ISSN:
1432-2013
Keywords:
Intracellular pH
;
K+ channel
;
NH4 +/NH3 Patch clamp
;
BCECF
Source:
Springer Online Journal Archives 1860-2000
Topics:
Medicine
Notes:
Abstract The K+ channels of the principal cells of rat cortical collecting duct (CCD) are pH sensitive in excised membranes. K+ secretion is decreased with increased H+ secretion during acidosis. We examined whether the pH sensitivity of these K+ channels is present also in the intact cell and thus could explain the coupling between K+ and H+ secretion. Membrane voltages (V m), whole-cell conductances (g c), and single-channel currents of K+ channels were recorded from freshly isolated CCD cells or isolated CCD segments with the patch-clamp method. Intracellular pH (pHi) was measured using the pH-sensitive fluorescent dye 2′-7′-bis(carboxyethyl)-5-6-carboxyfluorescein (BCECF). Acetate (20 mmol/l) had no effect on V m, g c, or the activity of the K+ channels in these cells. Acetate, however, acidified pHi slightly by 0.17±0.04 pH units (n=19). V m depolarized by 12±3 mV (n=26) and by 23±2 mV (n=66) and g c decreased by 26±5% (n=13) and by 55±5% (n=12) with 3–5 or 8–10% CO2, respectively. The same CO2 concentrations decreased pHi by 0.49±0.07 (n=15) and 0.73±0.11 pH units (n=12), respectively. Open probability (P o) of all four K+ channels in the intact rat CCD cells was reversibly inhibited by 8–10% CO2. pHi increased with the addition of 20 mmol/l NH4 +/NH3 by a maximum of 0.64±0.08 pH units (n=33) and acidified transiently by 0.37±0.05 pH units (n=33) upon NH4 +/NH3 removal. In the presence of NH4 +/NH3 V m depolarized by 16±2 mV (n=66) and g c decreased by 26±7% (n=16). The activity of all four K+ channels was also strongly inhibited in the presence of NH4 +/NH3. The effect of NH4 +/NH3 on V m and g c was markedly increased when the pH of the NH4 +/NH3-containing solution was set to 8.5 or 9.2. From these data we conclude that cellular acidification in rat CCD principal cells down-regulates K+ conductances, thus reduces K+ secretion by direct inhibition of K+ channel activity. This pH dependence is present in all four K+ channels of the rat CCD. The inhibition of K+ channels by NH4 +/NH3 is independent of changes in pHi and rather involves an effect of NH3.
Type of Medium:
Electronic Resource
URL:
http://dx.doi.org/10.1007/BF00374587
