Indirect Pathway Neurons Research Articles

Haploinsufficiency of Syngap1 in striatal indirect pathway neurons alters motor and goal-directed behaviors in mice.

SYNGAP1 is a high-confidence autism spectrum disorder (ASD) risk gene and mutations in SYNGAP1 lead to a neurodevelopmental disorder (NDD) that presents with epilepsy, ASD, motor developmental delay, and intellectual disability. SYNGAP1 codes for Ras/Rap GTP-ase activating protein SynGAP (SynGAP). In mice, SynGAP is located in the postsynaptic density of glutamatergic synapses and regulates glutamate receptor trafficking in an activity-dependent manner. In addition to forebrain glutamatergic neurons, Syngap1 is highly expressed in the striatum, although the functions of SynGAP in the striatum have not been extensively studied. Here we show that Syngap1 is expressed in both direct and indirect pathway striatal projection neurons (dSPNs and iSPNs) in mice of both sexes. In a mouse model of Syngap1 haploinsufficiency, dendritic spine density, morphology, and intrinsic excitability are altered primarily in iSPNs, but not dSPNs. At the behavioral level, SynGAP reduction alters striatal-dependent motor learning and goal-directed behavior. Several behavioral phenotypes are reproduced by iSPN-specific Syngap1 reduction and, in turn, prevented by iSPN-specific Syngap1 rescue. These results establish the importance of SynGAP to striatal neuron function and pinpoint the indirect pathway as a key circuit in the neurobiology of SYNGAP1-related NDD.Significance statement SYNGAP1 mutations cause a neurodevelopmental disorder presenting with intellectual disability, motor problems, epilepsy, autism spectrum disorder, and a constellation of other behavioral and psychiatric conditions. SynGAP protein is highly expressed in the striatum but its functions in this brain region have not yet been explored. This study shows that loss of one copy of the Syngap1 gene from striatal indirect, but not direct, pathway neurons alters synaptic properties, cellular excitability, motor behaviors, and goal-directed responding in mice. This work provides a new perspective on the functions of SynGAP and suggests that altered activity in striatal circuits may be an important driver of the motor and learning alterations in people with SYNGAP1 disorder.

LRRK2 mediates haloperidol-induced changes in indirect pathway striatal projection neurons.

Haloperidol is used to manage psychotic symptoms in several neurological disorders through mechanisms that involve antagonism of dopamine D2 receptors that are highly expressed in the striatum. Significant side effects of haloperidol, known as extrapyramidal symptoms, lead to motor deficits similar to those seen in Parkinson's disease and present a major challenge in clinical settings. The underlying molecular mechanisms responsible for these side effects remain poorly understood. Parkinson's disease-associated LRRK2 kinase has an important role in striatal physiology and a known link to dopamine D2 receptor signaling. Here, we systematically explore convergent signaling of haloperidol and LRRK2 through pharmacological or genetic inhibition of LRRK2 kinase, as well as knock-in mouse models expressing pathogenic mutant LRRK2 with increased kinase activity. Behavioral assays show that LRRK2 kinase inhibition ameliorates haloperidol-induced motor changes in mice. A combination of electrophysiological and anatomical approaches reveals that LRRK2 kinase inhibition interferes with haloperidol-induced changes, specifically in striatal neurons of the indirect pathway. Proteomic studies and targeted intracellular pathway analyses demonstrate that haloperidol induces a similar pattern of intracellular signaling as increased LRRK2 kinase activity. Our study suggests that LRRK2 kinase plays a key role in striatal dopamine D2 receptor signaling underlying the undesirable motor side effects of haloperidol. This work opens up new therapeutic avenues for dopamine-related disorders, such as psychosis, also furthering our understanding of Parkinson's disease pathophysiology.

Optochemical control of slow-wave sleep in the nucleus accumbens of male mice by a photoactivatable allosteric modulator of adenosine A2A receptors.

