Dopamine was first identified as a neurotransmitter localized to the midbrain over 50 years ago. The dopamine transporter (DAT; SLC6A3) and the vesicular monoamine transporter 2 (VMAT2; SLC18A2) are ...regulators of dopamine homeostasis in the presynaptic neuron. DAT transports dopamine from the extracellular space into the cytosol of the presynaptic terminal. VMAT2 then packages this cytosolic dopamine into vesicular compartments for subsequent release upon neurotransmission. Thus, DAT and VMAT2 act in concert to move the transmitter efficiently throughout the neuron. Accumulation of dopamine in the neuronal cytosol can trigger oxidative stress and neurotoxicity, suggesting that the proper compartmentalization of dopamine is critical for neuron function and risk of disease. For decades, studies have examined the effects of reduced transporter function in mice (e.g. DAT‐KO, VMAT2‐KO, VMAT2‐deficient). However, we have only recently been able to assess the effects of elevated transporter expression using BAC transgenic methods (DAT‐tg, VMAT2‐HI mice). Complemented with in vitro work and neurochemical techniques to assess dopamine compartmentalization, a new focus on the importance of transporter proteins as both models of human disease and potential drug targets has emerged. Here, we review the importance of DAT and VMAT2 function in the delicate balance of neuronal dopamine.
This review describes the importance of the dopamine transporter (DAT) and the vesicular monoamine transporter 2 (VMAT2) on dopamine compartmentalization and neuronal health (A–C). While theoretical, this schematic highlights emerging evidence from mouse models of varying transporter levels. We predict that the continuum of transporter function in these animal models will allow for new discoveries concerning endogenous dopamine handling, pharmacological manipulation of the transporters, and dopamine‐dependent behaviors (D).
N-methyl-D-aspartate receptors (NMDARs) are required to shape activity-dependent connections in the developing and adult brain. Impaired NMDAR signalling through genetic or environmental insults ...causes a constellation of neurodevelopmental disorders that manifest as intellectual disability, epilepsy, autism, or schizophrenia. It is not clear whether the developmental impacts of NMDAR dysfunction can be overcome by interventions in adulthood. This question is paramount for neurodevelopmental disorders arising from mutations that occur in the GRIN genes, which encode NMDAR subunits, and the broader set of mutations that disrupt NMDAR function. We developed a mouse model where a congenital loss-of-function allele of Grin1 can be restored to wild type by gene editing with Cre recombinase. Rescue of NMDARs in adult mice yields surprisingly robust improvements in cognitive functions, including those that are refractory to treatment with current medications. These results suggest that neurodevelopmental disorders arising from NMDAR deficiency can be effectively treated in adults.
In the last four to five years, the view that G protein-coupled receptors (GPCRs) function as monomeric proteins has been challenged by numerous studies, which suggests that GPCRs exist as dimers or ...even higher-structure oligomers. Recently, biophysical methods based on luminescence and fluorescence energy transfer have confirmed the existence of such oligomeric complexes in living cells. Although no consensus exists on the role of receptor dimerization, converging evidence suggests potential roles in various aspects of receptor biogenesis and function. In several cases, receptors appear to fold as constitutive dimers early after biosynthesis, whereas ligand-promoted dimerization at the cell surface has been proposed for others. The reports of heterodimerization between receptor subtypes suggest a potential level of receptor complexity that could account for previously unexpected pharmacological diversities. In addition to fundamentally changing our views on the structure and activation processes of GPCRs, the concept of homo- and heterodimerization could have dramatic impacts on drug development and screening.
