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Regulation of intracellular ph in llc pk 1 cells by sodium hydrogen exchange


, : Regulation of intracellular ph in llc pk 1 cells by sodium hydrogen exchange. Journal of Membrane Biology 93(1): 33-42

Suspensions of LLC-PK1 cells (a continuous epitheliod cell line with renal characteristics) are examined for mechanisms of intracellular pH regulation using the fluorescent probe BCECF. Initial experiments determine suitable calibration procedures for use of the BCECF fluorescent signal. They also determine that the cell suspension contains cells which (after 4 hr in suspension) have Na+ and K+ gradients comparable to those of cells in monolayer culture. The steady-state intracellular pH (7.05 .+-. 0.01, n=5) of cells which have recovered in (pH 7.4) Na+-containing medium is not affected over several minutes by addition of 100 .mu.M amiloride or removal of extracellular Na+ .**GRAPHIC**. < 1mM. In contrast, when the cells recover from an acid load (caused by NH4 preincubation and removal), the recovery is largely Na+ dependent and is sensitive to 100 .mu.M amiloride. These results suggest that with resting pH near neutrality, both .**GRAPHIC**. and .**GRAPHIC**. exchange reactions are functionally inactive (compared to cellular buffering capacity). In contrast .**GRAPHIC**. exchange is activated by an increased cellular acid load. This activation may be observed directly either as a stimulation of net H+ efflux or net Na+ influx with decreasing intracellular pH. The extrapolation of this latter data suggests a "set point" of Na+/H+ exchange of approximately pH 7.0, consistent with the observed resting intracellular pH of approximately 7.05.

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