Ventral hippocampal parvalbumin interneurons gate the acute anxiolytic action of the serotonergic psychedelic DOI.

There has been a recent renewal of interest in the therapeutic potential of serotonergic psychedelics. Here, we uncover the essential role of ventral hippocampus (vHpc) GABAergic interneurons in the anxiolytic effect evoked by the serotonergic psychedelic 2,5-dimethoxy-4-iodoamphetamine (DOI). Integrating anatomical, pharmacological, and genetic approaches, we show that 5-HT2A receptors in the CA1/subiculum (CA1/sub) region of the vHpc are required for the anxiolytic action of DOI. In vivo electrophysiology and opto-tagging experiments indicate that DOI enhances the firing rate of hippocampal fast-spiking parvalbumin (PV)-positive interneurons, most of which express the 5-HT2A receptors. Restoration of 5-HT2A receptors in PV-positive interneurons in a loss-of-function background reinstated the anxiolytic responses evoked by DOI in the vHpc CA1/sub region. Collectively, our results localize the acute anxiolytic action of a serotonergic psychedelic to 5-HT2A receptors in the ventral hippocampus and specifically identify PV-positive fast-spiking cells as a cellular trigger for the psychedelic-induced relief of anxiety-like behavior.

Psilocybin Facilitates Fear Extinction: Importance of Dose, Context, and Serotonin Receptors.

A variety of classic psychedelics and MDMA have been shown to enhance fear extinction in rodent models. This has translational significance because a standard treatment for post-traumatic stress disorder (PTSD) is prolonged exposure therapy. However, few studies have investigated psilocybin's potential effect on fear learning paradigms. More specifically, the extents to which dose, timing of administration, and serotonin receptors may influence psilocybin's effect on fear extinction are not understood. In this study, we used a delay fear conditioning paradigm to determine the effects of psilocybin on fear extinction, extinction retention, and fear renewal in male and female mice. Psilocybin robustly enhances fear extinction when given acutely prior to testing for all doses tested. Psilocybin also exerts long-term effects to elevate extinction retention and suppress fear renewal in a novel context, although these changes were sensitive to dose. Analysis of sex differences showed that females may respond to a narrower range of doses than males. Administration of psilocybin prior to fear learning or immediately after extinction yielded no change in behavior, indicating that concurrent extinction experience is necessary for the drug's effects. Cotreatment with a 5-HT2A receptor antagonist blocked psilocybin's effects for extinction, extinction retention, and fear renewal, whereas 5-HT1A receptor antagonism attenuated only the effect on fear renewal. Collectively, these results highlight dose, context, and serotonin receptors as crucial factors in psilocybin's ability to facilitate fear extinction. The study provides preclinical evidence to support investigating psilocybin as a pharmacological adjunct for extinction-based therapy for PTSD.

Classification of psychedelic drugs based on brain-wide imaging of cellular c-Fos expression.

Psilocybin, ketamine, and MDMA are psychoactive compounds that exert behavioral effects with distinguishable but also overlapping features. The growing interest in using these compounds as therapeutics necessitates preclinical assays that can accurately screen psychedelics and related analogs. We posit that a promising approach may be to measure drug action on markers of neural plasticity in native brain tissues. We therefore developed a pipeline for drug classification using light sheet fluorescence microscopy of immediate early gene expression at cellular resolution followed by machine learning. We tested male and female mice with a panel of drugs, including psilocybin, ketamine, 5-MeO-DMT, 6-fluoro-DET, MDMA, acute fluoxetine, chronic fluoxetine, and vehicle. In one-versus-rest classification, the exact drug was identified with 67% accuracy, significantly above the chance level of 12.5%. In one-versus-one classifications, psilocybin was discriminated from 5-MeO-DMT, ketamine, MDMA, or acute fluoxetine with >95% accuracy. We used Shapley additive explanation to pinpoint the brain regions driving the machine learning predictions. Our results support a novel approach for screening psychoactive drugs with psychedelic properties.

Spatiotemporal Organization of Prefrontal Norepinephrine Influences Neuronal Activity.