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DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, UILJ, UKNU, UL, UM, UPUK
Disruption of neurotransmitter vesicle dynamics (transport, capacity, release) has been implicated in a variety of neurodegenerative and neuropsychiatric conditions. Here, we report a novel mouse ...model of enhanced vesicular function via bacterial artificial chromosome (BAC)-mediated overexpression of the vesicular monoamine transporter 2 (VMAT2; Slc18a2). A twofold increase in vesicular transport enhances the vesicular capacity for dopamine (56%), dopamine vesicle volume (33%), and basal tissue dopamine levels (21%) in the mouse striatum. The elevated vesicular capacity leads to an increase in stimulated dopamine release (84%) and extracellular dopamine levels (44%). VMAT2-overexpressing mice show improved outcomes on anxiety and depressive-like behaviors and increased basal locomotor activity (41%). Finally, these mice exhibit significant protection from neurotoxic insult by the dopaminergic toxicant 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP), as measured by reduced dopamine terminal damage and substantia nigra pars compacta cell loss. The increased release of dopamine and neuroprotection from MPTP toxicity in the VMAT2-overexpressing mice suggest that interventions aimed at enhancing vesicular capacity may be of therapeutic benefit in Parkinson disease.
Since the unexpected discovery of the antipsychotic activity of chlorpromazine, a variety of therapeutic agents have been developed for the treatment of schizophrenia. Despite differences in their ...activities at various neurotransmitter systems, all clinically effective antipsychotics share the ability to interact with D2 class dopamine receptors (D2R). D2R mediate their physiological effects via both G protein-dependent and independent (β-arrestin 2-dependent) signaling, but the role of these D2R-mediated signaling events in the actions of antipsychotics remains unclear. We demonstrate here that while different classes of antipsychotics have complex pharmacological profiles at G protein-dependent D2R long isoform (D2LR) signaling, they share the common property of antagonizing dopamine-mediated interaction of D2LR with β-arrestin 2. Using two cellular assays based on a bioluminescence resonance energy transfer (BRET) approach, we demonstrate that a series of antipsychotics including haloperidol, clozapine, aripiprazole, chlorpromazine, quetiapine, olanzapine, risperidone, and ziprasidone all potently antagonize the β-arrestin 2 recruitment to D2LR induced by quinpirole. However, these antipsychotics have various effects on D2LR mediated Gi/o protein activation ranging from inverse to partial agonists and antagonists with highly variable efficacies and potencies at quinpirole-induced cAMP inhibition. These results suggest that the different classes of clinically effective antipsychotics share a common molecular mechanism involving inhibition of D2LR/β-arrestin 2 mediated signaling. Thus, selective targeting of D2LR/β-arrestin 2 interaction and related signaling pathways may provide new opportunities for antipsychotic development.
The ability of dopamine receptors to interact with other receptor subtypes may provide mechanisms for modulating dopamine-related functions and behaviors. In particular, there is evidence suggesting ...that the trace amine-associated receptor 1 (TAAR1) affects the dopaminergic system by regulating the firing rate of dopaminergic neurons or by altering dopamine D2 receptor (D2R) responsiveness to ligands. TAAR1 is a Gα(s) protein-coupled receptor that is activated by biogenic amines, "trace amines," such as β-phenylethylamine (β-PEA) and tyramine that are normally found at low concentrations in the mammalian brain. In the present study, we investigated the biochemical mechanism of interaction between TAAR1 and D2R and the role this interaction plays in D2R-related signaling and behaviors. Using a bioluminescence resonance energy transfer biosensor for cAMP, we demonstrated that the D2R antagonists haloperidol, raclopride, and amisulpride were able to enhance selectively a TAAR1-mediated β-PEA increase of cAMP. Moreover, TAAR1 and D2R were able to form heterodimers when coexpressed in human embryonic kidney 293 cells, and this direct interaction was disrupted in the presence of haloperidol. In addition, in mice lacking TAAR1, haloperidol-induced striatal c-Fos expression and catalepsy were significantly reduced. Taken together, these data suggest that TAAR1 and D2R have functional and physical interactions that could be critical for the modulation of the dopaminergic system by TAAR1 in vivo.
