Regulation of Amphetamine-stimulated Dopamine Efflux by Protein Kinase C β

Evidence suggests that protein kinase C (PKC) and intracellular calcium are important for amphetamine-stimulated outward transport of dopamine in rat striatum. In this study, we examined the effect of select PKC isoforms on amphetamine-stimulated dopamine efflux, focusing on Ca2+-dependent forms of PKC. Efflux of endogenous dopamine was measured in superfused rat striatal slices; dopamine was measured by high performance liquid chromatography. The non-selective classical PKC inhibitor Gö6976 inhibited amphetamine-stimulated dopamine efflux, whereas rottlerin, a specific inhibitor of PKCδ, had no effect. A highly specific PKCβ inhibitor, LY379196, blocked dopamine efflux that was stimulated by either amphetamine or the PKC activator, 12-O-tetradecanoylphorbol-13-acetate. None of the PKC inhibitors significantly altered [3H]dopamine uptake. PKCβI and PKCβII, but not PKCα or PKCγ, were co-immunoprecipitated from rat striatal membranes with the dopamine transporter (DAT). Conversely, antisera to PKCβI and PKCβII but not PKCα or PKCγ were able to co-immunoprecipitate DAT. Amphetamine-stimulated dopamine efflux was significantly enhanced in hDAT-HEK 293 cells transfected with PKCβII as compared with hDAT-HEK 293 cells alone, or hDAT-HEK 293 cells transfected with PKCα or PKCβI. These results suggest that classical PKCβII is physically associated with DAT and is important in maintaining the amphetamine-stimulated outward transport of dopamine in rat striatum. Evidence suggests that protein kinase C (PKC) and intracellular calcium are important for amphetamine-stimulated outward transport of dopamine in rat striatum. In this study, we examined the effect of select PKC isoforms on amphetamine-stimulated dopamine efflux, focusing on Ca2+-dependent forms of PKC. Efflux of endogenous dopamine was measured in superfused rat striatal slices; dopamine was measured by high performance liquid chromatography. The non-selective classical PKC inhibitor Gö6976 inhibited amphetamine-stimulated dopamine efflux, whereas rottlerin, a specific inhibitor of PKCδ, had no effect. A highly specific PKCβ inhibitor, LY379196, blocked dopamine efflux that was stimulated by either amphetamine or the PKC activator, 12-O-tetradecanoylphorbol-13-acetate. None of the PKC inhibitors significantly altered [3H]dopamine uptake. PKCβI and PKCβII, but not PKCα or PKCγ, were co-immunoprecipitated from rat striatal membranes with the dopamine transporter (DAT). Conversely, antisera to PKCβI and PKCβII but not PKCα or PKCγ were able to co-immunoprecipitate DAT. Amphetamine-stimulated dopamine efflux was significantly enhanced in hDAT-HEK 293 cells transfected with PKCβII as compared with hDAT-HEK 293 cells alone, or hDAT-HEK 293 cells transfected with PKCα or PKCβI. These results suggest that classical PKCβII is physically associated with DAT and is important in maintaining the amphetamine-stimulated outward transport of dopamine in rat striatum. The plasmalemmal dopamine transporter (DAT) 1The abbreviations used are: DAT, dopamine transporter; PKC, protein kinase C; hDAT, human DAT; hDAT-HEK-293, human DAT-transfected human embryonic kidney cells; KRB, Kreb's-Ringer buffer; KRH, Krebs-Ringer Hepes buffer; TPA, 12-O-tetradecanoylphorbol-13-acetate. is a pre-synaptic protein that belongs to the SLC6 family of Na+/Cl–-dependent neurotransmitter transporters. It is the primary regulator of the duration and strength of the dopamine signal in the synapse (1Giros B. Jaber M. Jones S.R. Wightman R.M. Caron M.G. Nature. 