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In the classical model of the basal ganglia, direct pathway striatal projection neurons (dSPNs) send projections to the substantia nigra (SNr) and entopeduncular nucleus to regulate motor function. Recent studies have re-established that dSPNs also possess “bridging” collaterals within the globus pallidus (GPe), yet the significance of these collaterals for behavior is unknown. Here we use in vivo optical and chemogenetic tools combined with deep learning approaches to dissect the roles of bridging collaterals in motor function. We find that dSPNs projecting to the SNr send synchronous motor-related information to the GPe via axon collaterals. Inhibition of native activity in dSPN GPe terminals impairs motor activity and function via regulation of pallidostriatal Npas1 neurons. We propose a model by which dSPN GPe collaterals (“striatopallidal Go pathway”) act in concert with the canonical terminals in the SNr to support motor control by inhibiting Npas1 signals going back to the striatum.

Cholinergic interneurons (CINs) in the striatum respond to salient stimuli with a multiphasic response, including a pause, in neuronal activity. Slice-physiology experiments have shown the importance of dopamine D2 receptors (D2Rs) in regulating CIN pausing, yet the behavioral significance of the CIN pause and its regulation by dopamine in vivo is still unclear. Here, we show that D2R upregulation in CINs of the nucleus accumbens (NAc) lengthens the pause in CIN activity ex vivo and enlarges a stimulus-evoked decrease in acetylcholine (ACh) levels during behavior. This enhanced dip in ACh levels is associated with a selective deficit in the learning to inhibit responding in a Go/No-Go task. Our data demonstrate, therefore, the importance of CIN D2Rs in modulating the CIN response induced by salient stimuli and point to a role of this response in inhibitory learning. This work has important implications for brain disorders with altered striatal dopamine and ACh function, including schizophrenia and attention-deficit hyperactivity disorder (ADHD).

It was first posited, more than five decades ago, that the etiology of schizophrenia involves overstimulation of dopamine receptors. Since then, advanced clinical research methods, including brain imaging, have refined our understanding of the relationship between striatal dopamine and clinical phenotypes as well as disease trajectory. These studies point to striatal dopamine D2 receptors, the main target for all current antipsychotic medications, as being involved in both positive and negative symptoms. Simultaneously, animal models have been central to investigating causal relationships between striatal dopamine D2 receptors and behavioral phenotypes relevant to schizophrenia. We begin this article by reviewing the circuit, cell-type and subcellular locations of dopamine D2 receptors and their downstream signaling pathways. We then summarize results from several mouse models in which D2 receptor levels were altered in various brain regions, cell-types and developmental periods. Behavioral, electrophysiological and anatomical consequences of these D2 receptor perturbations are reviewed with a selective focus on striatal circuit function and alterations in motivated behavior, a core negative symptom of schizophrenia. These studies show that D2 receptors serve distinct physiological roles in different cell types and at different developmental time points, regulating motivated behaviors in sometimes opposing ways. We conclude by considering the clinical implications of this complex regulation of striatal circuit function by D2 receptors.

Ventral striatal dopamine is thought to be important for associative learning. Dopamine exerts its role via activation of dopamine D1 and D2 receptors in the ventral striatum. Upregulation of dopamine D2R in ventral striatopallidal neurons impairs incentive motivation via inhibiting synaptic transmission to the ventral pallidum. Here, we determined whether upregulation of D2Rs and the resulting impairment in ventral striatopallidal pathway function modulates associative learning in an auditory Pavlovian reward learning task as well as Go/No-Go learning in an operant based reward driven Go/No-Go task. We found that upregulation of D2Rs did not affect Pavlovian learning or the extinction of Pavlovian responses, and neither did it alter No-Go learning. We however observed a delay in the Go component of the task which may indicate a deficit in learning though it could also be attributed to the established locomotor hyperactivity of the mice. In combination with previously published findings, our data suggest that D2Rs in ventral striatopallidal neurons play a specific role in regulating motivation by balancing cost/benefit computations but do not necessarily affect associative learning. (PsycInfo Database Record (c) 2021 APA, all rights reserved).

The ability to rapidly adapt to novel situations is essential for survival, and this flexibility is impaired in many neuropsychiatric disorders1. Thus, understanding whether and how novelty prepares, or primes, brain circuitry to facilitate cognitive flexibility has important translational relevance. Exposure to novelty recruits the hippocampus and medial prefrontal cortex (mPFC)2 and may prime hippocampal–prefrontal circuitry for subsequent learning-associated plasticity. Here we show that novelty resets the neural circuits that link the ventral hippocampus (vHPC) and the mPFC, facilitating the ability to overcome an established strategy. Exposing mice to novelty disrupted a previously encoded strategy by reorganizing vHPC activity to local theta (4–12 Hz) oscillations and weakening existing vHPC–mPFC connectivity. As mice subsequently adapted to a new task, vHPC neurons developed new task-associated activity, vHPC–mPFC connectivity was strengthened, and mPFC neurons updated to encode the new rules. Without novelty, however, mice adhered to their established strategy. Blocking dopamine D1 receptors (D1Rs) or inhibiting novelty-tagged cells that express D1Rs in the vHPC prevented these behavioural and physiological effects of novelty. Furthermore, activation of D1Rs mimicked the effects of novelty. These results suggest that novelty promotes adaptive learning by D1R-mediated resetting of vHPC–mPFC circuitry, thereby enabling subsequent learning-associated circuit plasticity. Exposure to a novel experience can ‘reset’ connections between the hippocampus and prefrontal cortex in mice, allowing them to overcome an existing learned behaviour and to replace it with a new one.

