5-HT1B receptors mediate dopaminergic inhibition of vesicular fusion and GABA release from striatonigral synapses.

The substantia nigra pars reticulata (SNr), a crucial basal ganglia output nucleus, contains a dense expression of dopamine D1 receptors (D1Rs), along with dendrites belonging to dopaminergic neurons of substantia nigra pars compacta. These D1Rs are primarily located on the terminals of striatonigral medium spiny neurons, suggesting their involvement in the regulation of neurotransmitter release from the direct pathway in response to somatodendritic dopamine release. To explore the hypothesis that D1Rs modulate GABA release from striatonigral synapses, we conducted optical recordings of striatonigral activity and postsynaptic patch-clamp recordings from SNr neurons in the presence of dopamine and D1R agonists. We found that dopamine inhibits optogenetically triggered striatonigral GABA release by modulating vesicle fusion and Ca 2+ influx in striatonigral boutons. Notably, the effect of DA was independent of D1R activity but required activation of 5-HT1B receptors. Our results suggest a serotonergic mechanism involved in the therapeutic actions of dopaminergic medications for Parkinson's disease and psychostimulant-related disorders.

Dysregulated acetylcholine-mediated dopamine neurotransmission in the eIF4E Tg mouse model of autism spectrum disorders.

Autism Spectrum Disorders (ASD) consist of diverse neurodevelopmental conditions where core behavioral symptoms are critical for diagnosis. Altered dopamine neurotransmission in the striatum has been suggested to contribute to the behavioral features of ASD. Here, we examine dopamine neurotransmission in a mouse model of ASD characterized by elevated expression of the eukaryotic initiation factor 4E (eIF4E), a key regulator of cap-dependent translation, using a comprehensive approach that encompasses genetics, behavior, synaptic physiology, and imaging. The results indicate that increased eIF4E expression leads to behavioral inflexibility and impaired striatal dopamine release. The loss of normal dopamine neurotransmission is due to a defective nicotinic receptor signaling that regulates calcium dynamics in dopaminergic axons. These findings reveal an intricate interplay between eIF4E, DA neurotransmission, and behavioral flexibility, provide a mechanistic understanding of ASD symptoms and offer a foundation for targeted therapeutic interventions.

Deficiency of the Heterogeneous Nuclear Ribonucleoprotein U locus leads to delayed hindbrain neurogenesis.

Genetic variants affecting Heterogeneous Nuclear Ribonucleoprotein U (HNRNPU) have been identified in several neurodevelopmental disorders (NDDs). HNRNPU is widely expressed in the human brain and shows the highest postnatal expression in the cerebellum. Recent studies have investigated the role of HNRNPU in cerebral cortical development, but the effects of HNRNPU deficiency on cerebellar development remain unknown. Here, we describe the molecular and cellular outcomes of HNRNPU locus deficiency during in vitro neural differentiation of patient-derived and isogenic neuroepithelial stem cells with a hindbrain profile. We demonstrate that HNRNPU deficiency leads to chromatin remodeling of A/B compartments, and transcriptional rewiring, partly by impacting exon inclusion during mRNA processing. Genomic regions affected by the chromatin restructuring and host genes of exon usage differences show a strong enrichment for genes implicated in epilepsies, intellectual disability, and autism. Lastly, we show that at the cellular level HNRNPU downregulation leads to an increased fraction of neural progenitors in the maturing neuronal population. We conclude that the HNRNPU locus is involved in delayed commitment of neural progenitors to differentiate in cell types with hindbrain profile.

Involvement of Autophagy in Levodopa-Induced Dyskinesia.

