Dopamine and Amphetamine Rapidly Increase Dopamine Transporter Trafficking to the Surface: Live-Cell Imaging Using Total Internal Reflection Fluorescence Microscopy Export

@article{citeulike:4284190, abstract = {Rapid treatment (1 min) of rat striatal synaptosomes with low-dose amphetamine increases surface expression of the dopamine transporter (DAT). Using mouse neuroblastoma N2A cells, stably transfected with green fluorescent protein-DAT, we demonstrate the real-time substrate-induced rapid trafficking of DAT to the plasma membrane using total internal reflection fluorescence microscopy (TIRFM). Both the physiological substrate, dopamine, and amphetamine began to increase surface DAT within 10 s of drug addition and steadily increased surface DAT until removal 2 min later. The substrate-induced rise in surface DAT was dose-dependent, was blocked by cocaine, and abated after drug removal. Although individual vesicle fusion was not visually detectable, exocytosis of DAT was blocked using both tetanus neurotoxin and botulinum neurotoxin C to cleave soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) proteins. Notably, the dopamine-induced increase in surface DAT was cocaine-sensitive but D2-receptor independent. TIRFM data were confirmed in human DAT-N2A cells using biotinylation, and similar effects were detected in rat striatal synaptosomes. A specific inhibitor of protein kinase C-{beta} blocked the substrate-mediated increase in surface DAT in both DAT-N2A cells and rat striatal synaptosomes. These data demonstrate that the physiological substrate, dopamine, and amphetamine rapidly increase the trafficking of DAT to the surface by a mechanism dependent on SNARE proteins and protein kinase C-{beta} but independent of dopamine D2 receptor activation. Importantly, this study suggests that the reuptake system is poised to rapidly increase its function during dopamine secretion to tightly regulate dopaminergic neurotransmission. 10.1523/JNEUROSCI.5386-08.2009}, author = {Furman, Cheryse A. and Chen, Rong and Guptaroy, Bipasha and Zhang, Minjia and Holz, Ronald W. and Gnegy, Margaret}, citeulike-article-id = {4284190}, citeulike-linkout-0 = {http://dx.doi.org/10.1523/JNEUROSCI.5386-08.2009}, citeulike-linkout-1 = {http://www.jneurosci.org/cgi/content/abstract/29/10/3328?rss=1&ssource=mfc}, citeulike-linkout-2 = {http://view.ncbi.nlm.nih.gov/pubmed/19279270}, citeulike-linkout-3 = {http://www.hubmed.org/display.cgi?uids=19279270}, day = {11}, doi = {10.1523/JNEUROSCI.5386-08.2009}, journal = {J. Neurosci.}, keywords = {amphetamine, dat, dopamine}, month = {March}, number = {10}, pages = {3328--3336}, posted-at = {2009-04-07 14:02:22}, priority = {3}, title = {Dopamine and Amphetamine Rapidly Increase Dopamine Transporter Trafficking to the Surface: Live-Cell Imaging Using Total Internal Reflection Fluorescence Microscopy}, url = {http://dx.doi.org/10.1523/JNEUROSCI.5386-08.2009}, volume = {29}, year = {2009} }

TY - JOUR ID - citeulike:4284190 L3 - citeulike-article-id:4284190 N2 - Rapid treatment (1 min) of rat striatal synaptosomes with low-dose amphetamine increases surface expression of the dopamine transporter (DAT). Using mouse neuroblastoma N2A cells, stably transfected with green fluorescent protein-DAT, we demonstrate the real-time substrate-induced rapid trafficking of DAT to the plasma membrane using total internal reflection fluorescence microscopy (TIRFM). Both the physiological substrate, dopamine, and amphetamine began to increase surface DAT within 10 s of drug addition and steadily increased surface DAT until removal 2 min later. The substrate-induced rise in surface DAT was dose-dependent, was blocked by cocaine, and abated after drug removal. Although individual vesicle fusion was not visually detectable, exocytosis of DAT was blocked using both tetanus neurotoxin and botulinum neurotoxin C to cleave soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) proteins. Notably, the dopamine-induced increase in surface DAT was cocaine-sensitive but D2-receptor independent. TIRFM data were confirmed in human DAT-N2A cells using biotinylation, and similar effects were detected in rat striatal synaptosomes. A specific inhibitor of protein kinase C-{beta} blocked the substrate-mediated increase in surface DAT in both DAT-N2A cells and rat striatal synaptosomes. These data demonstrate that the physiological substrate, dopamine, and amphetamine rapidly increase the trafficking of DAT to the surface by a mechanism dependent on SNARE proteins and protein kinase C-{beta} but independent of dopamine D2 receptor activation. Importantly, this study suggests that the reuptake system is poised to rapidly increase its function during dopamine secretion to tightly regulate dopaminergic neurotransmission. 10.1523/JNEUROSCI.5386-08.2009 IS - 10 JF - J. Neurosci. EP - 3336 TI - Dopamine and Amphetamine Rapidly Increase Dopamine Transporter Trafficking to the Surface: Live-Cell Imaging Using Total Internal Reflection Fluorescence Microscopy VL - 29 SP - 3328 KW - amphetamine KW - dat KW - dopamine AU - Furman, Cheryse AU - Chen, Rong AU - Guptaroy, Bipasha AU - Zhang, Minjia AU - Holz, Ronald AU - Gnegy, Margaret PY - 2009/03/11/ UR - http://dx.doi.org/10.1523/JNEUROSCI.5386-08.2009 ER -