Stimulation of Na+-alanine cotransport activates a voltage-dependent conductance in single proximal tubule cells isolated from frog kidney

摘要

class="enumerated" style="list-style-type:decimal">The swelling induced by Na⁺-alanine cotransport in proximal tubule cells of the frog kidney is followed by regulatory volume decrease (RVD). This RVD is inhibited by gadolinium (Gd³⁺), an inhibitor of stretch-activated channels, but is independent of extracellular Ca²⁺.In this study, the whole cell patch clamp technique was utilized to examine the effect of Na⁺-alanine cotransport on two previously identified volume- and Gd³⁺-sensitive conductances. One conductance is voltage dependent and anion selective (GVD) whilst the other is voltage independent and cation selective (GVI).Addition of 5 mM L-alanine to the bathing solution increased the whole cell conductance and gave a positive (depolarizing) shift in the reversal potential (Vrev, equivalent to the membrane potential in current-clamped cells) consistent with activation of Na⁺-alanine cotransport. Vrev shifted from -36 ± 4·9 to +12·9 ± 4·2 mV (n= 15).In the presence of alanine, the total whole cell conductance had several components including the cotransporter conductance and GVD and GVI. These conductances were separated using Gd³⁺, which inhibits both GVD and GVI, and the time dependency of GVD. Of these two volume-sensitive conductances, L-alanine elicited a specific increase in GVD, whereas GVI was unaffected.The L-alanine-induced activation of GVD was significantly reduced when cells were incubated in a hypertonic bathing solution.In summary, in single proximal tubule cells isolated from frog kidney, on stimulation of Na⁺-alanine cotransport GVD is activated, while GVI is unaffected. Taken with other evidence, this suggests that GVD is activated by cell swelling, consequent upon alanine entry, and may play a role as an anion efflux pathway during alanine-induced volume regulation.