sucrose prefeed) reduced cue-induced appetitive behavior. Inhibition of NTS neurons only attenuated the suppressive effect of foot shock stress, but not that of outcome specific satiety, indicating that NTS is required for stress-induced suppression of cue-induced appetitive behavior. Further investigation into the contributing neural phenotype revealed that stimulation of NTS A2 neurons reduced conditioned approach and suppressed cue-evoked VTA dopamine neural activity, without any effects on lateral hypothalamus (LH) neuron activity. Together, these findings suggest that NTS neurons mediate the effects of foot shock stress, but not outcome specific satiety, on cue-induced appetitive behavior, in part through activation of NTS A2 neurons and modulation of cue-evoked VTA dopamine neural activity. These results provide an NTS-mediated mechanism through which stress suppresses cue-induced feeding behavior.

Developing safe and effective pain medications is an ongoing challenge for human health. Agonists for the µ-opioid receptor (MOR) are essential pain medications, but their high intrinsic efficacy also induces adverse side effects, including respiratory depression, constipation, tolerance, dependence, withdrawal and addiction 1–7 . Strategies to limit adverse effects traditionally include developing MOR agonists that have low intrinsic efficacy or that preferentially activate G-protein signalling over β-arrestin signalling 8 . Here we identify a novel MOR agonist with supramaximal intrinsic efficacy and a unique pharmacological profile that produced effective analgesia in rodents with minimal adverse effects. N -desethyl-fluornitrazene (DFNZ) was derived from a class of synthetic benzimidazole opioids called nitazenes. DFNZ has impaired brain penetrance, a unique spatiotemporal MOR cellular signalling profile, and diminished efficacy at the MOR–galanin 1 receptor (GAL1) heteromer. DFNZ does not induce respiratory depression, tolerance or MOR downregulation after repeated exposure. Compared with other MOR agonists, DFNZ has limited effects on dopamine neurotransmission in nucleus accumbens and weaker reinforcing effects in the drug self-administration procedure. These results provide novel insights about MOR and nitazene pharmacology, have important implications for pain and addiction treatment, and challenge the prevailing dogma that high-efficacy MOR agonists cannot constitute safe and effective therapeutic agents.

Glucose-dependent insulinotropic polypeptide (GIP) is a gut-derived incretin hormone, and pharmacologic modulation of central GIP receptors (GIPRs) improves energy homeostasis and prevents conditioned taste avoidance (CTA). However, the mechanisms by which GIPR signaling impact food intake and aversion are incompletely understood. Here, we show that GIPR agonism abrogates the aversive and enhances the anorexigenic effects of the pro-inflammatory cytokine interleukin-1β (IL-1β). Aversion-encoding parabrachial calcitonin gene-related peptide (CGRP) neurons were required for IL-1β-induced CTA but not anorexia. Moreover, systemic IL-1β increased CGRP neural activity in vivo, and this was significantly attenuated by co-administration of a GIPR agonist. By contrast, GIPR in the dorsal vagal complex was required for the acute anorectic effect of GIPR agonism but not its anti-aversive effect. Taken together, our data suggest that GIPR agonism reduces food intake and prevents aversion via distinct circuits and that GIPR agonism may represent an effective approach to alleviate inflammation-induced aversion.

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 (NAc ms ) 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. Highlights CB1R inactivation attenuates cocaine-evoked DA in NAc ms , preferentially in females 2-AG augmentation via MAGL inhibition enhances cocaine-evoked DA, with female bias Estrous phase modulates the dopamine response to a high dose of cocaine in females Male and female rats show similar baseline DA and locomotor responses to cocaine