Optochemistry, an emerging pharmacologic approach in which light is used to selectively activate or deactivate molecules, has the potential to alleviate symptoms, cure diseases, and improve quality of life while preventing uncontrolled drug effects. The development of in-vivo applications for optochemistry to render brain cells photoresponsive without relying on genetic engineering has been progressing slowly. The nucleus accumbens (NAc) is a region for the regulation of slow-wave sleep (SWS) through the integration of motivational stimuli. Adenosine emerges as a promising candidate molecule for activating indirect pathway neurons of the NAc expressing adenosine A2A receptors (A2ARs) to induce SWS. Here, we developed a brain-permeable positive allosteric modulator of A2ARs (A2AR PAM) that can be rapidly photoactivated with visible light (λ > 400 nm) and used it optoallosterically to induce SWS in the NAc of freely behaving male mice by increasing the activity of extracellular adenosine derived from astrocytic and neuronal activity.

Dopamine depletion weakens direct pathway modulation of SNr neurons

Neurons in the substantia nigra reticulata (SNr) transmit information about basal ganglia output to dozens of brain regions in thalamocortical and brainstem motor networks. Activity of SNr neurons is regulated by convergent input from upstream basal ganglia nuclei, including GABAergic inputs from the striatum and the external globus pallidus (GPe). GABAergic inputs from the striatum convey information from the direct pathway, while GABAergic inputs from the GPe convey information from the indirect pathway. Chronic loss of dopamine, as occurs in Parkinson's disease, disrupts the balance of direct and indirect pathway neurons at the level of the striatum, but the question of how dopamine loss affects information propagation along these pathways outside of the striatum is less well understood. Using a combination of in vivo and slice electrophysiology, we find that dopamine depletion selectively weakens the direct pathway's influence over neural activity in the SNr due to changes in the decay kinetics of GABA-mediated synaptic currents. GABAergic signaling from GPe neurons in the indirect pathway was not affected, resulting in an inversion of the normal balance of inhibitory control over basal ganglia output through the SNr. These results highlight the contribution of cellular mechanisms outside of the striatum that impact the responses of basal ganglia output neurons to the direct and indirect pathways in disease.

Comparison of unitary synaptic currents generated by indirect and direct pathway neurons of the mouse striatum.

Two subtypes of striatal spiny projection neurons, iSPNs and dSPNs, whose axons form the "indirect" and "direct" pathways of the basal ganglia, respectively, both make synaptic connections in the external globus pallidus (GPe) but are usually found to have different effects on behavior. Activation of the terminal fields of iSPNs or dSPNs generated compound currents in almost all GPe neurons. To determine whether iSPNs and dSPNs have the same or different effects on pallidal neurons, we studied the unitary synaptic currents generated in GPe neurons by action potentials in single striatal neurons. We used optogenetic excitation to elicit repetitive firing in a small number of nearby SPNs, producing sparse barrages of inhibitory postsynaptic currents (IPSCs) in GPe neurons. From these barrages, we isolated sequences of IPSCs with similar time courses and amplitudes, which presumably arose from the same SPN. There was no difference between the amplitudes of unitary IPSCs generated by the indirect and direct pathways. Most unitary IPSCs were small, but a subset from each pathway were much larger. To determine the effects of these unitary synaptic currents on the action potential firing of GPe neurons, we drove SPNs to fire as before and recorded the membrane potential of GPe neurons. Large unitary potentials from iSPNs and dSPNs perturbed the spike timing of GPe neurons in a similar way. Most SPN-GPe neuron pairs are weakly connected, but a subset of pairs in both pathways are strongly connected.NEW & NOTEWORTHY This is the first study to record the synaptic currents generated by single identified direct or indirect pathway striatal neurons on single pallidal neurons. Each GPe neuron receives synaptic inputs from both pathways. Most striatal neurons generate small synaptic currents that become influential when occurring together, but a few are powerful enough to be individually influential.

Enhancing reinforcement learning models by including direct and indirect pathways improves performance on striatal dependent tasks.