1996; 379: 606-612Crossref PubMed Scopus (2076) Google Scholar). DAT binds extracellular dopamine, Na+, and Cl– and transports them intracellularly, clearing dopamine from the synaptic cleft. The transporter is also the site of action of psychostimulant drugs such as amphetamine, which is a DAT substrate. Following its transport into the synaptic terminal, amphetamine stimulates a reversal of the transporter eliciting dopamine efflux. DAT is characterized as having 12 transmembrane segments and a large second extracellular loop with the N- and C-terminals located intracellularly. There are also putative phosphorylation sites for protein kinase C (PKC), protein kinase A, and Ca2+/calmodulin-stimulated protein kinase II (2Giros B. Caron M.G. Trends Pharmacol. Sci. 1993; 14: 43-49Abstract Full Text PDF PubMed Scopus (490) Google Scholar). PKC has been implicated in various aspects of DAT function and regulation, such as trafficking (3Melikian H.E. Buckley K.M. J. Neurosci. 1999; 19: 7699-7710Crossref PubMed Google Scholar), transport activity (4Doolen S. Zahniser N.R. FEBS Lett. 2002; 516: 187-190Crossref PubMed Scopus (29) Google Scholar, 5Daniels G.M. Amara S.G. J. Biol. Chem. 1999; 274: 35794-35801Abstract Full Text Full Text PDF PubMed Scopus (292) Google Scholar), and direct phosphorylation (Refs. 6Foster J.D. Pananusorn B. Vaughan R.A. J. Biol. Chem. 2002; 277: 25178-25186Abstract Full Text Full Text PDF PubMed Scopus (126) Google Scholar and 7Vaughan R.A. Huff R.A. Uhl G.R. Kuhar M.J. J. Biol. Chem. 1997; 272: 15541-15546Abstract Full Text Full Text PDF PubMed Scopus (220) Google Scholar, and see references in Ref. 8Zahniser N.R. Doolen S. Pharmacol. Ther. 2001; 92: 21-55Crossref PubMed Scopus (243) Google Scholar). Evidence strongly suggests that PKC activity is important for amphetamine-stimulated outward transport. General PKC inhibitors blocked dopamine efflux induced by amphetamine in rat striatal slices (9Kantor L. Gnegy M.E. J. Pharmacol. Exp. Ther. 1998; 284: 594-598Google Scholar). Accordingly, a PKC inhibitor blocked amphetamine-mediated locomotor behavior when injected directly into rat nucleus accumbens (10Browman K.E. Kantor L. Richardson S. Badiani A. Robinson T.E. Gnegy M.E. Brain Res. 1998; 814: 112-119Crossref PubMed Scopus (48) Google Scholar). The phorbol ester PKC activator, 12-O-tetradecanoylphorbol-13-acetate (TPA), mimicked the effect of amphetamine on the reverse transport by eliciting an efflux of dopamine that was cocaine-sensitive and independent of extracellular Ca2+ (11Cowell R.M. Kantor L. Hewlett G.H. Frey K.A. Gnegy M.E. Eur. J. Pharmacol. 2000; 389: 59-65Crossref PubMed Scopus (69) Google Scholar). Finally, a deletion of the N-terminal 22 amino acids of DAT, containing the distal N-terminal serines that are strongly considered to be substrate sites of PKC (6Foster J.D. Pananusorn B. Vaughan R.A. J. Biol. Chem. 2002; 277: 25178-25186Abstract Full Text Full Text PDF PubMed Scopus (126) Google Scholar), abrogated amphetamine-induced dopamine efflux from hDAT-HEK 293 cells (human embryonic kidney 293 cells) without altering inward transport (12Khoshbouei H. Sen N. Guptaroy B. Johnson L. Lund D. Gnegy M.E. Galli A. Javitch J.A. PLoS Biol. 2004; 2: E78Crossref PubMed Scopus (207) Google Scholar). Mutation of the distal N-terminal serines of DAT to alanines similarly abolished amphetamine-stimulated dopamine efflux, whereas mutation of the N-terminal serines to aspartates restored sensitivity of the reverse transport to amphetamine. The PKC isoform altering DAT-mediated dopamine efflux is not known, but it could be a Ca2+-sensitive isoform. Although amphetamine-induced dopamine efflux does not require extracellular Ca2+, it does require intracellular Ca2+ (13Kantor L. Hewlett G.H. Park Y.H. Richardson-Burns S.M. Mellon M.J. Gnegy M.E. J. Pharmacol. Exp. Ther. 2001; 297: 1016-1024PubMed Google Scholar, 14Gnegy M.E. Khoshbouei H. Berg K.A. Javitch J.A. Clarke W.P. Zhang M. Galli A. Mol. Pharmacol. 2004; 66: 137-143Crossref PubMed Scopus (79) Google Scholar). Moreover, amphetamine, acting at DAT, increases intracellular Ca2+ from thapsigargin-sensitive stores (14Gnegy M.E. Khoshbouei H. Berg K.A. Javitch J.A. Clarke W.P. Zhang M. Galli A. Mol. Pharmacol. 