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Addiction Biology Nov 2022As well known, cocaine induces stimulant effects and dopamine transporter (DAT) trafficking to the plasma membrane of dopaminergic neurons. In the present study, we...
As well known, cocaine induces stimulant effects and dopamine transporter (DAT) trafficking to the plasma membrane of dopaminergic neurons. In the present study, we examined cocaine-induced hyperactivity along with cocaine-induced DAT trafficking and the recovery rate of the dopaminergic system in female rats in comparison with male rats, demonstrating interesting gender differences. Female rats are initially more sensitive to cocaine than male rats in terms of both the DAT trafficking and hyperactivity induced by cocaine. Particularly, intraperitoneal (i.p.) administration of 5 mg/kg cocaine induced significant hyperactivity and DAT trafficking in female rats but not in male rats. After repeated cocaine exposures (i.e., i.p. administration of 20 mg/kg cocaine every other day from Day 0 to Day 32), cocaine-induced hyperactivity in female rats gradually became a clear pattern of two phases, with the first phase of the hyperactivity lasting for only a few minutes and the second phase lasting for over an hour beginning at ~30 min, which is clearly different from that of male rats. It has also been demonstrated that the striatal DAT distribution of female rats may recover faster than that of male rats after multiple cocaine exposures. Nevertheless, despite the remarkable gender differences, our recently developed long-acting cocaine hydrolase, known as CocH5-Fc(M6), can similarly and effectively block cocaine-induced DAT trafficking and hyperactivity in both male and female rats.
Topics: Male; Female; Rats; Animals; Dopamine Plasma Membrane Transport Proteins; Cocaine; Sex Factors; Dopamine Uptake Inhibitors; Cell Membrane
PubMed: 36301205
DOI: 10.1111/adb.13236 -
Journal of Neurogenetics Mar 2016The dopamine transporter (DAT) plays an important homeostatic role in the control of both the extracellular and intraneuronal concentrations of dopamine, thereby... (Review)
Review
The dopamine transporter (DAT) plays an important homeostatic role in the control of both the extracellular and intraneuronal concentrations of dopamine, thereby providing effective control over activity of dopaminergic transmission. Since brain dopamine is known to be involved in numerous neuropsychiatric disorders, investigations using mice with genetically altered DAT function and thus intensity of dopamine-mediated signaling have provided numerous insights into the pathology of these disorders and novel pathological mechanisms that could be targeted to provide new therapeutic approaches for these disorders. In this brief overview, we discuss recent investigations involving animals with genetically altered DAT function, particularly focusing on translational studies providing new insights into pathology and pharmacology of dopamine-related disorders. Perspective applications of these and newly developed models of DAT dysfunction are also discussed.
Topics: Animals; Disease Models, Animal; Dopamine Plasma Membrane Transport Proteins; Mice; Translational Research, Biomedical
PubMed: 27276191
DOI: 10.3109/01677063.2016.1144751 -
The Journal of Biological Chemistry Apr 2020Following its evoked release, dopamine (DA) signaling is rapidly terminated by presynaptic reuptake, mediated by the cocaine-sensitive DA transporter (DAT). DAT surface...
Following its evoked release, dopamine (DA) signaling is rapidly terminated by presynaptic reuptake, mediated by the cocaine-sensitive DA transporter (DAT). DAT surface availability is dynamically regulated by endocytic trafficking, and direct protein kinase C (PKC) activation acutely diminishes DAT surface expression by accelerating DAT internalization. Previous cell line studies demonstrated that PKC-stimulated DAT endocytosis requires both Ack1 inactivation, which releases a DAT-specific endocytic brake, and the neuronal GTPase, Rit2, which binds DAT. However, it is unknown whether Rit2 is required for PKC-stimulated DAT endocytosis in DAergic terminals or whether there are region- and/or sex-dependent differences in PKC-stimulated DAT trafficking. Moreover, the mechanisms by which Rit2 controls PKC-stimulated DAT endocytosis are unknown. Here, we directly examined these important questions. studies revealed that PKC activation acutely decreased DAT surface expression selectively in ventral, but not dorsal, striatum. AAV-mediated, conditional Rit2 knockdown in DAergic neurons impacted baseline DAT surface:intracellular distribution in DAergic terminals from female ventral, but not dorsal, striatum. Further, Rit2 was required for PKC-stimulated DAT internalization in both male and female ventral striatum. FRET and surface pulldown studies in cell lines revealed that PKC activation drives DAT-Rit2 surface dissociation and that the DAT N terminus is required for both PKC-mediated DAT-Rit2 dissociation and DAT internalization. Finally, we found that Rit2 and Ack1 independently converge on DAT to facilitate PKC-stimulated DAT endocytosis. Together, our data provide greater insight into mechanisms that mediate PKC-regulated DAT internalization and reveal unexpected region-specific differences in PKC-stimulated DAT trafficking in DAergic terminals.
