The whole-cell patch-clamp technique was used to examine nonselective conductances in single proximal tubule cells isolated from mouse kidney. Single cells were isolated in either the presence or absence of a cocktail designed to stimulate cAMP. Patches were obtained with Na+ Ringer in the bath and Cs+ Ringer in the pipette. On initially achieving the whole-cell configuration, whole-cell currents were small. In cAMP-stimulated cells, with 5 niM ATP in the pipette solution, whole-cell currents increased with time. The activated current was linear, slightly cation-selective, did not discriminate between Na+ and K+ and was inhibited by 100 µm gadolinium. These properties are consistent with the activation of a nonselective conductance, designated Gns. Activation of Gns was abolished with pipette AMP-PNP, ATP plus alkaline phosphatase or in the absence of ATP. In unstimulated cells GNS was activated by pipette ATP together with PKA. These data support the hypothesis that GNS is activated by a PKA-mediated phosphorylation event. GNs was also activated by a hypertonic shock. However, GNS does not appear to be involved in regulatory volume increase (RVI), as RVI was unaffected in the presence of the GNS blocker gadolinium. Instead, the ATP sensitivity of GNs suggests that it may be regulated by the metabolic state of the renal proximal tubule cell.
The Journal of Membrane Biology – Springer Journals
Published: Mar 25, 2002
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