Evidence for ammonium conductance in a mouse thick ascending limb cell line

Abstract

In this study, we examined an ammonium conductance in the mouse thick ascending limb cell line ST‐1. Whole cell patch clamp was performed to measure currents evoked by NH 4Cl in the presence of BaCl2, tetraethylammonium, and BAPTA. Application of 20 mmol/L NH 4Cl induced an inward current (−272 ± 79 pA, n = 9). In current‐voltage (I–V) relationships, NH 4Cl application caused the I–V curve to shift down in an inward direction. The difference in current before and after NH 4Cl application, which corresponds to the current evoked by NH 4Cl, was progressively larger at more negative potentials. The reversal potential for NH4Cl was +15 mV, higher than the equilibrium potential for chloride, indicating that the current should be due to NH 4 +. We then injected ST‐1 poly(A) RNA into Xenopus oocytes and performed two‐electrode voltage clamp. NH 4Cl application in the presence of BaCl2 caused the I–V curve to be steeper. The NH 4 + current was retained at pH 6.4, where endogenous oocyte current was abolished. The NH 4 + current was unaffected by 10 μmol/L amiloride but abolished after incubation in Na+‐free media. These results demonstrate that the renal cell line ST‐1 produces an NH 4 + conductance.

Lack of Charge Interaction in the Ion Binding Site Determines Anion Selectivity in the Sodium Bicarbonate Cotransporter NBCe1

“…Oocyte pH i was measured using a proton-selective glass electrode, as described before [ 36 ]. Briefly, a pH electrode was made with a borosilicate glass capillary that was silanized, filled with the proton ionophore 1 cocktail B (cat no: 95293, MilliporeSigma, St. louis, MOP, USA), and back-filled with pH 7.0 phosphate buffer.…”