The increasing interest in manipulating neural circuits in developing brains has created a demand for reliable and accurate methods for delivering viruses to newborn mice. Here we describe a novel 3D-printed mouse neonatal stereotaxic adaptor for intracerebral viral injection that provides enhanced precision and reliability. Using this device, we injected A2a-Cre mice with a Cre-dependent hM4D-mCherry viral construct at postnatal day 1 (P1) and demonstrated selective expression in the striatal indirect pathway neurons on days P7, P11 and P25. Similarly, dopaminergic midbrain neurons were selectively targeted with a Cre-dependent green fluorescent protein virus in Dat-IRES-Cre neonates and expression examined at P25. Our open-source neonatal stereotaxic mouse adaptor facilitates neonatal neuronal targeting, which should improve the ability to label and modify neural circuits in developing mouse brains., METHOD SUMMARY The neonatal mouse stereotaxic adaptor, designed and easily 3D printed in-house, was fitted into the ear and tooth bars of a mouse digital stereotaxic apparatus and held the head of the ice-anesthetized newborn pups in a cone-shaped cradle during the virus injection procedure. After the secured heads were balanced on the AP and ML axes by adjusting the ear and tooth bars, Cre-dependent viruses (AAV5-hSyn-DIO-hM4D-mCherry or AAV5-hSyn-DIO-GFP) were injected to the desired brain region of Cre-expressing transgenic mice, while the animals were partially covered in ice. After completion of each surgery, the neonate was removed from the adaptor and returned to the den for recovery. Neuroanatomical viral expression was verified by immunofluorescence 6, 10 and 24 days after the procedure.

Low dopamine D2 receptor (D2R) availability in the striatum can predispose for cocaine abuse; though how low striatal D2Rs facilitate cocaine reward is unclear. Overexpression of D2Rs in striatal neurons or activation of D2Rs by acute cocaine suppresses striatal Penk mRNA. Conversely, low D2Rs in D2-striatal neurons increases striatal Penk mRNA and enkephalin peptide tone, an endogenous mu-opioid agonist. In brain slices, met-enkephalin and inhibition of enkephalin catabolism suppresses intra-striatal GABA transmission. Pairing cocaine with intra-accumbens met-enkephalin during place conditioning facilitates acquisition of preference, while mu-opioid receptor antagonist blocks preference in wild-type mice. We propose that heightened striatal enkephalin potentiates cocaine reward by suppressing intra-striatal GABA to enhance striatal output. Surprisingly, a mu-opioid receptor antagonist does not block cocaine preference in mice with low striatal D2Rs, implicating other opioid receptors. The bidirectional regulation of enkephalin by D2R activity and cocaine offers insights into mechanisms underlying the vulnerability for cocaine abuse.

In the striatum, acetylcholine (ACh) neuron activity is modulated co-incident with dopamine (DA) release in response to unpredicted rewards and reward predicting cues and both neuromodulators are thought to regulate each other. While this co-regulation has been studied using stimulation studies, the existence of this mutual regulation in vivo during natural behavior is still largely unexplored. One long-standing controversy has been whether striatal DA is responsible for the induction of the cholinergic pause or whether D2R modulate a pause that is induced by other mechanisms. Here, we used genetically encoded sensors in combination with pharmacological and genetic inactivation of D2Rs from cholinergic interneurons (CINs) to simultaneously measure ACh and DA levels after CIN D2R inactivation. We found that CIN D2Rs are not necessary for the induction of cue induced dips in ACh levels but regulate dip lengths and rebound ACh levels. Importantly, D2R inactivation strongly decreased the temporal correlation between DA and Ach signals not only at cue presentation but also during the intertrial interval. This points to a general mechanism by which D2Rs coordinate both signals. At the behavioral level D2R antagonism increased the latency to lever press, which was not observed in CIN-selective D2R knock out mice. This latency correlated with the cue evoked dip length supporting a role of the ACh dip and it’s regulation by D2Rs in motivated behavior. Overall, our data indicate that striatal DA coordinate phasic ACh and DA signals via CIN D2Rs which is important for the regulation of motivated behavior.

Adolescence is a period of increased vulnerability for the development of psychiatric disorders, including schizophrenia. The prefrontal cortex (PFC) undergoes substantial maturation during this period, and PFC dysfunction is central to cognitive impairments in schizophrenia. As a result, impaired adolescent maturation of the PFC has been proposed as a mechanism in the etiology of the disorder and its cognitive symptoms. In adulthood, PFC function is tightly linked to its reciprocal connections with the thalamus, and acutely inhibiting thalamic inputs to the PFC produces impairments in PFC function and cognitive deficits. Here, we propose that thalamic activity is equally important during adolescence because it is required for proper PFC circuit development. Because thalamic abnormalities have been observed early in the progression of schizophrenia, we further postulate that adolescent thalamic dysfunction can have long-lasting consequences for PFC function and cognition in patients with schizophrenia.