Topics: Animals; Binding Sites; Cell Line, Tumor; Corpus Striatum; Dopamine Plasma Membrane Transport Proteins; Dopaminergic Neurons; Endocytosis; Female; HEK293 Cells; Humans; Male; Mice; Mice, Inbred C57BL; Monomeric GTP-Binding Proteins; Protein Binding; Protein Kinase C
PubMed: 32132171
DOI: 10.1074/jbc.RA120.012628 -
Neurochemical Research Jan 2022The dopamine transporter (DAT) mediates the inactivation of released dopamine (DA) through its reuptake, and thereby plays an important homeostatic role in dopaminergic...
The dopamine transporter (DAT) mediates the inactivation of released dopamine (DA) through its reuptake, and thereby plays an important homeostatic role in dopaminergic neurotransmission. Amphetamines exert their stimulant effects by targeting DAT and inducing the reverse transport of DA, leading to a dramatic increase of extracellular DA. Animal models have proven critical to investigating the molecular and cellular mechanisms underlying transporter function and its modulation by psychostimulants such as amphetamine. Here we establish a behavioral model for amphetamine action using adult Drosophila melanogaster. We use it to characterize the effects of amphetamine on sleep and sleep architecture. Our data show that amphetamine induces hyperactivity and disrupts sleep in a DA-dependent manner. Flies that do not express a functional DAT (dDAT null mutants) have been shown to be hyperactive and to exhibit significantly reduced sleep at baseline. Our data show that, in contrast to its action in control flies, amphetamine decreases the locomotor activity of dDAT null mutants and restores their sleep by modulating distinct aspects of sleep structure. To begin to explore the circuitry involved in the actions of amphetamine on sleep, we also describe the localization of dDAT throughout the fly brain, particularly in neuropils known to regulate sleep. Together, our data establish Drosophila as a robust model for studying the regulatory mechanisms that govern DAT function and psychostimulant action.
Topics: Amphetamine; Animals; Dopamine Plasma Membrane Transport Proteins; Drosophila; Drosophila melanogaster; Sleep
PubMed: 33630236
DOI: 10.1007/s11064-021-03275-4 -
Nature Communications May 2022The dopamine transporter facilitates dopamine reuptake from the extracellular space to terminate neurotransmission. The transporter belongs to the...
The dopamine transporter facilitates dopamine reuptake from the extracellular space to terminate neurotransmission. The transporter belongs to the neurotransmitter:sodium symporter family, which includes transporters for serotonin, norepinephrine, and GABA that utilize the Na gradient to drive the uptake of substrate. Decades ago, it was shown that the serotonin transporter also antiports K, but investigations of K-coupled transport in other neurotransmitter:sodium symporters have been inconclusive. Here, we show that ligand binding to the Drosophila- and human dopamine transporters are inhibited by K, and the conformational dynamics of the Drosophila dopamine transporter in K are divergent from the apo- and Na-states. Furthermore, we find that K increases dopamine uptake by the Drosophila dopamine transporter in liposomes, and visualize Na and K fluxes in single proteoliposomes using fluorescent ion indicators. Our results expand on the fundamentals of dopamine transport and prompt a reevaluation of the impact of K on other transporters in this pharmacologically important family.
Topics: Animals; Dopamine; Dopamine Plasma Membrane Transport Proteins; Drosophila; Ion Transport; Ions; Neurotransmitter Agents; Potassium; Serotonin Plasma Membrane Transport Proteins; Sodium; Symporters
PubMed: 35508541
DOI: 10.1038/s41467-022-30154-5 -
Drug and Alcohol Dependence Feb 2015Treatment of stimulant-use disorders remains a formidable challenge, and the dopamine transporter (DAT) remains a potential target for antagonist or agonist-like... (Review)
Review
BACKGROUND
Treatment of stimulant-use disorders remains a formidable challenge, and the dopamine transporter (DAT) remains a potential target for antagonist or agonist-like substitution therapies.
