A5089868900- Neurotransmitter Receptor Influence on Behavior
- Neuroscience and Neuropharmacology Research
- Receptor Mechanisms and Signaling
- Amino Acid Enzymes and Metabolism
- Ion channel regulation and function
- Drug Transport and Resistance Mechanisms
- Ion Transport and Channel Regulation
- Lipid Membrane Structure and Behavior
- Analytical Chemistry and Chromatography
- Molecular Sensors and Ion Detection
- Various Chemistry Research Topics
- Pancreatic function and diabetes
- Chemical Synthesis and Analysis
- Bacterial Genetics and Biotechnology
- Neuroscience, Education and Cognitive Function
- Forensic Toxicology and Drug Analysis
- RNA and protein synthesis mechanisms
- GABA and Rice Research
- Cellular transport and secretion
- Nitric Oxide and Endothelin Effects
- DNA and Nucleic Acid Chemistry
- Obsessive-Compulsive Spectrum Disorders
- Neuropeptides and Animal Physiology
- Pharmacology and Obesity Treatment
- ATP Synthase and ATPases Research
Yale University
2015-2024
University of Kansas
2012-2021
Medical University of Vienna
2015
University of Vienna
2015
National Institute on Drug Abuse
2015
National Institutes of Health
2015
Icahn School of Medicine at Mount Sinai
2003
Emory University
1994
Hebrew University of Jerusalem
1980-1990
La Roche College
1975-1976
MDMA ("ecstasy") has been widely reported as a drug of abuse and neurotoxin. This report describes the mechanism action at serotonin transporters from plasma membranes secretory vesicles. stimulates efflux both types membrane vesicle. In vesicles isolated human platelets, inhibits transport [3H]imipramine binding by direct interaction with Na(+)-dependent transporter. radiolabel preloaded [3H]serotonin in stereo-specific, Na(+)-dependent, imipramine-sensitive manner characteristic...
We have constructed stable cell lines expressing transporters for dopamine (DA), norepinephrine (NE), and serotonin (5-HT) by transfection with cloned cDNAs. The parental LLC-PK1 does not express any of these neurotransmitter transporters. Therefore, monoamine transport activities in each are due to the transfected DNA only, allowing comparison same background. Drug inhibition profiles line distinct as expected transporter. LLC-NET LLC-DAT cells transported both NE DA types exhibited a lower...
Crystal structures of LeuT, a bacterial homologue mammalian neurotransmitter transporters, show molecule bound substrate that is essentially exposed to the extracellular space but occluded from cytoplasm. Thus, there must exist an alternate conformation for LeuT in which accessible cytoplasm and corresponding mechanism switches accessibility one side membrane other. Here, we identify cytoplasmic pathway serotonin transporter (SERT) (a member same family as LeuT). We also propose model...
We have used the pH-dependent fluorochrome fluorescein-dextran (FD) to study acidification of prelysosomal vacuoles (endosomes) and lysosomes isolated from cultured macrophages fibroblasts. FD was internalized by pinocytosis under conditions that allowed its selective localization in endosomes (1- 5-min pulse) or (5-min pulse, 30-min chase). Fibroblasts were also exposed at 20 degrees C, which temperature endosome-lysosome fusion is inhibited. Cells homogenized labeled organelles separated...
Two forms of serotonin transporter (SERT) were prepared with different epitope tags. When co-expressed in HeLa cells, the form containing a FLAG tag (Res-FLAG) was associated c-myc (Sens-myc). Antibody against precipitated Res-FLAG from detergent extracts cells expressing both forms, but not when expressed alone. The specificity interaction demonstrated by observation that anti-myc antibodies did precipitate unrelated vesicular stomatitis virus coat glycoprotein it Sens-myc. Sens-myc...
Plasma membrane vesicles isolated from human platelets accumulate 5-hydroxytryptamine when an electrical potential (interior negative) or Na+ gradient (out greater than in) is imposed across the vesicle membrane. Kinetic studies reveal a Km of 0.5 micronM for transport process. Uptake inhibited strongly by tricyclic antidepressants and ionophores such as gramicidin which catalyze transmembrane exchange K+. Transport absolutely dependent upon external Cl- only mildly, if at all, reserpine,...
Twenty residues in the third transmembrane domain of serotonin transporter (SERT) were mutated, one at a time, to cysteine. Almost all these mutants fully active for (5-HT) transport and insensitive inactivation by positively charged cysteine reagent [2-(trimethylammonium)ethyl]methanethiosul-fonate (MTSET). Two mutants, I172C I179C, sensitive rapid MTSET but relatively negatively (2-sulfonatoethyl)methanethiosulfonate (MTSES). Inactivation was blocked 5-HT cocaine, I179C not similarly...