BACKGROUND: Autophagy is intensively studied in cancer, metabolic and neurodegenerative diseases, but little is known about its role in pathological conditions linked to altered neurotransmission. We examined the involvement of autophagy in levodopa (l-dopa)-induced dyskinesia, a frequent motor complication developed in response to standard dopamine replacement therapy in parkinsonian patients. METHODS: We used mouse and non-human primate models of Parkinson's disease to examine changes in autophagy associated with chronic l-dopa administration and to establish a causative link between impaired autophagy and dyskinesia. RESULTS: We found that l-dopa-induced dyskinesia is associated with accumulation of the autophagy-specific substrate p62, a marker of autophagy deficiency. Increased p62 was observed in a subset of projection neurons located in the striatum and depended on l-dopa-mediated activation of dopamine D1 receptors, and mammalian target of rapamycin. Inhibition of mammalian target of rapamycin complex 1 with rapamycin counteracted the impairment of autophagy produced by l-dopa, and reduced dyskinesia. The anti-dyskinetic effect of rapamycin was lost when autophagy was constitutively suppressed in D1 receptor-expressing striatal neurons, through inactivation of the autophagy-related gene protein 7. CONCLUSIONS: These findings indicate that augmented responsiveness at D1 receptors leads to dysregulated autophagy, and results in the emergence of l-dopa-induced dyskinesia. They further suggest the enhancement of autophagy as a therapeutic strategy against dyskinesia. © 2021 The Authors. Movement Disorders published by Wiley Periodicals LLC on behalf of International Parkinson and Movement Disorder Society.

mTOR Suppresses Macroautophagy During Striatal Postnatal Development and Is Hyperactive in Mouse Models of Autism Spectrum Disorders.

Macroautophagy (hereafter referred to as autophagy) plays a critical role in neuronal function related to development and degeneration. Here, we investigated whether autophagy is developmentally regulated in the striatum, a brain region implicated in neurodevelopmental disease. We demonstrate that autophagic flux is suppressed during striatal postnatal development, reaching adult levels around postnatal day 28 (P28). We also find that mTOR signaling, a key regulator of autophagy, increases during the same developmental period. We further show that mTOR signaling is responsible for suppressing autophagy, via regulation of Beclin-1 and VPS34 activity. Finally, we discover that autophagy is downregulated during late striatal postnatal development (P28) in mice with in utero exposure to valproic acid (VPA), an established mouse model of autism spectrum disorder (ASD). VPA-exposed mice also display deficits in striatal neurotransmission and social behavior. Correction of hyperactive mTOR signaling in VPA-exposed mice restores social behavior. These results demonstrate that neurons coopt metabolic signaling cascades to developmentally regulate autophagy and provide additional evidence that mTOR-dependent signaling pathways represent pathogenic signaling cascades in ASD mouse models that are active during specific postnatal windows.

Cell-type-specific regulation of neuronal intrinsic excitability by macroautophagy.

The basal ganglia are a group of subcortical nuclei that contribute to action selection and reinforcement learning. The principal neurons of the striatum, spiny projection neurons of the direct (dSPN) and indirect (iSPN) pathways, maintain low intrinsic excitability, requiring convergent excitatory inputs to fire. Here, we examined the role of autophagy in mouse SPN physiology and animal behavior by generating conditional knockouts of Atg7 in either dSPNs or iSPNs. Loss of autophagy in either SPN population led to changes in motor learning but distinct effects on cellular physiology. dSPNs, but not iSPNs, required autophagy for normal dendritic structure and synaptic input. In contrast, iSPNs, but not dSPNs, were intrinsically hyperexcitable due to reduced function of the inwardly rectifying potassium channel, Kir2. These findings define a novel mechanism by which autophagy regulates neuronal activity: control of intrinsic excitability via the regulation of potassium channel function.

Dopamine Triggers the Maturation of Striatal Spiny Projection Neuron Excitability during a Critical Period.

Neural circuits are formed and refined during childhood, including via critical changes in neuronal excitability. Here, we investigated the ontogeny of striatal intrinsic excitability. We found that dopamine neurotransmission increases from the first to the third postnatal week in mice and precedes the reduction in spiny projection neuron (SPN) intrinsic excitability during the fourth postnatal week. In mice developmentally deficient for striatal dopamine, direct pathway D1-SPNs failed to undergo maturation of excitability past P18 and maintained hyperexcitability into adulthood. We found that the absence of D1-SPN maturation was due to altered phosphatidylinositol 4,5-biphosphate dynamics and a consequent lack of normal ontogenetic increases in Kir2 currents. Dopamine replacement corrected these deficits in SPN excitability when provided from birth or during a specific period of juvenile development (P18-P28), but not during adulthood. These results identify a sensitive period of dopamine-dependent striatal maturation, with implications for the pathophysiology and treatment of neurodevelopmental disorders.