We utilized forebrain organoids generated from induced pluripotent stem cells of patients with a syndromic form of Autism Spectrum Disorder (ASD) with a homozygous protein-truncating mutation in CNTNAP2, to study its effects on embryonic cortical development. Patients with this mutation present with clinical characteristics of brain overgrowth. Patient-derived forebrain organoids displayed an increase in volume and total cell number that is driven by increased neural progenitor proliferation. Single-cell RNA sequencing revealed PFC-excitatory neurons to be the key cell types expressing CNTNAP2. Gene ontology analysis of differentially expressed genes (DEgenes) corroborates aberrant cellular proliferation. Moreover, the DEgenes are enriched for ASD-associated genes. The cell-type-specific signature genes of the CNTNAP2-expressing neurons are associated with clinical phenotypes previously described in patients. The organoid overgrowth phenotypes were largely rescued after correction of the mutation using CRISPR-Cas9. This CNTNAP2-organoid model provides opportunity for further mechanistic inquiry and development of new therapeutic strategies for ASD., Mutations in CNTNAP2 have been associated with a syndromic form of Autism Spectrum Disorder. Here the authors show that forebrain organoids generated from induced pluripotent stem cells of patients with a syndromic form of ASD with a homozygous truncating mutation in CNTNAP2 displayed an increase in volume and total cell number, which is driven by abnormal cellular proliferation and neurogenesis.

Impaired cortical maturation is a postulated mechanism in the etiology of neurodevelopmental disorders, including schizophrenia. In sensory cortex, activity relayed by the thalamus during a postnatal sensitive period is essential for proper cortical maturation. Whether thalamic activity also shapes prefrontal cortical maturation is unknown. Here, we show that inhibiting the midline thalamus during adolescence leads to a long-lasting decrease in thalamo-prefrontal projection density and cortical excitation. Adolescent thalamic inhibition also causes prefrontal-dependent cognitive deficits during adulthood that are associated with disrupted prefrontal cross-correlations and task outcome encoding. In contrast, thalamic inhibition during adulthood has no long-lasting consequences. Strikingly, exciting the thalamus in adulthood during a cognitive task rescues prefrontal cross-correlations, task outcome encoding, and cognitive deficits. These data point to adolescence as a sensitive window of thalamo-cortical circuit maturation. Furthermore, by supporting prefrontal network activity, boosting thalamic activity provides a potential therapeutic strategy for rescuing cognitive deficits in neurodevelopmental disorders.

Impaired cortical maturation is a postulated mechanism in the etiology of neurodevelopmental disorders, including schizophrenia. In the sensory cortex, activity relayed by the thalamus during a postnatal sensitive period is essential for proper cortical maturation. Whether thalamic activity also shapes prefrontal cortical maturation is unknown. We show that inhibiting the mediodorsal and midline thalamus in mice during adolescence leads to a long-lasting decrease in thalamo-prefrontal projection density and reduced excitatory drive to prefrontal neurons. It also caused prefrontal-dependent cognitive deficits during adulthood associated with disrupted prefrontal cross-correlations and task outcome encoding. Thalamic inhibition during adulthood had no long-lasting consequences. Exciting the thalamus in adulthood during a cognitive task rescued prefrontal cross-correlations, task outcome encoding and cognitive deficits. These data point to adolescence as a sensitive window of thalamocortical circuit maturation. Furthermore, by supporting prefrontal network activity, boosting thalamic activity provides a potential therapeutic strategy for rescuing cognitive deficits in neurodevelopmental disorders.

The medial prefrontal cortex (mPFC) integrates inputs from multiple subcortical regions including the mediodorsal nucleus of the thalamus (MD) and the ventral hippocampus (vHPC). How the mPFC differentially processes these inputs is not known. One possibility is that these two inputs target discreet populations of mPFC cells. Alternatively, individual prefrontal cells could receive convergent inputs but distinguish between both inputs based on synaptic differences, such as communication frequency. To address this, we utilized a dual wavelength optogenetic approach to stimulate MD and vHPC inputs onto single, genetically defined mPFC neuronal subtypes. Specifically, we compared the convergence and synaptic dynamics of both inputs onto mPFC pyramidal cells, and parvalbumin (PV)- and vasoactive intestinal peptide (VIP)-expressing interneurons. We found that all individual pyramidal neurons in layer 2/3 of the mPFC receive convergent input from both MD and vHPC. In contrast, PV neurons receive input biased from the MD, while VIP cells receive input biased from the vHPC. Independent of the target, MD inputs transferred information more reliably at higher frequencies (20 Hz) than vHPC inputs. Thus, MD and vHPC projections converge functionally onto mPFC pyramidal cells, but both inputs are distinguished by frequency-dependent synaptic dynamics and preferential engagement of discreet interneuron populations.

Cholinergic interneurons (CINs) in the striatum respond to salient stimuli with a multiphasic response, including a pause, in neuronal activity. Slice-physiology experiments have shown the importance of dopamine D2 receptors (D2Rs) in regulating CIN pausing, yet the behavioral significance of the CIN pause and its regulation by dopamine in vivo is still unclear. Here, we show that D2R upregulation in CINs of the nucleus accumbens (NAc) lengthens the pause in CIN activity ex vivo and enlarges a stimulus-evoked decrease in acetylcholine (ACh) levels during behavior. This enhanced dip in ACh levels is associated with a selective deficit in the learning to inhibit responding in a Go/No-Go task. Our data demonstrate, therefore, the importance of CIN D2Rs in modulating the CIN response induced by salient stimuli and point to a role of this response in inhibitory learning. This work has important implications for brain disorders with altered striatal dopamine and ACh function, including schizophrenia and attention-deficit hyperactivity disorder (ADHD).