Classic theories propose opponent functions for striatal dopamine (DA) and serotonin (5-hydroxytryptamine; 5HT), with DA promoting approach and 5HT promoting patience or avoidance. How these neuromodulators regulate downstream circuits to achieve such antagonistic effects remains mysterious. Here, we mapped striatal 5HT receptor expression in mice to reveal preferential enrichment of putatively excitatory, Gq-coupled 5HT receptors on medium spiny neurons expressing inhibitory D2 DA receptors (D2-MSNs). Acute slice recordings from genetically identified striatal neurons showed that DA and 5HT inversely regulate D2-MSN firing and confirmed that 5HT's excitatory effect on these cells is blocked by 5HT2a and 5HT2c receptor antagonists. Pharmacologically upregulating striatal 5HT release preferentially induced cfos expression in D2-MSNs, validating that 5HT's excitatory effect on these cells also occurs in vivo. Finally, 5HT2c receptor loss-of-function in D2-MSNs, but not D1-MSNs, enhanced animals' sensitivity to the behavioral effects of cocaine - a potent releaser of DA and 5HT - showing that 5HTergic excitation of D2-MSNs counteracts the reinforcing effects of striatal DA release. Altogether these results demonstrate that DA and 5HT inversely modulate D2-MSN activity to regulate cocaine reward, identifying a key circuit mechanism underlying the opponent behavioral effects of these important neuromodulators.

In uncertain, threatening environments, rapid and flexible behavioral decisions are essential for survival. The ventral CA1 region of the hippocampus encodes threatening contexts and can regulate avoidance-related behaviors. While activity patterns in excitatory neurons of vCA1 have been well characterized, the contributions of specific interneuron subtypes to avoidance decisions in threatening situations remain to be fully elucidated. Here, we show that somatostatin expressing (SST) interneurons in vCA1 are selectively recruited during avoidance behavior and play a causal role in shaping behavioral responses in aversive spaces. We find that vCA1 SST interneurons ramp their activity in anticipation of, and during, avoidance. This contrasted with parvalbumin (PV) and vasoactive intestinal peptide (VIP) interneurons, which are preferentially active during exploratory approach behaviors. Unlike other classes of vCA1 interneurons, SST neuron activity more reliably represented the animal’s intention to avoid rather than its spatial position. Moreover, optogenetic silencing of SST interneurons reduced the efficiency of approach-avoidance decisions. These findings identify vCA1 SST interneurons as key regulators of threat assessment, revealing a cell-type-specific mechanism by which vCA1 microcircuits govern avoidance behaviors. This work provides a new framework for understanding hippocampal control of avoidance behavior and highlights SST-expressing interneurons as key contributors to anxiety-related behaviors.

Using external cues to guide behavior is a core function that enables multiple aspects of cognition and attentional control, and deficits in this process are central to many theories of neuro-psychiatric, degenerative, and developmental disorders. Cue detection relies on the precise coordination of neural circuits, with the mediodorsal thalamus (MD) hypothesized to play a pivotal role in orchestrating the relay of cue-based associative information to the prefrontal cortex. The prefrontal cortex comprises multiple subregions, which are believed to differentially contribute to such associative cue-based behaviors. This regional specificity is likely seeded by projection-defined MD→PFC pathways, although the anatomical organization of these discrete channels and their dynamic roles in cue detection are still being defined. Here, we address this gap by combining anatomical circuit mapping of MD-PFC output pathways with in vivo calcium imaging during a cue-based reward conditioning task in mice. These experiments reveal that MD projections to distinct PFC subregions (prelimbic and anterior cingulate cortex) form topographically defined loops, that are characterized by unique patterns of activity across cue-reward learning. Using fiber photometry to monitor changes in calcium activity in axonal projections from the MD to the PFC, we show that during learning, MD projections to the prelimbic subregion are activated by cue presentation, and the dynamics of this activity remain stable across training days. In contrast, MD projections to the anterior cingulate exhibit a learning-dependent suppression of activity that predicts reward approach behavior in late training. Interestingly, the two pathways exhibit opposing activity patterns when the predictive validity of the cue is diminished by extinction training, suggesting distinct functional roles in detecting violations of learned contingencies. Together, these findings reveal previously unrecognized anatomical and functional distinctions within MD-PFC circuits and demonstrate that parallel thalamocortical pathways differentially support cue detection and behavioral flexibility. This work advances understanding of thalamocortical mechanisms underlying cue detection and may inform circuit-based approaches for treating cognitive dysfunction in psychiatric disorders.