Synaptic plasticity may underlie l-DOPA induced dyskinesia.

l-DOPA provides highly effective treatment for Parkinson's disease, but l-DOPA induced dyskinesia (LID) is a very debilitating response that eventually is presented by a majority of patients. A central issue in understanding the basis of LID is whether it is due to a response to chronic l-DOPA over years of therapy, and/or due to synaptic changes that follow the loss of dopaminergic neurotransmission and then triggered by acute l-DOPA administration. We review recent work that suggests that specific synaptic changes in the D1 dopamine receptor-expressing direct pathway striatal projection neurons due to loss of dopamine in Parkinson's disease are responsible for LID. Chronic l-DOPA may nevertheless modulate LID through priming mechanisms.

Voluntary adolescent drinking enhances excitation by low levels of alcohol in a subset of dopaminergic neurons in the ventral tegmental area.

Enhanced dopamine (DA) neurotransmission from the ventral tegmental area (VTA) to the ventral striatum is thought to drive drug self-administration and mediate positive reinforcement. We examined neuronal firing rates in slices of mouse midbrain following adolescent binge-like alcohol drinking and find that prior alcohol experience greatly enhanced the sensitivity to excitation by ethanol itself (10-50 mM) in a subset of ventral midbrain DA neurons located in the medial VTA. This enhanced response after drinking was not associated with alterations of firing rate or other measures of intrinsic excitability. In addition, the phenomenon appears to be specific to adolescent drinking, as mice that established a drinking preference only after the onset of adulthood showed no change in alcohol sensitivity. Here we demonstrate not only that drinking during adolescence induces enhanced alcohol sensitivity, but also that this DA neuronal response occurs over a range of alcohol concentrations associated with social drinking in humans.

Loss of Striatonigral GABAergic Presynaptic Inhibition Enables Motor Sensitization in Parkinsonian Mice.

Degeneration of dopamine (DA) neurons in Parkinson's disease (PD) causes hypokinesia, but DA replacement therapy can elicit exaggerated voluntary and involuntary behaviors that have been attributed to enhanced DA receptor sensitivity in striatal projection neurons. Here we reveal that in hemiparkinsonian mice, striatal D1 receptor-expressing medium spiny neurons (MSNs) directly projecting to the substantia nigra reticulata (SNr) lose tonic presynaptic inhibition by GABAB receptors. The absence of presynaptic GABAB response potentiates evoked GABA release from MSN efferents to the SNr and drives motor sensitization. This alternative mechanism of sensitization suggests a synaptic target for PD pharmacotherapy.

Presynaptic effects of levodopa and their possible role in dyskinesia.

Levodopa replacement therapy has long provided the most effective treatment for Parkinson's disease (PD). We review how this dopamine (DA) precursor enhances dopaminergic transmission by providing a greater sphere of neurotransmitter influence as a result of the confluence of increased quantal size and decreased DA reuptake, as well as loading DA as a false transmitter into surviving serotonin neuron synaptic vesicles. We further review literature on how presynaptic dysregulation of DA release after l-dopa might trigger dyskinesias in PD patients.

Fluorescent dopamine tracer resolves individual dopaminergic synapses and their activity in the brain.

We recently introduced fluorescent false neurotransmitters (FFNs) as optical tracers that enable the visualization of neurotransmitter release at individual presynaptic terminals. Here, we describe a pH-responsive FFN probe, FFN102, which as a polar dopamine transporter substrate selectively labels dopamine cell bodies and dendrites in ventral midbrain and dopaminergic synaptic terminals in dorsal striatum. FFN102 exhibits greater fluorescence emission in neutral than acidic environments, and thus affords a means to optically measure evoked release of synaptic vesicle content into the extracellular space. Simultaneously, FFN102 allows the measurement of individual synaptic terminal activity by following fluorescence loss upon stimulation. Thus, FFN102 enables not only the identification of dopamine cells and their processes in brain tissue, but also the optical measurement of functional parameters including dopamine transporter activity and dopamine release at the level of individual synapses. As such, the development of FFN102 demonstrates that, by bringing together organic chemistry and neuroscience, molecular entities can be generated that match the endogenous transmitters in selectivity and distribution, allowing for the study of both the microanatomy and functional plasticity of the normal and diseased nervous system.

Dopamine in the hippocampus is cleared by the norepinephrine transporter.