Cholinergic interneurons (CINs) in the striatum respond to salient stimuli with a multiphasic response, including a pause, in neuronal activity. Slice physiology experiments have shown the importance of dopamine D2 receptors (D2Rs) in regulating CIN pausing yet the behavioral significance of the CIN pause and its regulation by dopamine in vivo is still unclear. Here, we show that D2R upregulation in CINs of the nucleus accumbens (NAc) lengthens the pause in CIN activity ex vivo and enlarges a stimulus-evoked decrease in acetylcholine (ACh) levels during behavior. This enhanced dip in ACh levels is associated with a selective deficit in the learning to inhibit responding in a Go/No-Go task. Our data demonstrate, therefore, the importance of CIN D2Rs in modulating the CIN response induced by salient stimuli and points to a role of the pause in inhibitory learning. This work has important implications for brain disorders with altered striatal dopamine and ACh function, including schizophrenia and attention-deficit hyperactivity disorder (ADHD).

Ventral striatal dopamine is thought to be important for associative learning. Dopamine exerts its role via activation of dopamine D1 and D2 receptors in the ventral striatum. Upregulation of dopamine D2R in the indirect pathway of the nucleus accumbens (NAc) impairs incentive motivation via inhibiting synaptic transmission to the ventral pallidum. Here, we determined whether upregulation of D2Rs and the resulting impairment in indirect pathway function modulates associative learning in an auditory Pavlovian reward learning task as well as Go/No-Go learning in an operant based reward driven Go/No-Go task. We found that upregulation of D2Rs in indirect pathway neurons of the NAc did not affect Pavlovian learning or the extinction of Pavlovian responses, and neither did it alter No-Go learning. A delay in the Go component of the task however could indicate a deficit in learning though it may be attributed to locomotor hyperactivity of the mice. In combination with previously published findings our data suggest that D2Rs in the NAc core play a specific role in regulating motivation by balancing cost/benefit computations but do not necessarily affect associative learning.

Cognitive functions, such as working memory, are disrupted in most psychiatric disorders. Many of these processes are believed to depend on the medial prefrontal cortex (mPFC). Traditionally, maze-based behavioral tasks, which have a strong exploratory component, have been used to study the role of the mPFC in working memory in mice. In maze tasks, mice navigate through the environment and require a significant amount of time to complete each trial, thereby limiting the number of trials that can be run per day. Here, we show that an operant-based delayed nonmatch to sample (DNMS) working memory task, with shorter trial lengths and a smaller exploratory component, is also mPFC-dependent. We created excitotoxic lesions in the mPFC of mice and found impairments in both the acquisition of the task, with no delay, and in the performance with delays introduced. Importantly, we saw no differences in trial length, reward collection, or lever-press latencies, indicating that the difference in performance was not due to a change in motivation or mobility. Using this operant DNMS task will facilitate the analysis of working memory and improve our understanding of the physiology and circuit mechanisms underlying this cognitive process. (PsycInfo Database Record (c) 2020 APA, all rights reserved).

Anorexia nervosa (AN) is an eating disorder observed predominantly in women and girls that is characterized by a low body-mass index, hypophagia, and hyperactivity. Activity-based anorexia (ABA), which refers to the weight loss, hypophagia, and hyperactivity exhibited by rodents exposed to both running wheels and scheduled fasting, provides a model for aspects of AN. Increased dopamine D2/D3 receptor binding in the anteroventral striatum has been reported in AN patients. We virally overexpressed D2Rs on nucleus accumbens core (D2R-OENac) neurons that endogenously express D2Rs, and tested mice of both sexes in the open field test, ABA paradigm, and intraperitoneal glucose tolerance test (IGTT). D2R-OENac did not alter baseline body weight, but increased locomotor activity in the open field across both sexes. During constant access to food and running wheels, D2R-OENac mice of both sexes increased food intake and ran more than controls. However, when food was available only 7 hours a day, only female D2R-OENac mice rapidly lost 25% of their initial body weight, reduced food intake, and substantially increased wheel running. Surprisingly, female D2R-OENac mice also rapidly lost 25% of their initial body weight during scheduled fasting without wheel access and showed no changes in food intake. In contrast, male D2R-OENac mice maintained body weight during scheduled fasting. D2R-OENac mice of both sexes also showed glucose intolerance in the IGTT. In conclusion, D2R-OENac alters glucose metabolism in both sexes but drives robust weight loss only in females during scheduled fasting, implicating metabolic mechanisms in this sexually dimorphic effect.

Deficits in cognition are a core feature of many psychiatric conditions, including schizophrenia, where the severity of such deficits is a strong predictor of long-term outcome. Impairment in cognitive domains, such as working memory and behavioral flexibility, have classically been associated with prefrontal cortex (PFC) dysfunction. However, there is increasing evidence that the PFC cannot be dissociated from its main thalamic counterpart, the mediodorsal thalamus (MD). Since the causal relationships between MD-PFC abnormalities and cognitive impairment, as well as the neuronal mechanisms underlying them, are difficult to address in humans, animal models have been employed for mechanistic insight. In this review, we discuss anatomical, behavioral, and electrophysiological findings from animal studies that provide a new understanding on how MD-PFC circuits support higher-order cognitive function. We argue that the MD may be required for amplifying and sustaining cortical representations under different behavioral conditions. These findings advance a new framework for the broader involvement of distributed thalamo-frontal circuits in cognition and point to the MD as a potential therapeutic target for improving cognitive deficits in schizophrenia and other disorders.