Norepinephrine (NE), a neuromodulator released by locus ceruleus (LC) neurons throughout the cortex, influences arousal and learning through extrasynaptic vesicle exocytosis. While NE within cortical regions has been viewed as a homogenous field, recent studies have demonstrated heterogeneous axonal dynamics and advances in GPCR-based fluorescent sensors permit direct observation of the local dynamics of NE at cellular scale. To investigate how the spatiotemporal dynamics of NE release in the prefrontal cortex (PFC) affect neuronal firing, we employed in vivo two-photon imaging of layer 2/3 of the PFC in order to observe fine-scale neuronal calcium and NE dynamics concurrently. In this proof of principle study, we found that local and global NE fields can decouple from one another, providing a substrate for local NE spatiotemporal activity patterns. Optic flow analysis revealed putative release and reuptake events which can occur at the same location, albeit at different times, indicating the potential to create a heterogeneous NE field. Utilizing generalized linear models, we demonstrated that cellular Ca2+ fluctuations are influenced by both the local and global NE field. However, during periods of local/global NE field decoupling, the local field drives cell firing dynamics rather than the global field. These findings underscore the significance of localized, phasic NE fluctuations for structuring cell firing, which may provide local neuromodulatory control of cortical activity.

Frontal noradrenergic and cholinergic transients exhibit distinct spatiotemporal dynamics during competitive decision-making.

Norepinephrine (NE) and acetylcholine (ACh) are neuromodulators that are crucial for learning and decision-making. In the cortex, NE and ACh are released at specific sites along neuromodulatory axons, which would constrain their spatiotemporal dynamics at the subcellular scale. However, how the fluctuating patterns of NE and ACh signaling may be linked to behavioral events is unknown. Here, leveraging genetically encoded NE and ACh indicators, we use two-photon microscopy to visualize neuromodulatory signals in the superficial layer of the mouse medial frontal cortex during decision-making. Head-fixed mice engage in a competitive game called matching pennies against a computer opponent. We show that both NE and ACh transients carry information about decision-related variables including choice, outcome, and reinforcer. However, the two neuromodulators differ in their spatiotemporal pattern of task-related activation. Spatially, NE signals are more segregated with choice and outcome encoded at distinct locations, whereas ACh signals can multiplex and reflect different behavioral correlates at the same site. Temporally, task-driven NE transients were more synchronized and peaked earlier than ACh transients. To test functional relevance, using optogenetics we found that evoked elevation of NE, but not ACh, in the medial frontal cortex increases the propensity of the animals to switch and explore alternate options. Taken together, the results reveal distinct spatiotemporal patterns of rapid ACh and NE transients at the subcellular scale during decision-making in mice, which may endow these neuromodulators with different ways to impact neural plasticity to mediate learning and adaptive behavior.

Dynamic Foraging Behavior Performance Is Not Affected by Scn2a Haploinsufficiency.

Dysfunction in the gene SCN2A, which encodes the voltage-gated sodium channel Nav1.2, is strongly associated with neurodevelopmental disorders including autism spectrum disorder and intellectual disability (ASD/ID). This dysfunction typically manifests in these disorders as a haploinsufficiency, where loss of one copy of a gene cannot be compensated for by the other allele. Scn2a haploinsufficiency affects a range of cells and circuits across the brain, including associative neocortical circuits that are important for cognitive flexibility and decision-making behaviors. Here, we tested whether Scn2a haploinsufficiency has any effect on a dynamic foraging task that engages such circuits. Scn2a +/- mice and wild-type (WT) littermates were trained on a choice behavior where the probability of reward between two options varied dynamically across trials and where the location of the high reward underwent uncued reversals. Despite impairments in Scn2a-related neuronal excitability, we found that both male and female Scn2a +/- mice performed these tasks as well as wild-type littermates, with no behavioral difference across genotypes in learning or performance parameters. Varying the number of trials between reversals or probabilities of receiving reward did not result in an observable behavioral difference, either. These data suggest that, despite heterozygous loss of Scn2a, mice can perform relatively complex foraging tasks that make use of higher-order neuronal circuits.

Spatiotemporal organization of prefrontal norepinephrine influences neuronal activity.

Norepinephrine (NE), a neuromodulator released by locus coeruleus neurons throughout cortex, influences arousal and learning through extra-synaptic vesicle exocytosis. While NE within cortical regions has been viewed as a homogenous field, recent studies have demonstrated heterogeneous axonal dynamics and advances in GPCR-based fluorescent sensors permit direct observation of the local dynamics of NE at cellular scale. To investigate how the spatiotemporal dynamics of NE release in the PFC affect neuronal firing, we employed in-vivo two-photon imaging of layer 2/3 of PFC in order to observe fine-scale neuronal calcium and NE dynamics concurrently. We found that local and global NE fields can decouple from one another, providing a substrate for local NE spatiotemporal activity patterns. Optic flow analysis revealed putative release and reuptake events which can occur at the same location, albeit at different times, indicating the potential to create a heterogeneous NE field. Utilizing generalized linear models, we demonstrated that cellular Ca2+ fluctuations are influenced by both the local and global NE field. However, during periods of local/global NE field decoupling, the local field drives cell firing dynamics rather than the global field. These findings underscore the significance of localized, phasic NE fluctuations for structuring cell firing, which may provide local neuromodulatory control of cortical activity.

