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Tunde Molnar, Peter Barabas, Claudio Punzo, David Krizaj; Store-operated Calcium Entry Regulates Intracellular Calcium Homeostasis In Mouse Rod Photoreceptors. Invest. Ophthalmol. Vis. Sci. 2011;52(14):6581.
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To measure resting [Ca2+]i in mouse rods and to determine the properties of store-operated calcium entry (SOCE) within different rod compartments. We also assessed whether SOCE in mouse photoreceptors is mediated by TRPC1 and/or TRPC3 channels which can function as store-operated channels in non-excitable cells.
Optical imaging was performed in dissociated mouse rods prepared from wild type and Trpc1/Trpc3 (TRPC1/3-/-) double knockout mouse retinas. Cells were loaded with fura-2 AM, excited at 340/380 nm and visualized using high-resolution 14 bit cooled CCD cameras. Voltage-operated and store-operated signals were evoked with high KCl and prolonged depletion of intracellular Ca2+ stores within the endoplasmic reticulum (ER). RT-PCR and in situ hybridization were performed using primers for mouse SOC channel candidates.
Resting [Ca2+]i levels in wild type mouse rod somata were 85 ± 15 nM with a median concentration of 55 nM (N=449). Depolarization elevated perikaryal [Ca2+]i >400 nM, indicating the maintained excitability of dissociated rods. Depletion of ER stores in Ca2+-free saline supplemented with cyclopiazonic acid induced SOCE (371 ± 30 nM) manifested as sustained [Ca2+]i overshoots following the return to control Ca2+ -containing saline. Sustained divalent cation entry was visualized as La3+-, and 1-oleoyl-2-acetyl-glycerol (OAG)-sensitive quenching of Fura-2 by Mn2+. Mouse rods showed prominent Trpc1 and Trpc3 mRNA expression that was reduced in Pde6brd1 retinas. Genetic ablation of TRPC1 and TRPC3 channels had little effect on baseline [Ca2+]i (84 ± 7 nM) or SOCE (452 ± 29 nM) in cells isolated from knockout animals. However, analysis of distribution of [Ca2+]i in the TRPC1/3-/- cohort showed a significant decrease in number of rods with elevated (100-250 nM) [Ca2+]i levels.
We found that SOCE mediates significant Ca2+ influx in mouse rod photoreceptor cell bodies and synaptic terminals. This pathway was activated by low [Ca2+]i and depletion of ER stores, indicating that it may play a disproportionately prominent role under light-adapted conditions. TRPC1 and TRPC3 channels do not mediate rod SOCE but may contribute to Ca2+ overload in stressed rods.
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