A major advance in understanding learning behavior stems from experiments showing that reward learning requires dopamine inputs to striatal neurons and arises from synaptic plasticity of cortico-striatal synapses. Numerous reinforcement learning models mimic this dopamine-dependent synaptic plasticity by using the reward prediction error, which resembles dopamine neuron firing, to learn the best action in response to a set of cues. Though these models can explain many facets of behavior, reproducing some types of goal-directed behavior, such as renewal and reversal, require additional model components. Here we present a reinforcement learning model, TD2Q, which better corresponds to the basal ganglia with two Q matrices, one representing direct pathway neurons (G) and another representing indirect pathway neurons (N). Unlike previous two-Q architectures, a novel and critical aspect of TD2Q is to update the G and N matrices utilizing the temporal difference reward prediction error. A best action is selected for N and G using a softmax with a reward-dependent adaptive exploration parameter, and then differences are resolved using a second selection step applied to the two action probabilities. The model is tested on a range of multi-step tasks including extinction, renewal, discrimination; switching reward probability learning; and sequence learning. Simulations show that TD2Q produces behaviors similar to rodents in choice and sequence learning tasks, and that use of the temporal difference reward prediction error is required to learn multi-step tasks. Blocking the update rule on the N matrix blocks discrimination learning, as observed experimentally. Performance in the sequence learning task is dramatically improved with two matrices. These results suggest that including additional aspects of basal ganglia physiology can improve the performance of reinforcement learning models, better reproduce animal behaviors, and provide insight as to the role of direct- and indirect-pathway striatal neurons.

The respective activation and silencing of striatal direct and indirect pathway neurons support behavior encoding

The basal ganglia are known to control actions and modulate movements. Neuronal activity in the two efferent pathways of the dorsal striatum is critical for appropriate behavioral control. Previous evidence has led to divergent conclusions on the respective engagement of both pathways during actions. Using calcium imaging to evaluate how neurons in the direct and indirect pathways encode behaviors during self-paced spontaneous explorations in an open field, we observed that the two striatal pathways exhibit distinct tuning properties. Supervised learning algorithms revealed that direct pathway neurons encode behaviors through their activation, whereas indirect pathway neurons exhibit behavior-specific silencing. These properties remain stable for weeks. Our findings highlight a complementary encoding of behaviors with congruent activations in the direct pathway encoding multiple accessible behaviors in a given context, and in the indirect pathway encoding the suppression of competing behaviors. This model reconciles previous conflicting conclusions on motor encoding in the striatum.

The Secondary Motor Cortex-striatum Circuit Contributes to Suppressing Inappropriate Responses in Perceptual Decision Behavior

The secondary motor cortex (M2) encodes choice-related information and plays an important role in cue-guided actions. M2 neurons innervate the dorsal striatum (DS), which also contributes to decision-making behavior, yet how M2 modulates signals in the DS to influence perceptual decision-making is unclear. Using mice performing a visual Go/No-Go task, we showed that inactivating M2 projections to the DS impaired performance by increasing the false alarm (FA) rate to the reward-irrelevant No-Go stimulus. The choice signal of M2 neurons correlated with behavioral performance, and the inactivation of M2 neurons projecting to the DS reduced the choice signal in the DS. By measuring and manipulating the responses of direct or indirect pathway striatal neurons defined by M2 inputs, we found that the indirect pathway neurons exhibited a shorter response latency to the No-Go stimulus, and inactivating their early responses increased the FA rate. These results demonstrate that the M2-to-DS pathway is crucial for suppressing inappropriate responses in perceptual decision behavior.

A striatal circuit balances learned fear in the presence and absence of sensory cues.

During fear learning, defensive behaviors like freezing need to be finely balanced in the presence or absence of threat-predicting cues (conditioned stimulus, CS). Nevertheless, the circuits underlying such balancing are largely unknown. Here, we investigate the role of the ventral tail striatum (vTS) in auditory-cued fear learning of male mice. In vivo Ca2+ imaging showed that sizable sub-populations of direct (D1R+) and indirect pathway neurons (Adora+) in the vTS responded to footshocks, and to the initiation of movements after freezing; moreover, a sub-population of D1R+ neurons increased its responsiveness to an auditory CS during fear learning. In-vivo optogenetic silencing shows that footshock-driven activity of D1R+ neurons contributes to fear memory formation, whereas Adora+ neurons modulate freezing in the absence of a learned CS. Circuit tracing identified the posterior insular cortex (pInsCx) as an important cortical input to the vTS, and recording of optogenetically evoked EPSCs revealed long-term plasticity with opposite outcomes at the pInsCx synapses onto D1R+ - and Adora+ neurons. Thus, direct- and indirect pathways neurons of the vTS show differential signs of plasticity after fear learning, and balance defensive behaviors in the presence and absence of learned sensory cues.

Early TNF-Dependent Regulation of Excitatory and Inhibitory Synapses on Striatal Direct Pathway Medium Spiny Neurons in the YAC128 Mouse Model of Huntington's Disease.