Abnormal dopaminergic neurotransmission in the hippocampus may be involved in certain aspects of cognitive dysfunction. In the hippocampus, there is little, if any, expression of dopamine transporters (DAT), indicating that the mechanism for dopamine clearance differs from that in the striatum. Here, by means of in-vivo microdialysis in freely moving rats, we tested the hypothesis that the norepinephrine transporter (NET) is involved in dopamine clearance in the hippocampus. We found that systemic administration of the selective NET inhibitor reboxetine (3 mg/kg) and the psychostimulants amphetamine (0.5 mg/kg) and cocaine (10 mg/kg) increased hippocampal dopamine efflux. Local administration of reboxetine (300 µM) produced a large increase in hippocampal dopamine levels that could not be further enhanced by the addition of the NET/DAT inhibitor nomifensine (100 µM). Administration of the specific DAT inhibitor GBR12909 at a concentration (1 mM) that robustly increased dopamine in the nucleus accumbens had a comparably smaller effect in the hippocampus. In line with a minor role of DAT in the hippocampus, we detected very little DAT in this area using ligand binding with radiolabelled RTI-55. Moreover, in contrast to raclopride (100 µM), a dopamine D2-autoreceptor antagonist, local administration of the α2-adrenoceptor antagonist idazoxan (100 µM) increased hippocampal dopamine. Taken together, our data demonstrate an interaction between dopamine and norepinephrine systems in the hippocampus. It is proposed that this interaction originates from a shared uptake mechanism at the NET level.

Multiple personalities in the ventral tegmental area.

A small number of ventral tegmental area dopamine neurons engage in numerous and apparently contradictory functions--how can this be? A clue is provided by Lammel and colleagues in this issue of Neuron: some VTA dopamine neurons display synaptic plasticity in response to cocaine, and others in response to pain, and these populations are distinguished by their axonal projections and Ih.

Histone H3 phosphorylation is under the opposite tonic control of dopamine D2 and adenosine A2A receptors in striatopallidal neurons.

The antipsychotic agent haloperidol regulates gene transcription in striatal medium spiny neurons (MSNs) by blocking dopamine D2 receptors (D2Rs). We examined the mechanisms by which haloperidol increases the phosphorylation of histone H3, a key step in the nucleosomal response. Using bacterial artificial chromosome (BAC)-transgenic mice that express EGFP under the control of the promoter of the dopamine D1 receptor (D1R) or the D2R, we found that haloperidol induced a rapid and sustained increase in the phosphorylation of histone H3 in the striatopallidal MSNs of the dorsal striatum, with no change in its acetylation. This effect was mimicked by raclopride, a selective D2R antagonist, and prevented by the blockade of adenosine A2A receptors (A2ARs), or genetic attenuation of the A2AR-associated G protein, Galpha(olf). Mutation of the cAMP-dependent phosphorylation site (Thr34) of the 32-kDa dopamine and cAMP-regulated phosphoprotein (DARPP-32) decreased the haloperidol-induced H3 phosphorylation, supporting the role of cAMP in H3 phosphorylation. Haloperidol also induced extracellular signal-regulated kinase (ERK) phosphorylation in striatopallidal MSNs, but this effect was not implicated in H3 phosphorylation. The levels of mitogen- and stress-activated kinase 1 (MSK1), which has been reported to mediate ERK-induced H3 phosphorylation, were lower in striatopallidal than in striatonigral MSNs. Moreover, haloperidol-induced H3 phosphorylation was unaltered in MSK1-knockout mice. These data indicate that, in striatopallidal MSNs, H3 phosphorylation is controlled by the opposing actions of D2Rs and A2ARs. Thus, blockade of D2Rs promotes histone H3 phosphorylation through the A2AR-mediated activation of Galpha(olf) and inhibition of protein phosphatase-1 (PP-1) through the PKA-dependent phosphorylation of DARPP-32.

Lrrk2 and alpha-synuclein are co-regulated in rodent striatum.

LRRK2, alpha-synuclein, UCH-L1 and DJ-1 are implicated in the etiology of Parkinson's disease. We show for the first time that increase in striatal alpha-synuclein levels induce increased Lrrk2 mRNA levels while Dj-1 and Uch-L1 are unchanged. We also demonstrate that a mouse strain lacking the dopamine signaling molecule DARPP-32 has significantly reduced levels of both Lrrk2 and alpha-synuclein, while mice carrying a disabling mutation of the DARPP-32 phosphorylation site T34A or lack alpha-synuclein do not show any changes. To test if striatal dopamine depletion influences Lrrk2 or alpha-synuclein expression, we used the neurotoxin 6-hydroxydopamine in rats and MitoPark mice in which there is progressive degeneration of dopamine neurons. Because striatal Lrrk2 and alpha-synuclein levels were not changed by dopamine depletion, we conclude that Lrrk2 and alpha-synuclein mRNA levels are possibly co-regulated, but they are not influenced by striatal dopamine levels.

