Functional expression of the amiloride-sensitive sodium channel in Xenopus oocytes.

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Abstract
Expression of the amiloride-sensitive sodium channel was examined in Xenopus oocytes that were microinjected with A6 cell mRNA. Amiloride-inhibitable 22Na flux could be measured in intact oocytes 2-3 days after injection with 25 ng of poly(A)+ RNA isolated from aldosterone-treated A6 cells. The rate of 22Na uptake was .apprxeq. 15-fold greater in oocytes microinjected with 25 ng of poly(A)+ RNa than in water-injected control oocytes. An increase in 22Na uptake by mRNA-injected oocytes occurred whether the mRNA was isolated from A6 cells grown on a porous or nonporous support. In the presence of 4 mM NaCl, amiloride caused dose-dependent inhibition of 22Na uptake in mRNA-injected oocytes, which was half-maximal at 6 .times. 10-8 M. Both 1 .mu.M amiloride and 0.1 .mu.M benzamil inhibited 22Na uptake in mRNA-injected oocytes by > 95%, whereas < 50% inhibition occurred with 1 .mu.M 5-(N-ethyl-N-isopropyl)amiloride. When A6 cell mRNA was size fractionated by sucrose density-gradient centrifugation, amiloride-sensitive 22Na uptake was expressed predominantly by oocytes injected with mRNA from two contiguous fractions.