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S. Wimmers, O. Strauss; Basal Calcium Entry in Retinal Pigment Epithelial Cells. Invest. Ophthalmol. Vis. Sci. 2007;48(13):2512.
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The retinal pigment epithelium (RPE) is involved in a variety of processes that help to maintain normal retinal function. Many of these tasks depend on intracellular free calcium ([Ca2+]i), for example photoreceptor phagocytosis, growth factor secretion, transepithelial transport mechanisms and cell differentiation. All these processes need extra- or intracellular signals to initiate an increase in [Ca2+]i. While there is some evidence that voltage-gated Ca2+ channels and purinergic receptors are involved in these Ca2+ fluxes nothing is known about the basal Ca2+ entry under resting conditions. The purpose of this study was to identify the molecular identity of Ca2+ channels involved in basal Ca2+ entry in RPE cells.
Intracellular Ca2+ concentration measurements were performed with ARPE-19 cells using FURA-2 as fluorescent Ca2+ indicator. The data obtained with the RPE cell line were confirmed with human RPE primary cultures. Additionally, RT PCR experiments were made with mRNA extracted from freshly isolated RPE cells and from ARPE-19 cells with oligonucleotides specific for different Ca2+ channels.
ARPE-19 cells had an resting [Ca2+]i of 106.4 ± 16.1 nM. This resting concentration was not changed by the application of the voltage-gated Ca2+ channel blocker nifedipin (10 µM). Gadolinium (100 µM); lanthanum (100 µM) and nickel (2 mM) reduced the [Ca2+]i by 73, 76 and 98%, respectively. Additionally, the blocker of store-operated Ca2+ entry 2-aminoethoxydiphenyl borate (75 µM) reduced [Ca2+]i by 69%. As these blockers are known to block Ca2+ conducting channels of the TRPC family we performed RT PCR experiments and found TRPC1 and TRPC4 to be expressed in ARPE-19 cells. In freshly isolated RPE cells TRPC7 was expressed in addition to TRPC1 and TRPC4.
Our data show that the basal entry of Ca2+ in RPE cells is mainly driven by TRPC1 and TRPC4 channels. Their basal open probability provides the basal [Ca2+]i needed for the function of diverse Ca2+-dependent processes in RPE cells.
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