Delayed, context- and dopamine D1 receptor-dependent activation of ERK in morphine-sensitized mice.

Exposure to cues previously associated with intake of substances of abuse can promote drug related responses. In this study, we have examined the effect of exposure to a drug-associated context on the expression of morphine psychomotor sensitization. We show that sensitization is markedly increased in mice examined 4 weeks after the last morphine injection. In addition, this incubation period confers to the environment paired with morphine the ability to increase ERK phosphorylation in the shell (but not the core) of the nucleus accumbens. Using transgenic mice with enhanced green fluorescent protein (EGFP) expression under the control of the dopamine D1 receptor (D1R) (Drd1a-EGFP) or D2 receptor promoter (Drd2-EGFP) we show that context-dependent ERK phosphorylation is restricted to D1R-expressing medium spiny neurons. Furthermore, this effect depends on D1R activation. These data show that, following repeated morphine injections, a drug-free period induces context-dependent phosphorylation of ERK in a specific population of neurons within the nucleus accumbens shell. This activation is associated to enhanced psychomotor sensitization and may be implicated in context-elicited drug seeking induced by repeated exposure to drugs of abuse.

Antagonistic cannabinoid CB1/dopamine D2 receptor interactions in striatal CB1/D2 heteromers. A combined neurochemical and behavioral analysis.

In vitro results show the ability of the CB(1) receptor agonist CP 55,940 to reduce the affinity of D(2) receptor agonist binding sites in both the dorsal and ventral striatum including the nucleus accumbens shell. This antagonistic modulation of D(2) receptor agonist affinity was found to remain and even be enhanced after G-protein activation by Gpp(NH)p. Using the FRET technique in living HEK-293T cells, the formation of CB(1)-D(2) receptor heteromers, independent of receptor occupancy, was demonstrated. These data thereby indicate that the antagonistic intramembrane CB(1)/D(2) receptor-receptor interactions may occur in CB(1)/D(2) formed heteromers. Antagonistic CB(1)/D(2) interactions were also discovered at the behavioral level through an analysis of quinpirole-induced locomotor hyperactivity in rats. The CB(1) receptor agonist CP 55,940 at a dose that did not change basal locomotion was able to block quinpirole-induced increases in locomotor activity. In addition, not only the CB(1) receptor antagonist rimonobant but also the specific A(2A) receptor antagonist MSX-3 blocked the inhibitory effect of CB(1) receptor agonist on D(2)-like receptor agonist-induced hyperlocomotion. Taken together, these results give evidence for the existence of antagonistic CB(1)/D(2) receptor-receptor interactions within CB(1)/D(2) heteromers in which A(2A) receptors may also participate.

Regulation of DARPP-32 phosphorylation by Delta9-tetrahydrocannabinol.

CB1 receptor agonists increase the state of phosphorylation of the dopamine and cAMP-regulated phosphoprotein of 32 kDa (DARPP-32) at the cAMP-dependent protein kinase site, Thr 34. This effect, which occurs in the medium spiny neurons of the striatum, has been proposed to mediate the motor depressant action of cannabinoids. In this study, we have examined the effect produced by systemic administration of Delta(9)-tetrahydrocannabinol (THC), the major component of marihuana and hashish, on DARPP-32. We show that THC increases DARPP-32 phosphorylation at Thr 34 both in dorsal striatum and nucleus accumbens. Time-course and dose-response experiments indicate that DARPP-32 phosphorylation is maximal 30 min following administration of 10mg/kg of THC. The THC-mediated increase in DARPP-32 phosphorylation is reduced by administration of the CB1 receptor antagonist, SR141716A (3mg/kg). A similar attenuation of the effect of THC is also exerted by suppression of cAMP signaling achieved using the dopamine D1 receptor antagonist, SCH23390 (0.125 mg/kg), or the adenosine A2A receptor antagonist, KW6002 (3mg/kg). These results indicate that, in the striatum, THC promotes PKA-dependent phosphorylation of DARPP-32 in striatal medium spiny neurons expressing dopamine D1 and adenosine A2A receptors.