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INHIBITION OF CALCIUM‐INDEPENDENT LUMINAL UPTAKE OF L‐DOPA BY CALMODULIN ANTAGONISTS IN IMMORTALIZED RAT CAPILLARY CEREBRAL ENDOTHELIAL CELLS
Author(s) -
SampaioMaia B.,
SoaresDaSilva P.
Publication year - 2001
Publication title -
cell biology international
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.932
H-Index - 77
eISSN - 1095-8355
pISSN - 1065-6995
DOI - 10.1006/cbir.2000.0639
Subject(s) - chelerythrine , protein kinase c , calmodulin , trifluoperazine , staurosporine , protein kinase a , chemistry , forskolin , biochemistry , zaprinast , kinase , biology , microbiology and biotechnology , in vitro , enzyme , phosphodiesterase
The present study examined the involvement of protein kinase A (PKA), protein kinase G (PKG), protein kinase C (PKC), protein tyrosine kinase (PTK) and Ca 2+ /calmodulin mediated pathways on the luminal uptake of L‐DOPA through the L‐type amino acid transporter in immortalized rat capillary cerebral endothelial (REB‐4) cells. Non‐linear analysis of the saturation curve for L‐DOPA revealed a K m value (in μM) of 71±9 and a V max value of 17±1 (in nmol mg protein/6min). L‐DOPA uptake at the luminal cell border was a sodium‐independent process and insensitive to N‐(methylamino)‐isobutyric acid (MeAIB, 1m m ), but sensitive to 2‐aminobicyclo(2,2,1)‐heptane‐2‐carboxylic acid (BHC, IC 50 =140μ m ). The Ca 2+ /calmodulin inhibitors calmidazolium and trifluoperazine inhibited L‐DOPA (2.5μ m ) uptake with IC 50 s of 23 and 33μ m , respectively. The inhibitory effect of BHC on the accumulation of L‐DOPA was of the competitive type, whereas that of calmidazolium and trifluoperazine was of the non‐competitive type. Modulators of PKA (cyclic AMP, forskolin, isobutylmethylxanthine and cholera toxin), PKG (cyclic GMP, zaprinast, LY 83583 and sodium nitroprusside), PKC (phorbol 12,13‐dibutyrate, staurosporine and chelerythrine) and PTK (genistein and tyrphostin 25) failed to affect the accumulation of a non‐saturating (2.5μ m ) concentration of L‐DOPA. It is concluded that L‐DOPA uptake in RBE‐4 cells is promoted through the L‐type amino acid transporter and appears to be under the control of calmodulin mediated pathways.