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First published online September 19, 2008
Journal of Experimental Biology 211, 3167-3173 (2008)
Published by The Company of Biologists 2008
doi: 10.1242/jeb.019646

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Effects of [Ca²⁺]_i and pH on epithelial Na⁺ channel activity of cultured mouse cortical collecting ducts

Yuchun Gu

Department of Physiology, University of Birmingham, The Medical School, Edgbaston, B15 2TT, UK

View larger version (30K): [in this window] [in a new window]   Fig. 1. Epithelial Na+ channel (ENaC) currents revealed by single channel recording from cultured M1 cells. (A) In a cell-attached recording, inward currents were recorded when the pipette voltage (Vp) was held at +20 mV and +40 mV. (B) In an inside-out recording, the single channel currents were detected when the pipette voltage was held at +100 mV and +60 mV. (C) In an outside-out recording, the currents were monitored when the pipette voltage was held at –60 mV. Bath application of 5 µmol l–1 amiloride almost abolished the ENaC currents. The currents were reversed when amiloride was washed off. c indicates the closing level.

View larger version (12K): [in this window] [in a new window]   Fig. 2. Effect of elevation of [Ca2+]i on ENaC activity. The top panel shows single channel currents were detected when the pipette voltage was held at +20 mV, in a cell-attached recording. After the cell was incubated with 1 µmol l–1 thapsigargin (TG) for 5 min, ENaC activity were reduced as shown in the lower trace.

View larger version (30K): [in this window] [in a new window]   Fig. 3. Direct effects of [Ca2+]i on ENaC activity. In an inside-out recording, 500 nmol l–1 Ca2+ led to maximum ENaC open probability (Po) and even 5 µmol l–1 Ca2+ exerted significant inhibition on ENaC Po. The inhibitory effect of Ca2+ on ENaC activity depended on the Ca2+ concentration. In the lower panels selected currents are shown on an expanded time scale.

View larger version (34K): [in this window] [in a new window]   Fig. 4. Effect of high pHi on ENaC activity. In inside-out recordings, with pHi at 7.6 (A) and 8 (B), channel activity was significantly enhanced. This effect was reversed when pHi was returned to 7.2. The lower traces in each panel show the currents on an expanded time scale, as indicated by the bars.

View larger version (22K): [in this window] [in a new window]   Fig. 5. Effect of low pHi on ENaC activity. In inside-out recordings, with pHi at 6.8 (A) and 6.2 (B), channel activity was significantly decreased. This effect was reversed when pHi was returned to 7.2. The lower traces in each panel show the currents on an expanded time scale, as indicated by the bars.

View larger version (25K): [in this window] [in a new window]   Fig. 6. Effect of pHi on ENaC activity. In an inside-out recording, low pHi significantly and reversibly reduced ENaC activity whereas high pHi reversibly enhanced ENaC activity. Lower traces show selected parts of the current recordings as indicated (1–8).

View larger version (2K): [in this window] [in a new window]   Fig. 7. A summary of pHi effect on ENaC activity. The open probability (NPo) values at each pH are normalized to that at pH 7.2.

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