2004; 66: 137-143Crossref PubMed Scopus (79) Google Scholar, 15Kantor L. Zhang M. Guptaroy B. Park Y.H. Gnegy M.E. J. Pharmacol. Exp. Ther. 2004; 311: 1044-1051Crossref PubMed Scopus (22) Google Scholar). There are three major classes of PKC isoforms: classical, non-classical, and atypical. The classical PKC isoforms (α, β1, β2, γ) are Ca2+- and diacylglycerol-dependent. The non-classical PKC isoforms (δ, ϵ, η, θ) are Ca2+-independent but diacylglycerol-dependent. The atypical PKC isoforms (ζ and ι/λ) are insensitive to Ca2+ and diacylglycerol. There is a limited number of inhibitors that can distinguish among the different isoforms. Gö6976 is a potent inhibitor of the classical PKC isoforms (16Martiny-Baron G. Kazanietz M.G. Mischak H. Blumberg P.M. Kochs G. Hug H. Marme D. Schachtele C. J. Biol. Chem. 1993; 268: 9194-9197Abstract Full Text PDF PubMed Google Scholar). LY379196, structurally similar to LY333531, is a highly selective inhibitor of the PKCβ isoform (17Jirousek M.R. Gillig J.R. Gonzalez C.M. Heath W.F. McDonald III, J.H. Neel D.A. Rito C.J. Singh U. Stramm L.E. Melikian-Badalian A. Baevsky M. Ballas L.M. Hall S.E. Winneroski L.L. Faul M.M. J. Med. Chem. 1996; 39: 2664-2671Crossref PubMed Scopus (326) Google Scholar), and rottlerin is a selective inhibitor of the PKCδ isoform (18Gschwendt M. Muller H.J. Kielbassa K. Zang R. Kittstein W. Rincke G. Marks F. Biochem. Biophys. Res. Commun. 1994; 199: 93-98Crossref PubMed Scopus (762) Google Scholar). Using these selective inhibitors in combination with co-immunoprecipitation studies and the transfection of PKC isozymes into hDAT human embryonic kidney 293 (hDAT-HEK 293) cells, we investigated the role of two classical PKC isozymes in regulating outward transport of dopamine by amphetamine and their association with DAT. Materials—Gö6976 and rottlerin were purchased from Calbiochem. d-Amphetamine, dopamine, and TPA were purchased from Sigma. LY379196 was a generous gift from Lilly. hDAT cDNA was a gift from Dr. Z. B. Pristupa, Centre for Addiction and Mental Health, Department of Psychiatry, University of Toronto, Canada; cDNAs for PKCα, PKCβI, and PKCβII were generous gifts from Dr. Stephen Ferguson, John P. Robarts Research Institute, London, Ontario, Canada. Amphetamine- or TPA-mediated Dopamine Efflux—Rat striata from female Sprague-Dawley rats (175–200 g) were sliced, weighed, and placed in chambers of a Brandel perfusion apparatus (Brandel SF-12, Gaithersburg, MD) onto Whatman GF/B filter disks. The chambers were perfused at 37 °C with oxygenated Krebs-Ringer buffer (KRB, 145 mm NaCl, 2.7 mm KCl, 1.2 mm KH2PO4, 1.2 mm CaCl2, 1.0 mm MgCl2, 10 mm glucose, 0.255 mm ascorbic acid, 24.9 mm NaHCO3, 50 μm pargyline, and 1 mm tropolone) for 40 min at a rate of 100 μl/min. The slices were then perfused in the presence or absence of the drugs (130 nm Gö6976, 10 μm rottlerin, or 100 nm LY379196) for 40 min prior to the addition of a 2.5-min bolus of 3 μm amphetamine or 300 nm TPA. Eluates (500 μl, 5-min fractions) were collected into vials containing 25 μl of internal standard solution (0.05 m HClO4, 4.55 mm dihydroxybenzylamine (internal standard), 1 m metabisulfate and 0.1 m EDTA). Dopamine was measured by high performance liquid chromatography with electrochemical detection. To measure dopamine efflux from hDAT-HEK 293 cells, cells cultured in 100-mm plates were incubated with 15 μm unlabeled dopamine for 30 min at 37 °C. Following the incubation, cells were washed two times with a Krebs-Ringer-Hepes (KRH) buffer (25 mm Hepes, pH 7.4, 125 mm NaCl, 4.8 mm KCl, 1.2 mm KH2PO4, 1.3 mm CaCl2, 1.2 mm MgSO4, 5.6 mm glucose, and 1 mm tropolone) and resuspended in 300 μl of KRH. The cells (200 μl) were placed in the Brandel superperfusion apparatus and washed for 30 min at room temperature before the addition of 3 μm amphetamine for 2.5 min. Dopamine was measured in the as striata were in 10 of buffer containing m 1 mm and a of inhibitors pH were at for 10 min to The was and the was resuspended in buffer and The were for 30 