The lateral preoptic area (LPO) is a functionally heterogeneous hypothalamic structure that is increasingly recognised for its role in motivated and drug‐seeking behaviours. Although prior studies have shown that LPO neurons exhibit diverse activity patterns during cocaine self‐administration, the specific contributions of glutamatergic and GABAergic populations to conditioned responses to cocaine remain unclear. In this study, we recorded the activity of LPO glutamatergic ( vglut2 ‐expressing) and GABAergic ( vgat ‐expressing) neuronal subpopulations during cocaine conditioning to identify how these cell types respond to drug‐associated cues in awake, behaving mice. Our results revealed that, after cocaine conditioning, both glutamatergic and GABAergic populations showed reduced activity upon entry into the cocaine‐paired chamber. However, only glutamatergic neurons exhibited increased activity when mice entered the saline‐paired chamber. The magnitude of these activity changes was also correlated with behavioural outcomes: reduced GABAergic activity was associated with greater preference for the cocaine‐paired chamber, whereas stronger glutamatergic responses were associated with greater preference for the saline‐paired chamber. These findings indicate that LPO GABAergic and glutamatergic neurons exhibit distinct, cell type–specific patterns during cocaine‐conditioned place preference. Rather than indicating opposing motivational states, the results highlight that LPO activity relates to conditioned behaviours in a cell type– and context‐dependent manner. By linking activity in glutamatergic and GABAergic neurons with conditioned responses, this work provides initial evidence for how LPO circuits may contribute to drug‐associated learning.

Serotonin (5-hydroxytryptamine; 5HT) has long been considered anti-aggressive, but the mechanisms by which 5HT regulates downstream circuits to control aggression remain unclear. Combining fiber photometry, optogenetics, and miniaturized microscope recordings in double-transgenic male mice, we find that 5HT levels ramp up in the nucleus accumbens during aggression, inhibiting a subset of D1 medium spiny neurons to suppress attacks. Our results reveal a novel 5HT-mediated neuromodulatory mechanism for limiting aggressive behavior.

The bed nuclei of the stria terminalis (BST) is a nuclear complex that coordinates neuroendocrine, autonomic, and behavioral responses associated with maintaining homeostasis. Here, we demonstrate that melanocortin 3 receptor (MC3R) neurons in the BST (BST MC3R ) play a key role in modulating feeding behaviors and responses to stressful events. BST MC3R neurons are primarily GABAergic and colocalize with neuropeptides known to regulate feeding and affective behaviors. Chemogenetic activation of BST MC3R neurons causes reduced feeding, particularly in males. BST MC3R neurons are also robustly activated by stress and can control responses to stress and defensive behaviors in both sexes. Whole-brain monosynaptic rabies tracing identified multiple sexually dimorphic inputs to BST MC3R neurons that may contribute to observed functional sexual dimorphisms. Altogether, these data reveal that BST MC3R neurons participate in sexually dimorphic circuits that influence feeding and defensive behaviors, responses to stress, and may represent a potential therapeutic target for stress- and eating-related disorders.

Cytokines and their receptors play important roles in brain development and aging-related disease, but their functions within the healthy adult brain remain poorly understood. Here, we show that pyramidal neurons in hippocampal CA1 induce Fn14 expression in response to activity and environmental enrichment. Once expressed, Fn14 dampens the activity of these neurons most prominently at the daily transition between light and dark. Fn14 expression in CA1 neurons is regulated by the circadian transcription factor Bmal1, and mice lacking Fn14 exhibit disrupted sleep-wake patterns in vivo. At the cellular level, microglia contact fewer excitatory synapses in the absence of Fn14, while neuronal overexpression of Fn14 induces rod-like microglia and recruits them to excitatory synapses. Beyond a homeostatic context, mice lacking Fn14 exhibit heightened susceptibility to chemically induced seizures. These findings reveal that pro-inflammatory cytokine receptors such as Fn14 can play major roles in healthy neurological function in the adult brain.

