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R.P. Malchow, A.J. Molina, J. Bodily, K. Hammar, P.J. Smith; Calcium-dependent Regulation of Intracellular and Extracellular Hydrogen Ion Levels of Retinal Horizontal Cells . Invest. Ophthalmol. Vis. Sci. 2003;44(13):4176.
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Purpose: Changes in intracellular and extracellular hydrogen ion concentration alter a number of properties of retinal neurons, including the release of neurotransmitter and the function of many voltage-and neurotransmitter gated channels. We sought to examine the mechanisms in retinal horizontal cells that regulate internal and external pH. Methods: Intracellular pH measurements from enzymatically isolated horizontal cells of the skate (R. erinacea/ R. ocellata) were examined using the pH indicator dye BCECF. Extracellular pH fluxes were monitored using self-referencing pH-selective microelectrodes. Increases in calcium were induced directly in isolated cells by loading with the AM-ester analog of NP-EGTA and then causing the release of caged calcium with ultraviolet light. Results: The application of 20 uM of the glutamate analog kainic acid consistently induced an acidification of isolated skate retinal horizontal cells as measured using BCECF. This same compound produced a depression in the outward proton flux from these cells measured using self-referencing microelectrodes. These changes were dependent upon the presence of extracellular calcium. Depolarization of horizontal cells from -70 mV to 0 mV using the whole-cell voltage clamp technique also led to a depression in the outward proton flux from cells. Stimulation by ultraviolet light of cells preloaded with NP-EGTA induced an increase in free calcium as measured using the calcium indicator dye Oregon Green and a decrease in extracellular proton flux as measured using self-referencing pH-selective microelectrodes. Conclusions: The changes in intracellular and extracellular pH we have observed can best be explained by activation of a plasmalemma calcium/ H+ pump, which transports calcium out of the cell while taking in hydrogen ions from the external milieu. Kainate permits the influx of calcium ions through ionotropic glutamate receptors, while depolarization of the cell leads to influx of calcium through activation of voltage-gated calcium channels; both procedures will lead to activation of the Ca/H+ ATPase and consequent acidification of the interior of the cell and alkalinization of the external milieu near the plasma membrane of the horizontal cells.
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