In Parkinson's disease, dopamine‐containing nigrostriatal neurons undergo profound degeneration. Tyrosine hydroxylase (TH) is the rate‐limiting enzyme in dopamine biosynthesis. TH increases in vitro ...formation of reactive oxygen species, and previous animal studies have reported links between cytosolic dopamine build‐up and oxidative stress. To examine effects of increased TH activity in catecholaminergic neurons in vivo, we generated TH‐over‐expressing mice (TH‐HI) using a BAC‐transgenic approach that results in over‐expression of TH with endogenous patterns of expression. The transgenic mice were characterized by western blot, qPCR, and immunohistochemistry. Tissue contents of dopamine, its metabolites, and markers of oxidative stress were evaluated. TH‐HI mice had a 3‐fold increase in total and phosphorylated TH levels and an increased rate of dopamine synthesis. Coincident with elevated dopamine turnover, TH‐HI mice showed increased striatal production of H2O2 and reduced glutathione levels. In addition, TH‐HI mice had elevated striatal levels of the neurotoxic dopamine metabolites 3,4‐dihydroxyphenylacetaldehyde and 5‐S‐cysteinyl‐dopamine and were more susceptible than wild‐type mice to the effects of amphetamine and methamphetamine. These results demonstrate that increased TH alone is sufficient to produce oxidative stress in vivo, build up autotoxic dopamine metabolites, and augment toxicity.
This paper investigates the effect of increased activity of tyrosine hydroxylase (TH), the rate‐limiting enzyme in catecholamine synthesis, in a novel line of TH over‐expressing mice. Past studies suggest that in synucleinopathies, early pathological changes can result in decreased TH regulation and increased activity. Here, we show that increased TH activity is sufficient to increase H2O2 and elevate levels of cysteinylated dopamine (Cys‐DA) and 3,4‐dihydroxyphenylacetaldehyde (DOPAL)—respective autotoxic products of enzymatic and spontaneous dopamine oxidation—coincident with increased dopamine turnover. These findings suggest that TH dysregulation could present a source of dopamine‐related oxidative stress unique to cells most vulnerable in Parkinson's disease. This article is accompanied by an Editorial Highlight by Elisa Greggio (https://doi.org/10.1111/jnc.15442).
NMDA receptor dysfunction is central to the encephalopathies caused by missense mutations in the NMDA receptor subunit genes. Missense variants of GRIN1, GRIN2A, and GRIN2B cause similar syndromes ...with varying severity of intellectual impairment, autism, epilepsy, and motor dysfunction. To gain insight into possible biomarkers of NMDAR hypofunction, we asked whether a loss-of-function variant in the Grin1 gene would cause structural changes in the brain that could be detected by MRI. We also studied the developmental trajectory of these changes to determine whether structural changes coincided with reported cognitive impairments in the mice. We performed magnetic resonance imaging in male Grin1−/− knockdown mice (GluN1KD) that were three, six, or twelve weeks old. Deformation-based morphometry was used to assess neuroanatomical differences. Volumetric reductions were detected in substantia nigra and striatum of GluN1KD mice at all ages. Changes in limbic structures were only evident at six weeks of age. Reductions in white matter volumes were first evident at three weeks, and additional deficits were detected at six and twelve weeks. FluoroJade immunofluorescence revealed degenerating neurons in twelve-week old GluN1KD mice. We conclude that Grin1 loss-of-function mutations cause volume reductions in dopaminergic structures early in development, while changes to limbic and white matter structures are delayed and are more pronounced in post-adolescent ages. The evidence of degenerating neurons in the mature brain indicates an ongoing process of cell loss as a consequence of NMDAR hypofunction.
•Mice with a Grin1 loss-of-function mutation have microcephaly across three developmental ages.•The volume of dopaminergic brain structures is reduced in juvenile, adolescent, and adult mutant mice.•Reduced white matter volumes are detected in juvenile Grin1 mutant mice and become more pronouncedin adult mutant mice.•Adult Grin1 mutant mice show indications of ongoing neurodegeneration in the striatum.
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