Hyperconnectivity of Two Separate Long-Range Cholinergic Systems Contributes to the Reorganization of the Brain Functional Connectivity during Nicotine Withdrawal in Male Mice.

Chronic nicotine results in dependence with withdrawal symptoms on discontinuation of use, through desensitization of nicotinic acetylcholine receptors and altered cholinergic neurotransmission. Nicotine withdrawal is associated with increased whole-brain functional connectivity and decreased network modularity; however, the role of cholinergic neurons in those changes is unknown. To identify the contribution of nicotinic receptors and cholinergic regions to changes in the functional network, we analyzed the contribution of the main cholinergic regions to brain-wide activation of the immediate early-gene Fos during withdrawal in male mice and correlated these changes with the expression of nicotinic receptor mRNA throughout the brain. We show that the main functional connectivity modules included the main long-range cholinergic regions, which were highly synchronized with the rest of the brain. However, despite this hyperconnectivity, they were organized into two anticorrelated networks that were separated into basal forebrain-projecting and brainstem-thalamic-projecting cholinergic regions, validating a long-standing hypothesis of the organization of the brain cholinergic systems. Moreover, baseline (without nicotine) expression of Chrna2, Chrna3, Chrna10, and Chrnd mRNA of each brain region correlated with withdrawal-induced changes in Fos expression. Finally, by mining the Allen Brain mRNA expression database, we were able to identify 1755 gene candidates and three pathways (Sox2-Oct4-Nanog, JAK-STAT, and MeCP2-GABA) that may contribute to nicotine withdrawal-induced Fos expression. These results identify the dual contribution of the basal forebrain and brainstem-thalamic cholinergic systems to whole-brain functional connectivity during withdrawal; and identify nicotinic receptors and novel cellular pathways that may be critical for the transition to nicotine dependence.

A computational model for learning from repeated traumatic experiences under uncertainty.

Traumatic events can lead to lifelong, inflexible adaptations in threat perception and behavior, which characterize posttraumatic stress disorder (PTSD). This process involves associations between sensory cues and internal states of threat and then generalization of the threat responses to previously neutral cues. However, most formulations neglect adaptations to threat that are not specific to those associations. To incorporate nonassociative responses to threat, we propose a computational theory of PTSD based on adaptation to the frequency of traumatic events by using a reinforcement learning momentum model. Recent threat prediction errors generate momentum that influences subsequent threat perception in novel contexts. This model fits primary data acquired from a mouse model of PTSD, in which unpredictable footshocks in one context accelerate threat learning in a novel context. The theory is consistent with epidemiological data that show that PTSD incidence increases with the number of traumatic events, as well as the disproportionate impact of early life trauma. Because the theory proposes that PTSD relates to the average of recent threat prediction errors rather than the strength of a specific association, it makes novel predictions for the treatment of PTSD.

Competitive and cooperative games for probing the neural basis of social decision-making in animals.

In a social environment, it is essential for animals to consider the behavior of others when making decisions. To quantitatively assess such social decisions, games offer unique advantages. Games may have competitive and cooperative components, modeling situations with antagonistic and shared objectives between players. Games can be analyzed by mathematical frameworks, including game theory and reinforcement learning, such that an animal's choice behavior can be compared against the optimal strategy. However, so far games have been underappreciated in neuroscience research, particularly for rodent studies. In this review, we survey the varieties of competitive and cooperative games that have been tested, contrasting strategies employed by non-human primates and birds with rodents. We provide examples of how games can be used to uncover neural mechanisms and explore species-specific behavioral differences. We assess critically the limitations of current paradigms and propose improvements. Together, the synthesis of current literature highlights the advantages of using games to probe the neural basis of social decisions for neuroscience studies.

5-MeO-DMT modifies innate behaviors and promotes structural neural plasticity in mice.

Serotonergic psychedelics are gaining increasing interest as potential therapeutics for a range of mental illnesses. Compounds with short-lived subjective effects may be clinically useful because dosing time would be reduced, which may improve patient access. One short-acting psychedelic is 5-MeO-DMT, which has been associated with improvement in depression and anxiety symptoms in early phase clinical studies. However, relatively little is known about the behavioral and neural mechanisms of 5-MeO-DMT, particularly the durability of its long-term effects. Here we characterized the effects of 5-MeO-DMT on innate behaviors and dendritic architecture in mice. We showed that 5-MeO-DMT induces a dose-dependent increase in head-twitch response that is shorter in duration than that induced by psilocybin at all doses tested. 5-MeO-DMT also substantially suppresses social ultrasonic vocalizations produced during mating behavior. 5-MeO-DMT produces long-lasting increases in dendritic spine density in the mouse medial frontal cortex that are driven by an elevated rate of spine formation. However, unlike psilocybin, 5-MeO-DMT did not affect the size of dendritic spines. These data provide insights into the behavioral and neural consequences underlying the action of 5-MeO-DMT and highlight similarities and differences with those of psilocybin.