METHODS
This review focuses on DAT ligands, such as benztropine, GBR 12909, modafinil, and DAT substrates derived from phenethylamine or cathinone that have atypical DAT-inhibitor effects, either in vitro or in vivo. The compounds are described from a molecular mechanistic, behavioral, and medicinal-chemical perspective.
RESULTS
Possible mechanisms for atypicality at the molecular level can be deduced from the conformational cycle for substrate translocation. For each conformation, a crystal structure of a bacterial homolog is available, with a possible role of cholesterol, which is also present in the crystal of Drosophila DAT. Although there is a direct relationship between behavioral potencies of most DAT inhibitors and their DAT affinities, a number of compounds bind to the DAT and inhibit dopamine uptake but do not share cocaine-like effects. Such atypical behavior, depending on the compound, may be related to slow DAT association, combined sigma-receptor actions, or bias for cytosol-facing DAT. Some structures are sterically small enough to serve as DAT substrates but large enough to also inhibit transport. Such compounds may display partial DA releasing effects, and may be combined with release or uptake inhibition at other monoamine transporters.
CONCLUSIONS
Mechanisms of atypical DAT inhibitors may serve as targets for the development of treatments for stimulant abuse. These mechanisms are novel and their further exploration may produce compounds with unique therapeutic potential as treatments for stimulant abuse.
Topics: Animals; Benzhydryl Compounds; Benztropine; Central Nervous System Stimulants; Cocaine; Dopamine Plasma Membrane Transport Proteins; Drug Delivery Systems; Humans; Ligands; Modafinil; Protein Binding; Protein Structure, Secondary; Protein Structure, Tertiary
PubMed: 25548026
DOI: 10.1016/j.drugalcdep.2014.12.005 -
Journal of Nuclear Medicine Technology Mar 2019Idiopathic Parkinson disease (PD) is a progressive neurologic condition that affects the dopamine transporters in the substantia nigra of the brain. Currently, more than... (Review)
Review
Idiopathic Parkinson disease (PD) is a progressive neurologic condition that affects the dopamine transporters in the substantia nigra of the brain. Currently, more than 10 million people are living with this disease worldwide, with thousands of newly diagnosed and undiagnosed cases added every year. The disease is difficult to differentiate from other similar disorders, as symptoms widely vary and can mimic other conditions. Classic PD symptoms may look similar to essential tremor and other parkinsonian syndromes. The I-ioflupane dopamine transporter (DaT) protocol differentiates PD from essential tremor through in vivo testing with SPECT imaging. The DaT protocol commonly relies on a semiquantitative analysis and visual interpretations of the images, which may produce inaccurate results due to human error. DaTQUANT software (GE Healthcare) was created in 2013 as an adjunct processing tool with advanced quantitative uptake methods and a designated normals database for a more accurate assessment of a patient's case. DaTQUANT has proven to be a vital protocol component for an accurate differentiation of PD from essential tremor. Current use of the software has been rather limited, so a greater push for education and implementation will be key for its success.
Topics: Dopamine Plasma Membrane Transport Proteins; Humans; Image Processing, Computer-Assisted; Parkinson Disease; Software
PubMed: 30683690
DOI: 10.2967/jnmt.118.222349 -
Molecular Psychiatry Feb 2022The human dopamine transporter gene SLC6A3 has been consistently implicated in several neuropsychiatric diseases but the disease mechanism remains elusive. In this risk...
The human dopamine transporter gene SLC6A3 has been consistently implicated in several neuropsychiatric diseases but the disease mechanism remains elusive. In this risk synthesis, we have concluded that SLC6A3 represents an increasingly recognized risk with a growing number of familial mutants associated with neuropsychiatric and neurological disorders. At least five loci were related to common and severe diseases including alcohol use disorder (high activity variant), attention-deficit/hyperactivity disorder (low activity variant), autism (familial proteins with mutated networking) and movement disorders (both regulatory variants and familial mutations). Association signals depended on genetic markers used as well as ethnicity examined. Strong haplotype selection and gene-wide epistases support multimarker assessment of functional variations and phenotype associations. Inclusion of its promoter region's functional markers such as DNPi (rs67175440) and 5'VNTR (rs70957367) may help delineate condensate-based risk action, testing a locus-pathway-phenotype hypothesis for one gene-multidisease etiology.