Huntington's disease (HD) is a neurodegenerative disease caused by a polyglutamine expansion in the huntingtin gene. Neurodegeneration first occurs in the striatum, accompanied by an elevation in inflammatory cytokines. Using the presymptomatic male YAC128 HD model mouse, we examined the synaptic input onto the striatal medium spiny neurons to look for early changes that precede degeneration. We observed an increase in excitatory synaptic strength, as measured by AMPA/NMDA ratios, specifically on direct pathway D1 receptor expressing medium spiny neurons, with no changes on indirect pathway neurons. The changes in excitation were accompanied by a decrease in inhibitory synaptic strength, as measured by the amplitude of miniature inhibitory synaptic currents. The pro-inflammatory cytokine tumor necrosis factor alpha (TNF) was elevated in the striatum of YAC128 at the ages examined. Critically, the changes in excitatory and inhibitory inputs are both dependent on TNF signaling, as blocking TNF signaling genetically or pharmacological normalized synaptic strength. The observed changes in synaptic function are similar to the changes seen in D1 medium spiny neurons treated with high levels of TNF, suggesting that saturating levels of TNF exist in the striatum even at early stages of HD. The increase in glutamatergic synaptic strength and decrease in inhibitory synaptic strength would increase direct pathway neuronal excitability, which may potentiate excitotoxicity during the progress of HD.SIGNIFICANCE STATEMENT The striatum is the first structure to degenerate in Huntington's disease, but the early changes that presage the degeneration are not well defined. Here we identify early synaptic changes in the YAC128 mouse model of Huntington's disease specifically on a subpopulation of striatal neurons. These neurons have stronger excitatory synapses and weaker inhibitory inputs, and thus would increase the susceptibility to excitotoxicity. These changes are dependent on signaling by the pro-inflammatory cytokine TNFα. TNF is elevated even at early presymptomatic stages, and blocking TNF signaling even acutely will reverse the synaptic changes. This suggests early intervention could be important therapeutically.

Acute cannabinoids impair association learning via selectively enhancing synaptic transmission in striatonigral neurons

BackgroundCannabinoids and their derivatives attract strong interest due to the tremendous potential of their psychoactive effects for treating psychiatric disorders and symptoms. However, their clinical application is restricted by various side-effects such as impaired coordination, anxiety, and learning and memory disability. Adverse impact on dorsal striatum-dependent learning is an important side-effect of cannabinoids. As one of the most important forms of learning mediated by the dorsal striatum, reinforcement learning is characterized by an initial association learning phase, followed by habit learning. While the effects of cannabinoids on habit learning have been well-studied, little is known about how cannabinoids influence the initial phase of reinforcement learning.ResultsWe found that acute activation of cannabinoid receptor type 1 (CB1R) by the synthetic cannabinoid HU210 induced dose-dependent impairment of association learning, which could be alleviated by intra-dorsomedial striatum (DMS) injection of CB1R antagonist. Moreover, acute exposure to HU210 elicited enhanced synaptic transmission in striatonigral “direct” pathway medium spiny neurons (MSNs) but not indirect pathway neurons in DMS. Intriguingly, enhancement of synaptic transmission that is also observed after learning was abolished by HU210, indicating cannabinoid system might disrupt reinforcement learning by confounding synaptic plasticity normally required for learning. Remarkably, the impaired response-reinforcer learning was also induced by selectively enhancing the D1-MSN (MSN that selectively expresses the dopamine receptor type 1) activity by virally expressing excitatory hM3Dq DREADD (designer receptor exclusively activated by a designer drug), which could be rescued by specifically silencing the D1-MSN activity via hM4Di DREADD.ConclusionOur findings demonstrate dose-dependent deleterious effects of cannabinoids on association learning by disrupting plasticity change required for learning associated with the striatal direct pathway, which furthers our understanding of the side-effects of cannabinoids and the underlying mechanisms.

Non-Apoptotic Caspase-3 Activation Mediates Early Synaptic Dysfunction of Indirect Pathway Neurons in the Parkinsonian Striatum.

