Role of Ca(2+) and protein kinase C in the serotonin (5-HT) transport in human platelets.
Accumulation of serotonin (5-HT) into human platelets was not affected by the presence of the extra-cellular calcium chelator EGTA, while decreased by platelet incubation with the membrane permeant chelator BAPTA-AM. Serotonin uptake also diminished upon platelet exposure to EGTA/thapsigargin or EGTA/ionomycin which increased the cytosolic [Ca(2+)] to levels lower than those inducing secretion of dense granules. The latter inhibition depended in part on changes of intra-granular pH, since the accumulation of acridine orange, which is driven into the dense granules by the intra-granular acid pH gradient, was slightly decreased in the presence of EGTA/thapsigargin. These compounds also inhibited the 5-HT uptake in platelets pre-incubated with reserpine and bafilomycin that prevent 5-HT from entering into the dense granules. Inhibitors of protease, protein phosphatase, Na(+)/H(+) exchanger or ciclo-oxygenase activities did not modify the serotonin accumulation. Addition of EGTA/thapsigargin to reserpine-treated, [(14)C]5-HT-loaded, platelets caused an imipramine-insensitive release of labelled serotonin. This release was reduced by both BAPTA-AM or protein kinase C inhibitor bisindoylmaleimide (GF). The latter compound, either alone or together with EGTA/thapsigargin, inhibited the 5-HT accumulation in reserpine-treated platelets. It is concluded that both cytosolic [Ca(2+)] and protein kinase C are involved in the regulation of the plasma membrane 5-HT transport.
Turetta
L
L
Department of Biological Chemistry, C.N.R. Unit for the Study of Biomembranes, University of Padova, Padova, Italy.
Bazzan
E
E
Bertagno
K
K
Musacchio
E
E
Deana
R
R
D000165:Acridine Orange / Q000494:pharmacology; D001792:Blood Platelets / Q000187:drug effects* / Q000201:enzymology*; D002118:Calcium / Q000378:metabolism*; D020013:Calcium Signaling / Q000187:drug effects / Q000502:physiology; D002352:Carrier Proteins / Q000187:drug effects / Q000378:metabolism*; D002462:Cell Membrane / Q000187:drug effects / Q000201:enzymology*; D002478:Cells, Cultured; D002614:Chelating Agents / Q000494:pharmacology; D003600:Cytosol / Q000187:drug effects / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006801:Humans; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000378:metabolism*; D011500:Protein Synthesis Inhibitors / Q000494:pharmacology; D012110:Reserpine / Q000494:pharmacology; D012701:Serotonin / Q000378:metabolism*; D050486:Serotonin Plasma Membrane Transport Proteins
D002352:Carrier Proteins; D002614:Chelating Agents; D004791:Enzyme Inhibitors; D011500:Protein Synthesis Inhibitors; D050486:Serotonin Plasma Membrane Transport Proteins; D012701:Serotonin; D012110:Reserpine; D011493:Protein Kinase C; D000165:Acridine Orange; D002118:Calcium