Can we one day prevent mental disorders? Mukherjee et al. (2019) use a genetic mouse model of schizophrenia-risk with established abnormalities in adult hippocampal-prefrontal circuit function and cognitive behaviors to identify circuit-specific treatments during adolescence that prevent the onset of the adult deficits.

Obsessive-compulsive disorder (OCD) is a chronic, disabling condition with inadequate treatment options that leave most patients with substantial residual symptoms. Structural, neurochemical, and behavioral findings point to a significant role for basal ganglia circuits and for the glutamate system in OCD. Genetic linkage and association studies in OCD point to SLC1A1, which encodes the neuronal glutamate/aspartate/cysteine transporter excitatory amino acid transporter 3 (EAAT3)/excitatory amino acid transporter 1 (EAAC1). However, no previous studies have investigated EAAT3 in basal ganglia circuits or in relation to OCD-related behavior. Here, we report a model of Slc1a1 loss based on an excisable STOP cassette that yields successful ablation of EAAT3 expression and function. Using amphetamine as a probe, we found that EAAT3 loss prevents expected increases in (i) locomotor activity, (ii) stereotypy, and (iii) immediate early gene induction in the dorsal striatum following amphetamine administration. Further, Slc1a1-STOP mice showed diminished grooming in an SKF-38393 challenge experiment, a pharmacologic model of OCD-like grooming behavior. This reduced grooming is accompanied by reduced dopamine D1 receptor binding in the dorsal striatum of Slc1a1-STOP mice. Slc1a1-STOP mice also exhibit reduced extracellular dopamine concentrations in the dorsal striatum both at baseline and following amphetamine challenge. Viral-mediated restoration of Slc1a1/EAAT3 expression in the midbrain but not in the striatum results in partial rescue of amphetamine-induced locomotion and stereotypy in Slc1a1-STOP mice, consistent with an impact of EAAT3 loss on presynaptic dopaminergic function. Collectively, these findings indicate that the most consistently associated OCD candidate gene impacts basal ganglia-dependent repetitive behaviors.

Autism spectrum disorder (ASD) represents a major public health burden but translating promising treatment findings from preclinical non-human models of ASD to the clinic has remained challenging. The recent development of forebrain organoids generated from human induced pluripotent stem cells (hiPSCs) derived from subjects with brain disorders is a promising method to study human-specific neurobiology, and may facilitate the development of novel therapeutics.In this study, we utilized forebrain organoids generated from hiPSCs derived from patients from the Old Order Amish community with a rare syndromic form of ASD, carrying a homozygous c.3709DelG mutation in CNTNAP2 and healthy controls to investigate the effects of this mutation on cortical embryonic development.Patients carrying the c.3709DelG mutation in CNTNAP2 present with an increased head circumference and brain MRI reveals an increase in gray matter volume. Patient-derived organoids displayed an increase in total volume that was driven by an increased proliferation in neural progenitor cells, leading to an increase in the generation of cortical neuronal and non-neuronal cell types. The observed phenotypes were rescued after correction of the pathogenic mutation using CRISPR-Cas9. RNA sequencing revealed 339 genes differentially expressed between patient- and control-derived organoids of which a subset are implicated in cell proliferation and neurogenesis. Notably, these differentially expressed genes included previously identified ASD-associated genes and are enriched for genes in ASD-associated weighted gene co-expression networks.This work provides a critical step towards understanding the role of CNTNAP2 in human cortical development and has important mechanistic implications for ASD associated with brain overgrowth. This CNTNAP2 organoid model provides opportunity for further mechanistic inquiry and development of new therapeutic strategies for ASD.

Although long-studied in the central nervous system, there is increasing evidence that dopamine (DA) has important roles in the periphery including in metabolic regulation. Insulin-secreting pancreatic β-cells express the machinery for DA synthesis and catabolism, as well as all five DA receptors. In these cells, DA functions as a negative regulator of glucose-stimulated insulin secretion (GSIS), which is mediated by DA D2-like receptors including D2 (D2R) and D3 (D3R) receptors. However, the fundamental mechanisms of DA synthesis, storage, release, and signaling in pancreatic β-cells and their functional relevance in vivo remain poorly understood. Here, we assessed the roles of the DA precursor L-DOPA in β-cell DA synthesis and release in conjunction with the signaling mechanisms underlying DA's inhibition of GSIS. Our results show that the uptake of L-DOPA is essential for establishing intracellular DA stores in β-cells. Glucose stimulation significantly enhances L-DOPA uptake, leading to increased DA release and GSIS reduction in an autocrine/paracrine manner. Furthermore, D2R and D3R act in combination to mediate dopaminergic inhibition of GSIS. Transgenic knockout mice in which β-cell D2R or D3R expression is eliminated exhibit diminished DA secretion during glucose stimulation, suggesting a new mechanism where D2-like receptors modify DA release to modulate GSIS. Lastly, β-cell-selective D2R knockout mice exhibit marked postprandial hyperinsulinemia in vivo. These results reveal that peripheral D2R and D3R receptors play important roles in metabolism through their inhibitory effects on GSIS. This opens the possibility that blockade of peripheral D2-like receptors by drugs including antipsychotic medications may significantly contribute to the metabolic disturbances observed clinically.