Non-apoptotic caspase-3 activation is critically involved in dendritic spine loss and synaptic dysfunction in Alzheimer’s disease. It is, however, not known whether caspase-3 plays similar roles in other pathologies. Using a mouse model of clinically manifest Parkinson’s disease, we provide the first evidence that caspase-3 is transiently activated in the striatum shortly after the degeneration of nigrostriatal dopaminergic projections. This caspase-3 activation concurs with a rapid loss of dendritic spines and deficits in synaptic long-term depression (LTD) in striatal projection neurons forming the indirect pathway. Interestingly, systemic treatment with a caspase inhibitor prevents both the spine pruning and the deficit of indirect pathway LTD without interfering with the ongoing dopaminergic degeneration. Taken together, our data identify transient and non-apoptotic caspase activation as a critical event in the early plastic changes of indirect pathway neurons following dopamine denervation.

Striatal Indirect Pathway Dysfunction Underlies Motor Deficits in a Mouse Model of Paroxysmal Dyskinesia.

Abnormal involuntary movements, or dyskinesias, are seen in many neurologic diseases, including disorders where the brain appears grossly normal. This observation suggests that alterations in neural activity or connectivity may underlie dyskinesias. One influential model proposes that involuntary movements are driven by an imbalance in the activity of striatal direct and indirect pathway neurons (dMSNs and iMSNs, respectively). Indeed, in some animal models, there is evidence that dMSN hyperactivity contributes to dyskinesia. Given the many diseases associated with dyskinesia, it is unclear whether these findings generalize to all forms. Here, we used male and female mice in a mouse model of paroxysmal nonkinesigenic dyskinesia (PNKD) to assess whether involuntary movements are related to aberrant activity in the striatal direct and indirect pathways. In this model, as in the human disorder PNKD, animals experience dyskinetic attacks in response to caffeine or alcohol. Using optically identified striatal single-unit recordings in freely moving PNKD mice, we found a loss of iMSN firing during dyskinesia bouts. Further, chemogenetic inhibition of iMSNs triggered dyskinetic episodes in PNKD mice. Finally, we found that these decreases in iMSN firing are likely because of aberrant endocannabinoid-mediated suppression of glutamatergic inputs. These data show that striatal iMSN dysfunction contributes to the etiology of dyskinesia in PNKD, and suggest that indirect pathway hypoactivity may be a key mechanism for the generation of involuntary movements in other disorders.SIGNIFICANCE STATEMENT Involuntary movements, or dyskinesias, are part of many inherited and acquired neurologic syndromes. There are few effective treatments, most of which have significant side effects. Better understanding of which cells and patterns of activity cause dyskinetic movements might inform the development of new neuromodulatory treatments. In this study, we used a mouse model of an inherited human form of paroxysmal dyskinesia in combination with cell type-specific tools to monitor and manipulate striatal activity. We were able to narrow in on a specific group of neurons that causes dyskinesia in this model, and found alterations in a well-known form of plasticity in this cell type, endocannabinoid-dependent synaptic LTD. These findings point to new areas for therapeutic development.

Dysregulation of the Basal Ganglia Indirect Pathway in Early Symptomatic Q175 Huntington's Disease Mice.

The debilitating psychomotor symptoms of Huntington's disease (HD) are linked partly to degeneration of the basal ganglia indirect pathway. At early symptomatic stages, before major cell loss, indirect pathway neurons exhibit numerous cellular and synaptic changes in HD and its models. However, the impact of these alterations on circuit activity remains poorly understood. To address this gap, optogenetic- and reporter-guided electrophysiological interrogation was used in early symptomatic male and female Q175 HD mice. D2 dopamine receptor-expressing striatal projection neurons (D2-SPNs) were hypoactive during synchronous cortical slow-wave activity, consistent with known reductions in dendritic excitability and cortical input strength. Downstream prototypic parvalbumin-expressing external globus pallidus (PV+ GPe) neurons discharged at 2-3 times their normal rate, even during periods of D2-SPN inactivity, arguing that defective striatopallidal inhibition was not the only cause of their hyperactivity. Indeed, PV+ GPe neurons also exhibited abnormally elevated autonomous firing ex vivo Optogenetic inhibition of PV+ GPe neurons in vivo partially and fully ameliorated the abnormal hypoactivity of postsynaptic subthalamic nucleus (STN) and putative PV- GPe neurons, respectively. In contrast to STN neurons whose autonomous firing is impaired in HD mice, putative PV- GPe neuron activity was unaffected ex vivo, implying that excessive inhibition was responsible for their hypoactivity in vivo Together with previous studies, these data demonstrate that (1) indirect pathway nuclei are dysregulated in Q175 mice through changes in presynaptic activity and/or intrinsic cellular and synaptic properties; and (2) prototypic PV+ GPe neuron hyperactivity and excessive target inhibition are prominent features of early HD pathophysiology.SIGNIFICANCE STATEMENT The early symptoms of Huntington's disease (HD) are linked to degenerative changes in the action-suppressing indirect pathway of the basal ganglia. Consistent with this linkage, the intrinsic properties of cells in this pathway exhibit complex alterations in HD and its models. However, the impact of these changes on activity is poorly understood. Using electrophysiological and optogenetic approaches, we demonstrate that the indirect pathway is highly dysregulated in early symptomatic HD mice through changes in upstream activity and/or intrinsic properties. Furthermore, we reveal that hyperactivity of external globus pallidus neurons and excessive inhibition of their targets are key features of early HD pathophysiology. Together, these findings could help to inform the development and targeting of viral-based, gene therapeutic approaches for HD.