Topics: Attention Deficit Disorder with Hyperactivity; Dopamine Plasma Membrane Transport Proteins; Haplotypes; Humans; Mutation; Phenotype
PubMed: 34650206
DOI: 10.1038/s41380-021-01341-5 -
Journal of Nuclear Medicine Technology Jun 2020Racial and ethnic disparities in the prevalence of neurodegenerative diseases exist. This study examined the agreement between gold standard diagnosis and visual... (Comparative Study)
Comparative Study
Racial and ethnic disparities in the prevalence of neurodegenerative diseases exist. This study examined the agreement between gold standard diagnosis and visual assessment of dopamine transporter (DaT) imaging in Hispanic and non-Hispanic patients being evaluated for Parkinsonian syndromes (PS). A retrospective review of DaT imaging and demographic data was performed with institutional review board approval. Documented interpretation by visual assessment was used to classify scans as normal or abnormal. The gold standard for the final diagnosis of PS was determined by a neurologist after 2 or more years of clinical follow-up. Data were analyzed with a -test for uncorrelated samples. In 30 Hispanic patients, DaT imaging was abnormal in 17, normal in 12, and nondiagnostic in 1. Of those with abnormal imaging, PS was confirmed in 16 of 17. Of those with normal imaging, no PS was confirmed in any patient. Sensitivity was 100%, and specificity was 92%. The single patient with nondiagnostic imaging was excluded. Of 77 non-Hispanic patients, visual assessment of DaT imaging was abnormal in 51. Of those with abnormal imaging, PS was confirmed in 48 of 51. Of those with normal imaging, no PS was confirmed in 22 of 26. Sensitivity was 92%, and specificity was 88%. There was no statistically significant difference ( = 0.34) in the rates of agreement between the gold standard and DaT imaging in Hispanic versus non-Hispanic patients. The study sample size afforded a power of 0.60. No significant difference was found in the accuracy of DaT imaging between Hispanic and non-Hispanic patients. Accuracy was high for both groups.
Topics: Adult; Aged; Aged, 80 and over; Diagnostic Imaging; Dopamine Plasma Membrane Transport Proteins; Female; Hispanic or Latino; Humans; Iodine Radioisotopes; Male; Middle Aged; Parkinsonian Disorders; Retrospective Studies
PubMed: 31604897
DOI: 10.2967/jnmt.119.231423 -
The Journal of Biological Chemistry Nov 2015The dopamine transporter is a neuronal protein that drives the presynaptic reuptake of dopamine (DA) and is the major determinant of transmitter availability in the...
The dopamine transporter is a neuronal protein that drives the presynaptic reuptake of dopamine (DA) and is the major determinant of transmitter availability in the brain. Dopamine transporter function is regulated by protein kinase C (PKC) and other signaling pathways through mechanisms that are complex and poorly understood. Here we investigate the role of Ser-7 phosphorylation and Cys-580 palmitoylation in mediating steady-state transport kinetics and PKC-stimulated transport down-regulation. Using both mutational and pharmacological approaches, we demonstrate that these post-translational modifications are reciprocally regulated, leading to transporter populations that display high phosphorylation-low palmitoylation or low phosphorylation-high palmitoylation. The balance between the modifications dictates transport capacity, as conditions that promote high phosphorylation or low palmitoylation reduce transport Vmax and enhance PKC-stimulated down-regulation, whereas conditions that promote low phosphorylation or high palmitoylation increase transport Vmax and suppress PKC-stimulated down-regulation. Transitions between these functional states occur when endocytosis is blocked or undetectable, indicating that the modifications kinetically regulate the velocity of surface transporters. These findings reveal a novel mechanism for control of DA reuptake that may represent a point of dysregulation in DA imbalance disorders.
Topics: Cell Line; Dopamine; Dopamine Plasma Membrane Transport Proteins; Down-Regulation; Endocytosis; Humans; Kinetics; Lipoylation; Protein Kinase C
PubMed: 26424792
DOI: 10.1074/jbc.M115.667055