Effects of extracellular Mg2+ on transepithelial capacitance and Na+ transport in A6 cells under different osmotic conditions.
The electrophysiological characteristics of monolayers of cultured renal epithelial A6 cells were studied under short-circuit conditions. Replacing basolateral isosmotic (260 mOsm/kg H2O) media by hyposmotic (140 mOsm/kg H2O) solutions transiently increased the transepithelial capacitance (C(T)) by 57.3+/-2.3% after 16 min. The transepithelial Na+ current (I(Na)) increased concomitantly from 4.2+/-0.7 to 26.1+/-2.6 microA/cm2 with a time course that was noticeably slower, reaching its maximum after 60 min of hypotonicity. The transepithelial conductance (G(T)) increased synchronously with I(Na). Analysis of blocker-induced noise in I(Na), using the amiloride analogue 6-chloro-3,5-diaminopyrazine-2-carboxamide (CDPC), showed that the hypotonic shock increased Na+ channel density (N(T)) at the apical border. The presence of 10 mM Mg2+ on both sides of the epithelium suppressed the hypotonicity-induced C(T) increase to 14.3+/-0.5%, whereas the I(Na) increase was even larger than without Mg2+. Both effects of Mg2+ were located at an extracellular, basolateral site, because apical administration was without effect, whereas the acute basolateral addition of Mg2+ at the moment of the hypotonic shock was sufficient. Interaction between Mg2+ and Ca2+ influenced the behaviour of C(T). At constant osmolality (200 mOsm/kg H2O) 10 mM Mg2+ increased I(Na), leaving C(T) unaffected, whereas 10 mM Ca2+ stimulated both I(Na) and CT. In the presence of 1 mM Mg2+, however, the Ca(2+)-induced CT increase was abolished. The failure of CT to increase during stimulation of I(Na) by Mg2+ suggests that the divalent cation activates pre-existing channels in the apical membrane. Noise analysis showed that the natriferic effects of Mg2+ were also mediated by an increase in NT. The moderate initial increase in CT in the presence of Mg2+ under hypotonic conditions, occurring in parallel with increases in GT and I(Na), reflects most likely Na+ channel insertion induced by the hypotonic treatment. However, the large, transient, Mg(2+)-sensitive increase in CT, not correlated with increases in GT and I(Na), seems to be unrelated to Na+ channel recruitment.
Jans
D
D
Laboratory of Physiology, K.U. Leuven, Campus Gasthuisberg O/N, Belgium.
Simaels
J
J
Cucu
D
D
Zeiske
W
W
Van Driessche
W
W
D000818:Animals; D016477:Artifacts; D001692:Biological Transport / Q000187:drug effects / Q000502:physiology; D002118:Calcium / Q000494:pharmacology; D016764:Cell Polarity / Q000187:drug effects / Q000502:physiology; D004847:Epithelial Cells / Q000737:chemistry / Q000187:drug effects / Q000378:metabolism*; D053503:Epithelial Sodium Channels; D005089:Exocytosis / Q000502:physiology; D007038:Hypotonic Solutions / Q000494:pharmacology; D015640:Ion Channel Gating / Q000187:drug effects / Q000502:physiology; D007668:Kidney / Q000166:cytology*; D008274:Magnesium / Q000494:pharmacology*; D009994:Osmolar Concentration; D018408:Patch-Clamp Techniques; D012964:Sodium / Q000378:metabolism*; D015222:Sodium Channels / Q000378:metabolism; D014882:Water-Electrolyte Balance / Q000502:physiology*
D053503:Epithelial Sodium Channels; D007038:Hypotonic Solutions; D015222:Sodium Channels; D012964:Sodium; D008274:Magnesium; D002118:Calcium