Direct and indirect pathway neurons in ventrolateral striatum differentially regulate licking movement and nigral responses.

Drinking behavior in rodents is characterized by stereotyped, rhythmic licking movement, which is regulated by the basal ganglia. It is unclear how direct and indirect pathways control the lick bout and individual spout contact. We find that inactivating D1 and D2 receptor-expressing medium spiny neurons (MSNs) in the ventrolateral striatum (VLS) oppositely alters the number of licks in a bout. D1- and D2-MSNs exhibit different patterns of lick-sequence-related activity and different phases of oscillation time-locked to the lick cycle. On the timescale of a lick cycle, transient inactivation of D1-MSNs during tongue protrusion reduces spout contact probability, whereas transiently inactivating D2-MSNs has no effect. On the timescale of a lick bout, inactivation of D1-MSNs (D2-MSNs) causes rate increase (decrease) in a subset of basal ganglia output neurons that decrease firing during licking. Our results reveal the distinct roles of D1- and D2-MSNs in regulating licking at both coarse and fine timescales.

Optogenetic inhibition of indirect pathway neurons in the dorsomedial striatum reduces excessive grooming in Sapap3-knockout mice.

Excessive grooming of Sapap3-KO mice has been used as a model of obsessive-compulsive disorder (OCD). Previous studies suggest that dysregulation of cortico-striatal circuits is critically important in the generation of compulsive behaviors, and it has been proposed that the alteration in the activity patterns of striatal circuitry underlies the excessive grooming observed in Sapap3-KO mice. To test this hypothesis, we used in-vivo calcium imaging of individual cells to record striatal activity in these animals and optogenetic inhibition to manipulate this activity. We identified striatal neurons that are modulated during grooming behavior and found that their proportion is significantly larger in Sapap3-KO mice compared to wild-type littermates. Inhibition of striatal cells in Sapap3-KO mice increased the number of grooming episodes observed. Remarkably, the specific inhibition of indirect pathway neurons decreased the occurrence of grooming events. Our results indicate that there is striatal neural activity related to excessive grooming engagement in Sapap3-KO mice. We also demonstrate, for the first time, that specific inhibition of striatal indirect pathway neurons reduces this compulsive phenotype, suggesting that treatments that alleviate compulsive symptoms in OCD patients may exert their effects through this specific striatal population.

Striatal TRPV1 activation by acetaminophen ameliorates dopamine D2 receptor antagonist-induced orofacial dyskinesia.

Antipsychotics often cause tardive dyskinesia, an adverse symptom of involuntary hyperkinetic movements. Analysis of the US Food and Drug Administration Adverse Event Reporting System and JMDC insurance claims revealed that acetaminophen prevented the dyskinesia induced by dopamine D2 receptor antagonists. In vivo experiments further showed that a 21-day treatment with haloperidol increased the number of vacuous chewing movements (VCMs) in rats, an effect that was inhibited by oral acetaminophen treatment or intracerebroventricular injection of N-(4-hydroxyphenyl)-arachidonylamide (AM404), an acetaminophen metabolite that acts as an activator of the transient receptor potential vanilloid 1 (TRPV1). In mice, haloperidol-induced VCMs were also mitigated by treatment with AM404 applied to the dorsal striatum, an effect not seen in TRPV1-deficient mice. Acetaminophen prevented the haloperidol-induced decrease in the number of c-Fos+preproenkephalin+ striatal neurons in wild-type mice but not in TRPV1-deficient mice. Finally, chemogenetic stimulation of indirect pathway medium spiny neurons in the dorsal striatum decreased haloperidol-induced VCMs. These results suggest that acetaminophen activates the indirect pathway neurons by activating TRPV1 channels via AM404.