Dopamine neurotransmission is suspected to play important physiological roles in multiple sparsely innervated brain nuclei, but there has not been a means to measure synaptic dopamine release in such regions. The globus pallidus externa (GPe) is a major locus in the basal ganglia that displays a sparse innervation of en passant dopamine axonal fibers. Due to the low levels of innervation that preclude electrochemical analysis, it is unknown if these axons engage in neurotransmission. To address this, we introduce an optical approach using a pH-sensitive fluorescent false neurotransmitter, FFN102, that exhibits increased fluorescence upon exocytosis from the acidic synaptic vesicle to the neutral extracellular milieu. In marked contrast to the striatum, FFN102 transients in the mouse GPe were spatially heterogeneous and smaller than in striatum with the exception of sparse hot spots. GPe transients were also significantly enhanced by high frequency stimulation. Our results support hot spots of dopamine release from substantia nigra axons.

Schizophrenia is a heterogeneous and poorly understood mental disorder that is presently defined solely by its behavioral symptoms. Advances in genetic, epidemiological and brain imaging techniques in the past half century, however, have significantly advanced our understanding of the underlying biology of the disorder. In spite of these advances clinical research remains limited in its power to establish the causal relationships that link etiology with pathophysiology and symptoms. In this context, animal models provide an important tool for causally testing hypotheses about biological processes postulated to be disrupted in the disorder. While animal models can exploit a variety of entry points towards the study of schizophrenia, here we describe an approach that seeks to closely approximate functional alterations observed with brain imaging techniques in patients. By modeling these intermediate pathophysiological alterations in animals, this approach offers an opportunity to (1) tightly link a single functional brain abnormality with its behavioral consequences, and (2) to determine whether a single pathophysiology can causally produce alterations in other brain areas that have been described in patients. In this review we first summarize a selection of well-replicated biological abnormalities described in the schizophrenia literature. We then provide examples of animal models that were studied in the context of patient imaging findings describing enhanced striatal dopamine D2 receptor function, alterations in thalamo-prefrontal circuit function, and metabolic hyperfunction of the hippocampus. Lastly, we discuss the implications of findings from these animal models for our present understanding of schizophrenia, and consider key unanswered questions for future research in animal models and human patients.

Abnormalities in prefrontal gamma aminobutyric acid (GABA)ergic transmission, particularly in fast-spiking interneurons that express parvalbumin (PV), are hypothesized to contribute to the pathophysiology of multiple psychiatric disorders, including schizophrenia, bipolar disorder, anxiety disorders and depression. While primarily histological abnormalities have been observed in patients and in animal models of psychiatric disease, evidence for abnormalities in functional neurotransmission at the level of specific interneuron populations has been lacking in animal models and is difficult to establish in human patients. Using an animal model of a psychiatric disease risk factor, prenatal maternal immune activation (MIA), we found reduced functional GABAergic transmission in the medial prefrontal cortex (mPFC) of adult MIA offspring. Decreased transmission was selective for interneurons expressing PV, resulted from a decrease in release probability and was not observed in calretinin-expressing neurons. This deficit in PV function in MIA offspring was associated with increased anxiety-like behavior and impairments in attentional set shifting, but did not affect working memory. Furthermore, cell-type specific optogenetic inhibition of mPFC PV interneurons was sufficient to impair attentional set shifting and enhance anxiety levels. Finally, we found that in vivo mPFC gamma oscillations, which are supported by PV interneuron function, were linearly correlated with the degree of anxiety displayed in adult mice, and that this correlation was disrupted in MIA offspring. These results demonstrate a selective functional vulnerability of PV interneurons to MIA, leading to affective and cognitive symptoms that have high relevance for schizophrenia and other psychiatric disorders.

Dopaminergic signaling in the striatum, particularly at dopamine 2 receptors (D2R), has been a topic of active investigation in obesity research in the past decades. However, it still remains unclear whether variations in striatal D2Rs modulate the risk for obesity and if so in which direction. Human studies have yielded contradictory findings that likely reflect a complex nonlinear relationship, possibly involving a combination of causal effects and compensatory changes. Animal work indicates that although chronic obesogenic diets reduce striatal D2R function, striatal D2R down-regulation does not lead to obesity. In this study, we evaluated the consequences of striatal D2R up-regulation on body-weight gain susceptibility and energy balance in mice. We used a mouse model of D2R overexpression (D2R-OE) in which D2Rs were selectively up-regulated in striatal medium spiny neurons. We uncover a pathological mechanism by which striatal D2R-OE leads to reduced brown adipose tissue thermogenesis, reduced energy expenditure, and accelerated obesity despite reduced eating. We also show that D2R-OE restricted to development is sufficient to promote obesity and to induce energy-balance deficits. Together, our findings indicate that striatal D2R-OE during development persistently increases the propensity for obesity by reducing energy output in mice. This suggests that early alterations in the striatal dopamine system could represent a key predisposition factor toward obesity.