The recent determination of the crystal structure leucine transporter from Aquifex aeolicus (aaLeuT) has provided significant insights into function neurotransmitter:sodium symporters. Transport by aaLeuT is Cl(-) independent, whereas many symporters higher organisms depend on ions. However, only ion identified in interacts with nonconserved residues extracellular loops, and thus relevance this binding site unclear. Here, we use calculations pK(A)s homology modeling to predict location a...
LLC-PK1 cells have been stably transfected with cDNAs encoding the human norepinephrine transporter (NET), rat dopamine (DAT), and serotonin transporter. Using these cell lines, specificity of each toward agents that inhibit substrate influx stimulate efflux across plasma membrane was examined. With 1-methyl-4-phenylpyridinium as a for DAT NET transporter, demonstrated distinct pattern inhibition by panel amphetamine derivatives analogs, including amphetamine, methamphetamine (also known...
Platelet plasma membrane vesicles accumulate 5-hy- droxytryptamine in response to a gradient of NaCl (out > in). Although potassium ion is not required for this process, K’ concentration (in out) elevates the steady state level 5-hydroxytryptamine accu- mulation 5-fold. Transport independent gen- eration an electrical potential (interior negative) across vesicle when present inter- nally, but its absence trans- port apparently electrogenic, with net positive charge crossing 5-hydroxytrypt-...
Plasma membrane vesicles isolated from porcine blood platelets take up approximately 8 to 15 pmol of [3H]imipramine per mg protein. This apparent binding requires Na+ in the external medium and is reversed by 5-hydroxytryptamine fluoxetine. The KD for imipramine uptake 23 nM, which agrees well with KI competitive inhibition transport imipramine. In contrast transport, not dependent on transmembrane K+ gradients insensitive ionophores such as nigericin gramicidin dissipate these gradients....
We discuss the structural and morphological properties of galaxies in a z = 1.62 proto-cluster using near-IR imaging data from Hubble Space Telescope Wide Field Camera 3 Cosmic Assembly Near-IR Deep Extragalactic Legacy Survey (CANDELS). The cluster exhibit clear color–morphology relation: with colors quiescent stellar populations generally have morphologies consistent spheroids, ongoing star formation disk-like irregular morphologies. size distribution shows deficit compact (≲ 1 kpc),...
The transmembrane topology of the serotonin transporter (SERT) has been examined by measuring reactivity selected lysine and cysteine residues with extracellular reagents. An impermeant biotinylating reagent, sulfosuccinimidyl 2-(biotinamido)ethyl-1,3-dithiopropionate (NHS-SS-biotin), was shown to label SERT transiently expressed in cultured cells. Replacement four that were predicted lie external hydrophilic loops (eK-less) largely prevented biotinylation reaction. Likewise,...
Hydropathy analysis predicts three cysteines (C109, C200, and C209) in extracellular loops of the rat serotonin transporter (SERT). We mutated these residues, singly combination, to either alanine or serine expressed mutant transporters HeLa cells using vaccinia−T7 transient expression system. Mutation C109 had no effect on transport activity surface transporter. In Na+-containing solutions, methanethiosulfonate (MTS) reagents little wild type C109A mutant. When Na+ was replaced with Li+,...
The vaccinia/T7 transient expression system, which results in rapid, high-level of proteins encoded by plasmids bearing T7 promoters, provides a powerful strategy for the cloning membrane transporters. To test feasibility this approach, we introduced rabbit Na+/glucose transporter liposome-mediated transfection into vaccinia infected HeLa cells and determined characteristics sensitivity induced [14C]α-methyl d-glucopyranoside uptake. We observed rapid (4–12 h) saturable (Kt = 342 μm) uptake...
A rarely occurring variant of human serotonin transporter (hSERT) was tested for its functional consequences in HeLa and COS-7 cells. The variant, which Ile-425 is converted to Val, significantly different from wild type with respect catalytic properties. In both cell types, rates (5-HT) transport were higher the I425V variant. Both an increase Vmax a decrease KM caused this rate. not accounted by increases expression or distribution between surface intracellular pools. accompanied KD...
The third transmembrane domain (TM3) of serotonin transporter (SERT) contains two isoleucine residues previously proposed to be involved in binding and transport serotonin. When Ile-172 was replaced with cysteine, SERT became sensitive inactivation by externally added [2-(trimethylammonium)ethyl]methanethio-sulfonate (MTSET). disulfide product this not reduction sulfhydryl compounds, but apparently reacted intracellular reducing agents spontaneously regenerate active SERT. apparent...