Development of CVN424: A Selective and Novel GPR6 Inverse Agonist Effective in Models of Parkinson Disease.

GPR6 is an orphan G-protein-coupled receptor that has enriched expression in the striatopallidal, indirect pathway and medium spiny neurons of the striatum. This pathway is greatly impacted by the loss of the nigro-striatal dopaminergic neurons in Parkinson disease, and modulating this neurocircuitry can be therapeutically beneficial. In this study, we describe the in vitro and in vivo pharmacological characterization of (R)-1-(2-(4-(2,4-difluorophenoxy)piperidin-1-yl)-3-((tetrahydrofuran-3-yl)amino)-7,8-dihydropyrido[3,4-b]pyrazin-6(5H)-yl)ethan-1-one (CVN424), a highly potent and selective small-molecule inverse agonist for GPR6 that is currently undergoing clinical evaluation. CVN424 is brain-penetrant and shows dose-dependent receptor occupancy that attained brain 50% of receptor occupancy at plasma concentrations of 6.0 and 7.4 ng/ml in mice and rats, respectively. Oral administration of CVN424 dose-dependently increases locomotor activity and reverses haloperidol-induced catalepsy. Furthermore, CVN424 restored mobility in bilateral 6-hydroxydopamine lesion model of Parkinson disease. The presence and localization of GPR6 in medium spiny neurons of striatum postmortem samples from both nondemented control and patients with Parkinson disease were confirmed at the level of both RNA (using Nuclear Enriched Transcript Sort sequencing) and protein. This body of work demonstrates that CVN424 is a potent, orally active, and brain-penetrant GPR6 inverse agonist that is effective in preclinical models and is a potential therapeutic for improving motor function in patients with Parkinson disease. SIGNIFICANCE STATEMENT: CVN424 represents a nondopaminergic novel drug for potential use in patients with Parkinson disease.

Activation of the basal ganglia and indirect pathway neurons during frontal lobe seizures

There are no detailed descriptions of neuronal circuit active during frontal lobe motor seizures. Using activity reporter mice, local field potential recordings, tissue clearing, viral tracing, and super-resolution microscopy, we found neuronal activation after focal motor to bilateral tonic-clonic seizures in the striatum, globus pallidus externus, subthalamic nucleus, substantia nigra pars reticulata and neurons of the indirect pathway. Seizures preferentially activated dopamine D2 receptor-expressing neurons over D1 in the striatum, which have different projections. Furthermore, the D2 receptor agonist infused into the striatum exerted an anticonvulsant effect. Seizures activate structures via short and long latency loops, and anatomical connections of the seizure focus determine the seizure circuit. These studies, for the first time, show activation of neurons in the striatum, globus pallidus, subthalamic nucleus, and substantia nigra during frontal lobe motor seizures on the cellular level, revealing a complex neuronal activation circuit subject to modulation by the basal ganglia.

Striatal direct and indirect pathway neurons differentially control the encoding and updating of goal-directed learning.

The posterior dorsomedial striatum (pDMS) is necessary for goal-directed action; however, the role of the direct (dSPN) and indirect (iSPN) spiny projection neurons in the pDMS in such actions remains unclear. In this series of experiments, we examined the role of pDMS SPNs in goal-directed action in rats and found that whereas dSPNs were critical for goal-directed learning and for energizing the learned response, iSPNs were involved in updating that learning to support response flexibility. Instrumental training elevated expression of the plasticity marker Zif268 in dSPNs only, and chemogenetic suppression of dSPN activity during training prevented goal-directed learning. Unilateral optogenetic inhibition of dSPNs induced an ipsilateral response bias in goal-directed action performance. In contrast, although initial goal-directed learning was unaffected by iSPN manipulations, optogenetic inhibition of iSPNs, but not dSPNs, impaired the updating of this learning and attenuated response flexibility after changes in the action-outcome contingency.