SUMMARY The dopamine (DA) D2 receptor (D2R) is an important target for the treatment of neuropsychiatric disorders such as schizophrenia and Parkinson’s disease. However, the development of improved therapeutic strategies has been hampered by our incomplete understanding of this receptor’s downstream signaling processes in vivo and how these relate to the desired and undesired effects of drugs. D2R is a G protein-coupled receptor (GPCR) that activates G proteindependent as well as non-canonical arrestin-dependent signaling pathways. Whether these effector pathways act alone or in concert to facilitate specific D2R-dependent behaviors is unclear. Here, we report on the development of a D2R mutant that recruits arrestin but is devoid of G protein activity. When expressed virally in “indirect pathway” medium spiny neurons (iMSNs) in the ventral striatum of D2R knockout mice, this mutant restored basal locomotor activity and cocaine-induced locomotor activity in a manner indistinguishable from wildtype D2R, indicating that arrestin recruitment can drive locomotion in the absence of D2R-mediated G protein signaling. In contrast, incentive motivation was enhanced only by wildtype D2R, signifying a dissociation in the mechanisms that underlie distinct D2R-dependent behaviors, and opening the door to more targeted therapeutics.

Long-range synchronization of neural oscillations correlates with distinct behaviors, yet its causal role remains unproven. In mice, tests of anxiety-like avoidance behavior evoke increases in theta-frequency (~8 Hz) oscillatory synchrony between the ventral hippocampus (vHPC) and medial prefrontal cortex (mPFC). To test the causal role of this synchrony, we dynamically modulated vHPC-mPFC terminal activity using optogenetic stimulation. Oscillatory stimulation at 8 Hz maximally increased avoidance behavior compared to 2, 4 and 20 Hz. Moreover, avoidance behavior was selectively increased when 8 Hz stimulation was delivered in an oscillatory but not pulsatile manner. Furthermore, 8 Hz oscillatory stimulation enhanced vHPC-mPFC neurotransmission and entrained neural activity in the vHPC-mPFC network, resulting in increased synchrony between vHPC theta activity and mPFC spiking. These data suggest a privileged role for vHPC-mPFC theta-frequency communication in generating avoidance behavior, and provide direct evidence that synchronized oscillations play a role in facilitating neural transmission and behavior.

Summary Decreased hippocampal-prefrontal synchrony may mediate cognitive deficits in schizophrenia, but it remains unclear which cells orchestrate this long-range synchrony. Parvalbumin (PV)- and somatostatin (SOM)-expressing interneurons show histological abnormalities in individuals with schizophrenia and are hypothesized to regulate oscillatory synchrony within the prefrontal cortex. To examine the relationship between interneuron function, long-range hippocampal-prefrontal synchrony, and cognition, we optogenetically inhibited SOM and PV neurons in the medial prefrontal cortex (mPFC) of mice performing a spatial working memory task while simultaneously recording neural activity in the mPFC and the hippocampus (HPC). We found that inhibiting SOM, but not PV, interneurons during the encoding phase of the task impaired working memory accuracy. This behavioral impairment was associated with decreased hippocampal-prefrontal synchrony and impaired spatial encoding in mPFC neurons. These findings suggest that interneuron dysfunction may contribute to cognitive deficits associated with schizophrenia by disrupting long-range synchrony between the HPC and PFC.

The validity of rodent models for the study of psychiatric disorders is controversial. Despite great efforts from academic institutions and pharmaceutical companies, as of today, no major therapeutic intervention has been developed for the treatment of psychiatric disorders based on mechanistic insights from rodent models. Here, we argue that despite these historical shortcomings, rodent studies are nevertheless instrumental for identifying neuronal circuit mechanisms underlying behaviours that are affected in psychiatric disorders. Focusing on schizophrenia, we will give four examples of rodent models that were generated based on genetic and environmental risk factors or pathophysiological evidence as entry points. We will then discuss how circuit analysis in these specific examples can be used for testing hypotheses about neuronal mechanisms underlying symptoms of schizophrenia, which will then guide the development of new therapies. This article is part of a discussion meeting issue ‘Of mice and mental health: facilitating dialogue between basic and clinical neuroscientists’.

Current therapies for treating movement disorders such as Parkinson’s disease are effective but limited by undesirable and intractable side effects. Developing more effective therapies will require better understanding of what causes basal ganglia dys-regulation and why medication-induced side effects develop. Although basal ganglia have been extensively studied in the last decades, its circuit anatomy is very complex, and significant controversy exists as to how the interplay of different basal ganglia nuclei process motor information and output. We have recently identified the importance of an underappreciated collateral projection that bridges the striatal output direct pathway with the indirect pathway. These bridging collaterals are extremely plastic in the adult brain and are involved in the regulation of motor balance. Our findings add a new angle to the classical model of basal ganglia circuitry that could be exploited for the development of new therapies against movement disorders. In this Scientific Perspective, we describe the function of bridging collaterals and other recent discoveries that challenge the simplicity of the classical basal ganglia circuit model. We then discuss the potential implication of bridging collaterals in the pathophysiology of Parkinson’s disease and schizophrenia. Because dopamine D2 receptors and striatal neuron excitability have been found to regulate the density of bridging collaterals, we propose that targeting these projections downstream of D2 receptors could be a possible strategy for the treatment of basal ganglia disorders.