Hyperconnectivity of two separate long-range cholinergic systems contributes to the reorganization of the brain functional connectivity during nicotine withdrawal in male mice.

Chronic nicotine results in dependence with withdrawal symptoms upon discontinuation of use, through desensitization of nicotinic acetylcholine receptors and altered cholinergic neurotransmission. Nicotine withdrawal is associated with increased whole-brain functional connectivity and decreased network modularity, however, the role of cholinergic neurons in those changes is unknown. To identify the contribution of nicotinic receptors and cholinergic regions to changes in the functional network, we analyzed the contribution of the main cholinergic regions to brain-wide activation of the immediate early-gene FOS during withdrawal in male mice and correlated these changes with the expression of nicotinic receptor mRNA throughout the brain. We show that the main functional connectivity modules included the main long-range cholinergic regions, which were highly synchronized with the rest of the brain. However, despite this hyperconnectivity they were organized into two anticorrelated networks that were separated into basal forebrain projecting and brainstem-thalamic projecting cholinergic regions, validating a long-standing hypothesis of the organization of the brain cholinergic systems. Moreover, baseline (without nicotine) expression of Chrna2 , Chrna3 , Chrna10 , and Chrnd mRNA of each brain region correlated with withdrawal-induced changes in FOS expression. Finally, by mining the Allen Brain mRNA expression database, we were able to identify 1755 gene candidates and three pathways (Sox2-Oct4-Nanog, JAK-STAT, and MeCP2-GABA) that may contribute to nicotine withdrawal-induced FOS expression. These results identify the dual contribution of the basal forebrain and brainstem-thalamic cholinergic systems to whole-brain functional connectivity during withdrawal; and identify nicotinic receptors and novel cellular pathways that may be critical for the transition to nicotine dependence. Significance Statement: Discontinuation of nicotine use in dependent users is associated with increased whole-brain activation and functional connectivity and leads to withdrawal symptoms. Here we investigated the contribution of the nicotinic cholinergic receptors and main cholinergic projecting brain areas in the whole-brain changes associated with withdrawal. This not only allowed us to visualize and confirm the previously described duality of the cholinergic brain system using this novel methodology, but also identify nicotinic receptors together with 1751 other genes that contribute, and could thus be targets for treatments against, nicotine withdrawal and dependence.

Shared and Distinct Brain Regions Targeted for Immediate Early Gene Expression by Ketamine and Psilocybin.

Psilocybin is a psychedelic with therapeutic potential. While there is growing evidence that psilocybin exerts its beneficial effects through enhancing neural plasticity, the exact brain regions involved are not completely understood. Determining the impact of psilocybin on plasticity-related gene expression throughout the brain can broaden our understanding of the neural circuits involved in psychedelic-evoked neural plasticity. In this study, whole-brain serial two-photon microscopy and light sheet microscopy were employed to map the expression of the immediate early gene, c-Fos, in male and female mice. The drug-induced c-Fos expression following psilocybin administration was compared to that of subanesthetic ketamine and saline control. Psilocybin and ketamine produced acutely comparable elevations in c-Fos expression in numerous brain regions, including anterior cingulate cortex, locus coeruleus, primary visual cortex, central and basolateral amygdala, medial and lateral habenula, and claustrum. Select regions exhibited drug-preferential differences, such as dorsal raphe and insular cortex for psilocybin and the CA1 subfield of hippocampus for ketamine. To gain insights into the contributions of receptors and cell types, the c-Fos expression maps were related to brain-wide in situ hybridization data. The transcript analyses showed that the endogenous levels of Grin2a and Grin2b predict whether a cortical region is sensitive to drug-evoked neural plasticity for both ketamine and psilocybin. Collectively, the systematic mapping approach produced an unbiased list of brain regions impacted by psilocybin and ketamine. The data are a resource that highlights previously underappreciated regions for future investigations. Furthermore, the robust relationships between drug-evoked c-Fos expression and endogenous transcript distributions suggest glutamatergic receptors as a potential convergent target for how psilocybin and ketamine produce their rapid-acting and long-lasting therapeutic effects.

Psychedelics and Neural Plasticity: Therapeutic Implications.

