Stimulation of L-type Ca(2+) channels by increase of intracellular InsP3 in rat retinal pigment epithelial cells.
The purpose of this study was to investigate the role of voltage-dependent L-type Ca(2+)channels in intracellular Ca(2+)signaling of the retinal pigment epithelium (RPE). Patch-clamp techniques in conjunction with measurements of the intracellular free Ca(2+)using the Ca(2+)-sensitive fluorescence dye fura-2 were performed using cultured rat RPE cells. Intracellular application of inositol-1,4,5-trisphosphate (InsP3; 10 microM) via the patch-pipette during the whole-cell configuration led to an increase in the intracellular free Ca(2+)([Ca(2+)](i)). This effect could be reduced by the L-type Ca(2+)channel blocker nifedipine (2 microM). At the moment of the maximal rise in [Ca(2+)](i)L-type currents displayed an increase in the current density and shifts in the activation curve and of the steady-state inactivation. Comparable changes of L-type channel activity could be observed by induction of capacitative Ca(2+)entry, a maneuver to release Ca(2+)from intracellular Ca(2+)stores independently from InsP3. The increase in L-type Ca(2+)channel activity and [Ca(2+)](i)by intracellular application of InsP3 or induction of capacitative Ca(2+)entry could be inhibited by blocking tyrosine kinase activity using genistein (5 microM) or tyrphostin 51 (10 microM). It is concluded that L-type Ca(2+)channels are involved in the Ca(2+)/InsP3 second messenger system by generating an influx of extracellular Ca(2+)into the cell. This is enabled by depletion of cytosolic Ca(2+)stores and tyrosine kinase-dependent activation of L-type channels.
Mergler
Stefan
S
Universitätsklinikum Charité der Humboldt-Universität zu Berlin, Medizinische Klinik, Hepatologie und Gastroenterologie, Campus Virchow Klinikum, Augustenburger Platz 1, Berlin, 13353, Germany.
Strauss
Olaf
O
D000818:Animals; D002118:Calcium / Q000378:metabolism; D020746:Calcium Channels, L-Type / Q000187:drug effects / Q000502:physiology*; D002478:Cells, Cultured; D003600:Cytosol / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology; D019833:Genistein / Q000494:pharmacology; D015544:Inositol 1,4,5-Trisphosphate / Q000494:pharmacology*; D018408:Patch-Clamp Techniques; D010857:Pigment Epithelium of Eye / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D011505:Protein-Tyrosine Kinases / Q000037:antagonists & inhibitors / Q000502:physiology; D051381:Rats; D015290:Second Messenger Systems / Q000502:physiology; D020032:Tyrphostins / Q000494:pharmacology
D020746:Calcium Channels, L-Type; D004791:Enzyme Inhibitors; D020032:Tyrphostins; C118515:tyrphostin 51; D015544:Inositol 1,4,5-Trisphosphate; D019833:Genistein; D011505:Protein-Tyrosine Kinases; D002118:Calcium