min at at °C. The was and the was resuspended in for [3H]dopamine were resuspended in 1.2 of and incubated in the presence or absence of drugs (130 nm Gö6976, 10 μm rottlerin, or 100 nm LY379196) for 40 min at 37 °C. was for 3 min. was with of were on and washed with of were and then in a liquid was 10 μm of the DAT inhibitor of 25 nm [3H]dopamine was measured similarly in hDAT-HEK 293 cells, which were resuspended in from the striatal were in buffer mm pH mm NaCl, and by for 1 at was by at were incubated with either from or of In was with 40 of the of for at °C. were to protein for 30 min and at with buffer and with 10 pH 7.4, and NaCl, were from protein with buffer and on and of 293 were in with at 37 °C and the cells were at on plates and were transfected with of hDAT cDNA in a in to the 12 the solution was and was were for and was to select for a transfected of PKC isoforms PKCβII, or were by transfection of 15 of cDNAs for PKCα, PKCβI, or PKCβII in PKCβI in protein Zhang J. Caron M.G. J. Biol. Chem. 1998; Full Text Full Text PDF PubMed Scopus Google in into hDAT-HEK cells as The were by in the presence of and for PKCα, PKCβI, and of hDAT and PKCα, PKCβI, and PKCβII was by were by on a and to a were blocked with mm pH 7.4, mm NaCl, containing and incubated for 1 in containing DAT at by Dr. University of at PKCβI or PKCβII at in were with and incubated for 1 with at a were washed with and with of to of rat DAT was at the University of was in at the for University of the of C. R. Vaughan R.A. S. Uhl G.R. Kuhar M.J. J. PubMed Scopus Google Scholar). The was the from to the of of hDAT-HEK 293 cells and cells hDAT PKCα, hDAT PKCβI, and hDAT PKCβII was by the with the by C. C.J. Javitch J.A. U. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google with the were to on 100-mm from the plates by and resuspended in pH before with at °C for min in The was and the were as by C. C.J. Javitch J.A. U. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google that of were incubated with The were by and a of were at by detection. of the was by the was among three or was of with such as the effect of a on amphetamine-stimulated dopamine efflux, were a of PKC on Amphetamine-stimulated Dopamine effect of the selective PKC inhibitors Gö6976, LY379196, and rottlerin on amphetamine-stimulated dopamine efflux from rat striatal slices was The classical PKC inhibitor Gö6976 at nm of (16Martiny-Baron G. Kazanietz M.G. Mischak H. Blumberg P.M. Kochs G. Hug H. Marme D. Schachtele C. J. Biol. Chem. 1993; 268: 9194-9197Abstract Full Text PDF PubMed Google Scholar), the of dopamine efflux in to amphetamine from of protein in the absence of the to of protein in the presence of the of amphetamine-stimulated dopamine efflux was also nm Gö6976, a to the PKC LY379196 is a highly selective inhibitor of PKCβ of 30 (17Jirousek M.R. Gillig J.R. Gonzalez C.M. Heath W.F. McDonald III, J.H. Neel D.A. Rito C.J. Singh U. Stramm L.E. Melikian-Badalian A. Baevsky M. Ballas L.M. Hall S.E. Winneroski L.L. Faul M.M. J. Med. Chem. 1996; 39: 2664-2671Crossref PubMed Scopus (326) Google we used this to the role of a classical PKC LY379196 a classical and non-classical In the presence of 100 nm LY379196, amphetamine-stimulated dopamine efflux was from of protein in the absence of to of protein The for dopamine efflux in the presence of LY379196 was not different from To the activity of a PKC, we used rottlerin, a selective PKCδ inhibitor of (18Gschwendt M. Muller H.J. Kielbassa K. Zang R. Kittstein W. Rincke G. Marks F. Biochem. Biophys. Res. Commun. 1994; 199: 93-98Crossref PubMed Scopus (762) Google Scholar). at 10 not amphetamine-mediated dopamine efflux in rat striatal slices a but effect on efflux. of the PKC on the PKC inhibitors inward transport of [3H]dopamine was were incubated with Gö6976, LY379196, and rottlerin at 37 °C for 40 min prior to the addition of [3H]dopamine to the in the efflux of the with nm Gö6976, 100 nm LY379196, or 10 μm rottlerin had no effect on [3H]dopamine as compared with of LY379196 on TPA-mediated Dopamine that of PKC with TPA mimicked the of amphetamine to dopamine efflux from rat striatal slices (9Kantor L. Gnegy M.E. J. Pharmacol. Exp. Ther. 1998; 284: 594-598Google Scholar, R.M. Kantor L. Hewlett G.H. Frey K.A. Gnegy M.E. Eur. J. Pharmacol. 