Basal calcium entry in retinal pigment epithelial cells is mediated by TRPC channels.
OBJECTIVE
Ca(2+) is a major regulator of cell function. In the retinal pigment epithelium (RPE), intracellular free Ca(2+) concentration ([Ca(2+)](i)) is essential for the maintenance of normal retinal function. Therefore, accurate control of [Ca(2+)](i) is vital in these cells. Because Ca(2+) is permanently extruded from the cytosol, RPE cells need a basal Ca(2+) entry pathway that counteracts this Ca(2+) efflux. The purpose of this study was to identify the molecular basis of basal Ca(2+) entry into the RPE.


METHODS
[Ca(2+)](i) was measured using Fura-2-loaded ARPE-19 cells. The expression pattern of TRPC channels was investigated by RT-PCR with RNA extracted from ARPE-19 cells and freshly isolated RPE cells from human donor eyes.


RESULTS
In most cells, basal [Ca(2+)](i) is highly controlled by cell membranes that are only slightly permeable to Ca(2+) and by the activity of Ca(2+) pumps and transporters. The authors show here that RPE cells have a basal Ca(2+) conductance that is dose dependently blocked by La(3+). Basal [Ca(2+)](i) was also strongly reduced by the TRP channel blockers Gd(3+), Ni(2+), 2-APB, and SKF96365 and was insensitive to blockers of other Ca(2+) channels. In confirmation of this pharmacologic profile, RPE cells expressed TRPC1 and TRPC4 channels, as shown by RT-PCR experiments.


CONCLUSIONS
Ca(2+) is needed for several permanently occurring regulatory processes in RPE cells. The Ca(2+) influx pathway identified in this study is essential to define a resting basal [Ca(2+)](i). This resting [Ca(2+)](i) may contribute, for example, to basal cytokine secretion essential for the maintenance of normal retinal function.
Experimentelle Ophthalmologie, Klinik und Poliklinik für Augenheilkunde, Universitätsklinkikum Hamburg-Eppendorf, Hamburg, Germany. wimmers@uke.uni-hamburg.de
Sönke
S
Wimmers
Olaf
O
Strauss
D002118:Calcium / Q000378:metabolism*; D000252:Calcium-Transporting ATPases / Q000037:antagonists & inhibitors; D002460:Cell Line; D002462:Cell Membrane / Q000201:enzymology; D002463:Cell Membrane Permeability; D016257:Fura-2 / Q000378:metabolism; D006801:Humans; D010857:Pigment Epithelium of Eye / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D050052:TRPC Cation Channels / Q000502:physiology*; D014638:Vanadates / Q000494:pharmacology
D050052:TRPC Cation Channels; D014638:Vanadates; D000252:Calcium-Transporting ATPases; D002118:Calcium; D016257:Fura-2