The striatum is the major input nucleus of the basal ganglia involved in reward processing, goal-directed behaviors, habit learning, and motor control. The striatum projects to the basal ganglia output nuclei via the "direct" and "indirect" pathways, which can be distinguished by their projection fields and their opposing effects on behavior. In adult animals, the functional opposition is modulated by the differential actions of D1 and D2 dopamine receptors (D1R, D2R), the expression of which is largely separated between these pathways. To determine whether a similar degree of separation exists earlier in development, we used dual-label immunohistochemistry to map dorsal-striatal D1R and D2R expression at the promoter level in postnatal day 0 (PD0) Drd1a-tdTomato/Drd2-GFP BAC transgenic mice, and at the receptor level by costaining for native D1R and D2R in wildtype (WT) PD0 animals. To assess for potential molecular interactions between D1R and D2R we also employed a recently developed proximity-ligation assay (PLA). Limited coexpression and colocalization of the D1R and D2R proteins was found in clusters of neurons endemic to the "patch" compartment as identified by costaining with tyrosine hydroxylase, but not outside these clusters. Moreover, in contrast to our recent findings where we failed to detect a D1R-D2R PLA signal in the adult striatum, in PD0 striatum we did identify a clear PLA signal for this pair of receptors. This colocalization at close proximity points to a possible role for D1R/D2R-mediated crosstalk in early striatal ontogeny.

Brain imaging studies performed in humans have associated low striatal dopamine release and D2R binding with alcohol dependence. Conversely, high striatal D2R binding has been observed in unaffected members of alcoholic families suggesting that high D2R function may protect against alcohol dependence. A possible protective role of increased D2R levels in the striatum is further supported by preclinical studies in non-human primates and rodents. Here, we determined whether there is a causal relationship between D2R levels and alcohol intake. To this end, we upregulated D2R expression levels in the nucleus accumbens of the adult mouse, but selectively restricted the upregulation to the indirect striatal output pathway, which endogenously expresses D2Rs. After overexpression was established, mice were tested in two models of free-choice alcohol drinking: the continuous and intermittent access two-bottle choice models. As anticipated, we found that D2R upregulation leads to hyperactivity in the open field. Contrary to our expectation, D2R upregulation did not reduce alcohol intake during continuous or intermittent access or when alcohol drinking was tested in the context of aversive outcomes. These data argue against a protective role of accumbal indirect pathway D2Rs in alcohol consumption but emphasize their importance in promoting locomotor activity.

Background: Evidence indicating an increase in dopamine D2 receptor (D2R) density and occupancy in patients with schizophrenia comes from positron emission tomography studies using ligands that bind both D2Rs and dopamine D3 receptors (D3Rs), questioning the role of D3Rs in the pathophysiology of the disease. Dopamine D3 receptor positron emission tomography ligands have recently been developed and antagonists with preferential affinity for D3R versus D2R are undergoing clinical evaluation. To determine if an increase in D3Rs in the striatum could produce phenotypes relevant to schizophrenia, we generated a transgenic model of striatal D3R overexpression. Methods: A bi-transgenic system was used to generate mice with increased D3Rs selectively in the striatum. Mice with overexpression of D3R were subjected to an extensive battery of behavioral tests, including several relevant to schizophrenia. Ligand binding and quantitative reverse transcription polymerase chain reaction methods were used to quantify the effect of D3R overexpression on dopamine D1 receptors (D1Rs) in the striatum. Results: Mice with overexpression of D3R show no abnormalities in basic behavioral functions or cognitive tests but do display a deficit in incentive motivation. This was associated with a reduction in striatal D1R ligand binding, driven by a downregulation at the level of transcription. Both motivation and D1R expression were rescued by switching off the transgene in adulthood. Conclusions: Overexpression of D3Rs in the striatum of mice does not elicit cognitive deficits but disrupts motivation, suggesting that changes in D3Rs may be involved in the negative symptoms of schizophrenia. These data imply that it will be important to evaluate the effects of D3R antagonists on motivational symptoms, which are not improved by currently available antipsychotic medications.

Visual Abstract, Alterations in thalamic dopamine (DA) or DA D2 receptors (D2Rs) have been measured in drug addiction and schizophrenia, but the relevance of thalamic D2Rs for behavior is largely unknown. Using in situ hybridization and mice expressing green fluorescent protein (GFP) under the Drd2 promoter, we found that D2R expression within the thalamus is enriched in the paraventricular nucleus (PVT) as well as in more ventral midline thalamic nuclei. Within the PVT, D2Rs are inhibitory as their activation inhibits neuronal action potentials in brain slices. Using Cre-dependent anterograde and retrograde viral tracers, we further determined that PVT neurons are reciprocally interconnected with multiple areas of the limbic system including the amygdala and the nucleus accumbens (NAc). Based on these anatomical findings, we analyzed the role of D2Rs in the PVT in behaviors that are supported by these areas and that also have relevance for schizophrenia and drug addiction. Male and female mice with selective overexpression of D2Rs in the PVT showed attenuated cocaine locomotor sensitization, whereas anxiety levels, fear conditioning, sensorimotor gating, and food-motivated behaviors were not affected. These findings suggest the importance of PVT inhibition by D2Rs in modulating the sensitivity to cocaine, a finding that may have novel implications for human drug use.