2000; 389: 59-65Crossref PubMed Scopus (69) Google Scholar). TPA and amphetamine are the TPA-mediated dopamine efflux be inhibited by the selective PKCβ inhibitor, LY379196, in rat striatal in LY379196 TPA-mediated dopamine efflux from of dopamine to 1.3 dopamine PKCβI and PKCβII with DAT in the of co-immunoprecipitation to PKCβ was to DAT. DAT was a in The DAT were for PKCα, PKCβI, PKCβII, and To that the co-immunoprecipitation was specific for DAT, we the with the DAT N-terminal The in that PKCβI and PKCβII co-immunoprecipitated with DAT in the absence but not the presence of the the PKCα PKCγ co-immunoprecipitated with DAT. in forms of PKC were in the To the was antisera to the PKC and the presence of DAT in the was with in DAT co-immunoprecipitated with PKCβI and PKCβII but not with PKCα or these results with two different PKCβII also co-immunoprecipitation of hDAT and PKCβII in cells with either or not we in the of PKCβ isozymes that with DAT. with two to three different to PKCβI and PKCβII, and the results were we that the of the could on the of the for or of striatal with either or but not or DAT. striatal were in buffer and as 50 of protein were incubated with either or The protein from the was in and of protein for in and of protein in in were by for PKCβII Amphetamine-stimulated Dopamine PKCβI and PKCβII co-immunoprecipitated with DAT, or of the isozymes could be for amphetamine-stimulated dopamine efflux. LY379196 does not PKCβI and To distinguish the two PKCβ isozymes we hDAT-HEK 293 PKCα, PKCβI, or in hDAT-HEK 293 cells PKCα and PKCβI but the high of their PKC The cells were with 15 μm dopamine for 30 min to measure dopamine efflux by The dopamine efflux in to 3 μm amphetamine was similar in the hDAT-HEK 293 cells, the 293 cells, and the 293 cells the dopamine efflux in to amphetamine was in the 293 cells in the The of amphetamine in the 293 cells was whereas that in the was at hDAT-HEK 293 cells, 293 cells, and 1.3 293 cells, of of in the hDAT-HEK 293 cells and the 293 containing PKCα, PKCβI, and PKCβII were and The of dopamine in the cells at the of the perfusion in of in hDAT-HEK 293 cells; 1.2 in 293 cells; in 293 cells; and in 293 cells of of cells were used in the in amphetamine-stimulated dopamine efflux in the 293 cells was not to a significantly of dopamine in the To that transport of amphetamine into the different was not altered by the transfection of the PKC we measured of [3H]dopamine in the of 25 nm [3H]dopamine in of in hDAT-HEK 293 cells; in 293 cells; in 293 cells; and 0.1 in 293 cells of To the of hDAT on the of the was altered by transfection of the PKC of DAT was in the in amphetamine-stimulated dopamine efflux in 293 cells is also not to the presence of significantly of hDAT as compared with the that classical PKC isozymes the of action of amphetamine in eliciting outward transport of the that PKCβ PKCβII, amphetamine-induced outward transport the dopamine transporter and that PKCβ isoforms are associated with DAT. The of DAT by PKC is that with TPA efflux of dopamine that is blocked by but independent of extracellular Ca2+ (11Cowell R.M. Kantor L. Hewlett G.H. Frey K.A. Gnegy M.E. Eur. J. Pharmacol. 2000; 389: 59-65Crossref PubMed Scopus (69) Google Scholar). the with TPA inward trafficking of DAT N.R. Doolen S. Pharmacol. Ther. 2001; 92: 21-55Crossref PubMed Scopus (243) Google Scholar). The of DAT also classical Gö6976 of transport (4Doolen S. Zahniser N.R. FEBS Lett. 2002; 516: 187-190Crossref PubMed Scopus (29) Google Scholar), an effect to the enhanced of DAT. TPA can PKC and are of classical PKC isozymes and different classical PKC isozymes could be these two of by different of PKC and different PKC isozymes has been with TPA increases activity of the transporter in whereas inward transport J. Biol. PubMed Scopus Google Scholar). II on transport in the and to be by PKC J. Biol. PubMed Scopus Google Scholar, S. C. M. H. A. G. J. 14: PubMed Scopus Google Scholar, J. Pharmacol. Exp. Ther. 