Protein kinase C activation inhibits glutamate transport by endothelial cells.
The role of protein kinase C (PKC) in regulating endothelial cell glutamate transport was investigated. Glutamate transport studies were performed in confluent human umbilical vein endothelial cells which were treated with the phorbol ester 12-myristate 13-acetate (TPA, 0-1000 nM), a compound which directly activates PKC. TPA inhibited Na(+)-independent System xAG- glutamate transport by 70% but only slightly reduced Na(+)-dependent activity. The TPA-mediated reduction in transport activity was dose-dependent, beginning at 5 min and lasting for at least 24 hr. TPA inhibition of glutamate transport had two distinctive phases: an acute phase (< 1 hr, not affected by either cycloheximide or actinomycin D) in which TPA decreased System xAG- glutamate transporter affinity (TPA Km = 522 +/- 25 microM vs control Km = 329 +/- 85 microM, P < 0.01) but did not alter transporter capacity (TPA Vmax = 4426 +/- 230 pmole/mg/min vs control Vmax = 4535 +/- 750 pmole/mg/min, P = NS) and a chronic phase (4-24 hr) in which TPA inhibition of glutamate transport was due to a reduced transporter capacity (Vmax = 2895 +/- 570 pmole/mg/min) without altering transporter affinity (Km = 370 +/- 60 microM glutamate) and was abrogated by cycloheximide or actinomycin D. The protein kinase C inhibitor chelerythrine chloride abrogated TPA's inhibition effect in both the acute and chronic phases. These data indicate that protein kinase C activation decreases glutamate transport in human umbilical vein endothelial cells via protein synthesis dependent and independent mechanisms.
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Division of Surgical Oncology, Massachusetts General Hospital, Harvard Medical School, Boston 02114, USA.
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Souba
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D001692:Biological Transport / Q000187:drug effects; D002478:Cells, Cultured; D004730:Endothelium, Vascular / Q000378:metabolism*; D004789:Enzyme Activation; D018698:Glutamic Acid / Q000378:metabolism*; D005973:Glutamine / Q000378:metabolism; D006801:Humans; D011493:Protein Kinase C / Q000502:physiology*; D012964:Sodium / Q000378:metabolism; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology
D005973:Glutamine; D018698:Glutamic Acid; D012964:Sodium; D011493:Protein Kinase C; D013755:Tetradecanoylphorbol Acetate