Psychedelic drugs have reemerged as tools to treat several brain disorders. Cultural attitudes toward them are changing, and scientists are once again investigating the neural mechanisms through which these drugs impact brain function. The significance of this research direction is reflected by recent work, including work presented by these authors at the 2022 meeting of the Society for Neuroscience. As of 2022, there were hundreds of clinical trials recruiting participants for testing the therapeutic effects of psychedelics. Emerging evidence suggests that psychedelic drugs may exert some of their long-lasting therapeutic effects by inducing structural and functional neural plasticity. Herein, basic and clinical research attempting to elucidate the mechanisms of these compounds is showcased. Topics covered include psychedelic receptor binding sites, effects of psychedelics on gene expression, and on dendrites, and psychedelic effects on microcircuitry and brain-wide circuits. We describe unmet clinical needs and the current state of translation to the clinic for psychedelics, as well as other unanswered basic neuroscience questions addressable with future studies.

Activity of a direct VTA to ventral pallidum GABA pathway encodes unconditioned reward value and sustains motivation for reward.

Dopamine signaling from the ventral tegmental area (VTA) plays critical roles in reward-related behaviors, but less is known about the functions of neighboring VTA GABAergic neurons. We show here that a primary target of VTA GABA projection neurons is the ventral pallidum (VP). Activity of VTA-to-VP-projecting GABA neurons correlates consistently with size and palatability of the reward and does not change following cue learning, providing a direct measure of reward value. Chemogenetic stimulation of this GABA projection increased activity of a subset of VP neurons that were active while mice were seeking reward. Optogenetic stimulation of this pathway improved performance in a cue-reward task and maintained motivation to work for reward over days. This VTA GABA projection provides information about reward value directly to the VP, likely distinct from the prediction error signal carried by VTA dopamine neurons.

The neural basis of psychedelic action.

Psychedelics are serotonin 2A receptor agonists that can lead to profound changes in perception, cognition and mood. In this review, we focus on the basic neurobiology underlying the action of psychedelic drugs. We first discuss chemistry, highlighting the diversity of psychoactive molecules and the principles that govern their potency and pharmacokinetics. We describe the roles of serotonin receptors and their downstream molecular signaling pathways, emphasizing key elements for drug discovery. We consider the impact of psychedelics on neuronal spiking dynamics in several cortical and subcortical regions, along with transcriptional changes and sustained effects on structural plasticity. Finally, we summarize neuroimaging results that pinpoint effects on association cortices and thalamocortical functional connectivity, which inform current theories of psychedelic action. By synthesizing knowledge across the chemical, molecular, neuronal, and network levels, we hope to provide an integrative perspective on the neural mechanisms responsible for the acute and enduring effects of psychedelics on behavior.

Pupil Correlates of Decision Variables in Mice Playing a Competitive Mixed-Strategy Game.

In a competitive game involving an animal and an opponent, the outcome is contingent on the choices of both players. To succeed, the animal must continually adapt to competitive pressure, or else risk being exploited and lose out on rewards. In this study, we demonstrate that head-fixed male mice can be trained to play the iterative competitive game "matching pennies" against a virtual computer opponent. We find that the animals' performance is well described by a hybrid computational model that includes Q-learning and choice kernels. Comparing between matching pennies and a non-competitive two-armed bandit task, we show that the tasks encourage animals to operate at different regimes of reinforcement learning. To understand the involvement of neuromodulatory mechanisms, we measure fluctuations in pupil size and use multiple linear regression to relate the trial-by-trial transient pupil responses to decision-related variables. The analysis reveals that pupil responses are modulated by observable variables, including choice and outcome, as well as latent variables for value updating, but not action selection. Collectively, these results establish a paradigm for studying competitive decision-making in head-fixed mice and provide insights into the role of arousal-linked neuromodulation in the decision process.

Psychedelics.

Psychedelics are compounds that alter consciousness by acting on serotonin receptors in the brain. The term 'psychedelic', from the Greek for mind manifesting, refers to the drugs' subjective effects and was first proposed by Humphry Osmond in 1956. Other terms have been used to emphasize different aspects of the psychological experiences produced by various related compounds, including hallucinogens (perceptual), entheogens (spiritual), and empathogens or entactogens (social/emotional). The diversity in terminology reflects the existence of hundreds of potential psychedelic compounds with a spectrum of behavioral and neurobiological effects. Recent data on the effectiveness of psychedelics for treating mental illnesses has led to a resurgence of interest in their neurobiological effects. The purpose of this Primer is to provide those interested in the field of psychedelics with a concise and accessible overview of the scientific data.