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M. 2000; 39: PubMed Scopus Google Scholar). of TPA-mediated dopamine efflux by the highly specific PKCβ inhibitor LY379196 strongly suggests that PKCβ is the specific isoform that is for DAT-mediated efflux of The of PKCα and PKCγ on amphetamine-mediated dopamine efflux were not investigated of the of selective inhibitors for these isoforms. the co-immunoprecipitation studies and the studies with of PKCα strongly suggest that these isozymes to DAT DAT-mediated dopamine efflux. classical PKC isoforms are to in the dopamine of Sprague-Dawley the used in these studies J.D. 1998; PubMed Scopus Google Scholar), the was PKCβI. that the PKCβI and PKCβII isozymes co-immunoprecipitated with DAT that in the not directly to DAT or is an The for C has been to to the of DAT Res. 2004; PubMed Scopus Google Scholar). PKCβII with which the and activity of PKCβII D. Z. L. A. A. J. Biol. Chem. 1999; 274: Full Text Full Text PDF PubMed Scopus Google Scholar). The 1 J. R. J. Biol. PubMed Scopus Google Scholar), has been to a with the in the C of DAT H. K.M. J. Caron M.G. 2001; Full Text Full Text PDF PubMed Scopus Google Scholar). It is at this PKCα role in the of 1 to the number of H. K.M. J. Caron M.G. 2001; Full Text Full Text PDF PubMed Scopus Google Scholar). a of PKCα, and DAT not been or PKCα when DAT is PKCα with the transporter PKC in cells and rat and this with the of Robinson J. Neurosci. PubMed Google Scholar). suggests that PKCβ is associated with DAT studies protein and DAT as as the and S. S. Vaughan R.A. J. Neurosci. 2000; PubMed Google Scholar). it that DAT and can in in the that and signal hDAT-HEK 293 cells transfected with PKC isozymes suggest that the PKCβII is important for amphetamine-stimulated efflux of dopamine DAT. of hDAT-HEK 293 cells or not significantly the amphetamine-stimulated dopamine efflux from that of 293 Although could that of PKCα not a effect the 293 cells of PKCα, that not be for PKCβI. The 293 cells had of PKCβI and no with PKCβII significantly the dopamine efflux in to amphetamine. The by which PKCβ amphetamine-stimulated dopamine efflux is PKCβ could be directly DAT and altering efflux. A phosphorylation of DAT has been at sites that to be the distal N-terminal serines (6Foster J.D. Pananusorn B. Vaughan R.A. J. Biol. Chem. 2002; 277: 25178-25186Abstract Full Text Full Text PDF PubMed Scopus (126) Google Scholar). we that phosphorylation of the N-terminal serines in DAT is for amphetamine-induced dopamine efflux (12Khoshbouei H. Sen N. Guptaroy B. Johnson L. Lund D. Gnegy M.E. Galli A. Javitch J.A. PLoS Biol. 2004; 2: E78Crossref PubMed Scopus (207) Google Scholar), but [3H]dopamine is not is that PKCβ could in the transporter or of of DAT. PKCβ increases the of transporter 1 and R. J. Pharmacol. 2002; PubMed Scopus Google Scholar, R. FEBS Lett. 1999; PubMed Scopus Google Scholar). PKCβII, as to PKCβI, has been to with the D. S. J. Biol. 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Gnegy M.E. J. Pharmacol. Exp. Ther. 1996; Google Scholar). There is that is by PKCβII, as compared with PKCα or PKCγ R.M. A. Biochem. Biophys. Res. Commun. PubMed Scopus Google Scholar), that the amphetamine-stimulated in phosphorylation could be by of which PKC isoforms are in regulating amphetamine-mediated dopamine efflux we can the by which amphetamine to reverse the transport of These studies that PKCβII is with DAT and is important in the reverse transport of dopamine DAT. Stephen for the of this also Zhang for and

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