Reward processes have traditionally been ascribed to dopamine (DA)–associated circuits. While external stimuli, such as food and drugs of abuse, are activators of DA-neuron activity, growing evidence indicates that interoceptive signals also play a critical role. Among these, the gut-brain vagal axis has emerged as a key regulator, although its precise contribution to mesolimbic DA signaling and behavior remains unclear. Here, we combine complementary ex vivo and in vivo approaches across multiple scales to show that gut-brain vagal tone is essential for gating mesolimbic DA system activity and functions, modulating DA-dependent molecular and cellular processes, and scaling both food- and drug-induced reinforcement. These findings challenge the traditional brain-centric view of reward processing, supporting a more integrated model in which vagus-mediated interoceptive signals intrinsically shape motivation and reinforcement. By uncovering the influence of gut-brain vagal communication on DA functions, this work provides insights into the neurobiology of adaptive and maladaptive reward, with broad relevance for eating disorders and addiction.

How positive and negative affective stimuli interact in the brain to influence behavioral outcomes remains poorly understood. Here, we show that recall of a positively valenced reward-associated cue (reward-conditioned stimulus, CS Rew +) can prevent or reverse fear generalization in mice. Modification of generalized fear by recall of a CS Rew + is dependent on the midbrain dopamine system and the regulation of discriminatory fear encoding by the central amygdala (CeA). Precisely timed, transient elevations in dopamine are necessary to reverse fear generalization and nondiscriminatory fear encoding in the CeA. Recall of a positive association is also effective at enhancing the extinction of a conditioned fear response in a dopamine-dependent manner. These data demonstrate that recall of a positive experience can be an effective means to suppress generalized fear and show that dopamine projections to the CeA are an important neural substrate for this phenomenon.

Visual selective attention is not wired into the brain fully formed and immutable but is acquired and refined as animals learn by interacting with their environment. Here we investigated this process by studying neuronal activity in the superior colliculus (SC) of male and female mice that learned different meanings of the same visual stimuli. We recorded spiking activity of neurons across the superficial and deep SC layers in three cohorts of mice, each trained using a tailored set of task rules that attributed different levels of behavioral relevance to the visual stimuli. Experimental conditions across the three tasks were carefully matched for visual stimulation and task engagement. We found that several markers of attention-related activity depended on the level of learned behavioral relevance, with the strongest effects in deep SC. First, for activity evoked by cue onset, visual responses in superficial SC were larger in animals that exhibited higher learned relevance. In deep SC, the presence of visual onset responses depended entirely on the learned behavioral relevance. Second, traditional attention-related modulation, defined as enhanced steady-state activity for the visual stimulus at the cued location, was also stronger with higher learned relevance, and this effect was limited to deep SC. Finally, the response to the visual change at the cued location was virtually absent when learned behavioral relevance was low but was prominent after visual training. Together, these results reveal that several fundamental features of attention-related modulation depend on the behavioral relevance learned through task-specific training, including aspects of stimulus-driven attention.

Interoceptive predictions are internal models that guide behavior by anticipating physiological needs based on past experiences and environmental cues. The paraventricular nucleus of the thalamus (PVT), which receives both interoceptive and exteroceptive inputs, is well-positioned to integrate these signals. Using fiber photometry and bulk calcium imaging in mice performing a reward-based linear maze task, we found that dopamine receptor 2-expressing PVT neurons (PVTD2R+) are engaged in behaviors leading to physiologically relevant outcomes, with their activity highly dependent on the interoceptive state of the mice and the expected outcome. Furthermore, we show that the chronic inhibition of PVTD2R+ neurons impairs the long-term performance of interoceptive-guided motivated behavior. Collectively, these findings identify PVTD2R+ neurons as critical integrators of internal states, external cues, and experiential factors, all of which contribute to interoceptive predictions that facilitate adaptive, goal-directed behavior.

The mammillary body (MB) has traditionally been regarded as a relay station for the hippocampus and plays a pivotal role in the Papez circuit. However, its molecular and cellular organization remains inadequately characterized. This study focuses on the horizontally symmetrically distributed neurotensin (Nts)-expressing and nitric oxide synthase 1 (Nos1)-expressing neurons in the MB, demonstrating that Grik4 (encoding a high-affinity kainate receptor subunit) underlies their distinct electrophysiological properties. Within neural circuits, Nts and Nos1 neurons receive excitatory inputs from the ventral subiculum and send parallel excitatory projections to the dorsomedial and ventrolateral subdivisions of the anteroventral thalamus (AV). These 2 cell type-specific circuits are essential for working memory and exhibit selective activation during the maintenance phase with a marked temporal difference. Together, our findings establish a direct link from molecular identity to circuit architecture and cognitive processing by demonstrating that molecularly distinct Nts and Nos1 neurons constitute differential circuits with convergent inputs, divergent outputs, and dissociable roles in working memory maintenance. This work thus reveals a fundamental cross-scale organizational principle—molecule, cell, circuit, function—within the MB.

The ability to associate environmental stimuli with positive outcomes is a fundamental form of learning. While extensive research has focused on midbrain dopamine neurons during associative learning, less is known about learning-mediated changes in the afferents that shape dopamine neuron responses. We demonstrate in rats that during critical phases of learning, anion homeostasis in midbrain inhibitory GABA neurons - a primary source of input to dopamine neurons - is disrupted due to downregulation of the potassium chloride cotransporter KCC2. This alteration in GABA neurons preferentially impacted lateral mesoaccumbal dopamine pathways and was not observed after learning was established. At the network level, learning-mediated KCC2 downregulation was associated with enhanced synchronization between individual GABA neurons and increased dopamine responses to rewards and reward-related stimuli. Conversely, enhancing KCC2 function during learning reduced GABA synchronization, diminished relevant dopamine signaling, and prevented cue-reward associations. Thus, circuit-specific adaptations in midbrain GABA neurons are crucial for forming new reward-related behaviors.

Drugs of abuse in adolescence impact brain maturation and increase psychiatric risk, with differences in sensitivity between males and females. Amphetamine in early adolescence (postnatal day; PND 21 ± 1–32 ± 1) in male, but not female mice, causes dopamine axons intended to innervate the nucleus accumbens and to grow ectopically to the prefrontal cortex (PFC). This is mediated by drug-induced downregulation of the Netrin-1 receptor DCC. How off-target dopamine axons function in the adult PFC remains to be determined. Here we report that males and females show place preference for amphetamine in early adolescence. However, only in males, amphetamine increases PFC dopamine transporter expression in adulthood (PND 101 ± 15): leading to aberrant baseline dopamine transients, faster dopamine release, and exaggerated responses to acute methylphenidate. Upregulation of DCC in adolescence, using CRISPRa, prevents all these changes. Mesolimbic dopamine axons rerouted to the PFC in adolescence retain anatomical and functional phenotypes of their intended target, rendering males enduringly vulnerable to the harmful effects of drugs of abuse.

The hypothalamus is critical for regulating behaviors essential for survival and locomotion, but how it integrates internal needs and transmits locomotion commands to the spinal cord (SC) remains unclear. We found that glutamatergic neurons in lateral hypothalamic area (LHA) are essential for regulating motivated locomotor activity. Using single-neuron projectome analysis, trans-synaptic tracing, and optogenetic manipulation, we showed that LHA facilitates motivated locomotion during food seeking via pontine reticular nucleus, oral part (PnO) projection neurons, rather than direct SC projections or indirect stress signaling via medial septum and diagonal band. Activating PnO-SC projection neurons also initiated locomotion. Importantly, LHA-PnO projection neurons were crucial for regulating locomotor recovery following mouse spinal cord injury (SCI). Motor cortex signals gated LHA deep brain stimulation treatment markedly promoted long-term restoration of hindlimb motor functions after severe SCI. Thus, we have identified a hypothalamic-pontine-spinal pathway and the stimulation paradigm for potential therapeutic intervention after SCI.