Extracellular Ca²⁺ controls outward rectification by apical cation channels in toad urinary bladder: Patch-clamp and whole-bladder studies

Summary

Outward rectifying. cation channels were observed in the epithelial cells of the urinary bladder of the toad.Bufo marinus. As studied in isolated cells using the patch-clamp technique, the channel has an average conductance of 24 and 157 pS for pipette potentials between 0 and +60 mV and −60 to −100 mV, respectively, when the major cation in both bath and pipette solutions is K⁺. The conductance of the cannel decreasen with increasing dehydration energy of the permeant monovalent cation in the oder Rb⁺=K⁺>Na⁺>Li⁺. Reversal potentials near zero under biionic conditions imply that the permeabilities for all four of these cations are smiliar. The channel is sensitive to quinidine sulfate but not to amiloride. It shares several pharmacological and biophysical properties with an outwardly-rectifying, vasopressin-sensitive pical K⁺ conductive pathway described previously for the toad urinary bladder. We demonstrate, in both single-channel and whole-bladder studies, that the outward rectification is a consequence of interaction of the chanel with extracellular divalent cations, particularly Ca²⁺, which blocks inward but not outward current. Various divalent cations impart different degrees of outward rectification to the conductive pathway. Concentrations of Mg²⁺ and Ca²⁺ required for halfmaximal effect are 3×10⁻⁴ and 10⁻⁴ m, resopectively. For Co²⁺ the values are 10⁻⁶ m at +50 mV and a 10⁻⁴ m at +200 mV. The mechanism of blockade by divalent cations is not established, but does not seem to involve a voltage-dependent interaction in which the blocker penetrates the transmembrane electric field. In the absence of divalent cations in the mucosal solution, the magnitudes of inward current carried by Rb⁺, K⁺, Na⁺ and Li⁺ through the apical K⁺ pathway at any transepithelial voltage, are in the same order as in the single-channel studies. We propose that the cation channel observed by us in isolated epithelial cells is the single-channel correlate of the vasopressin-sensitive apical K⁺ conductive pathway in the toad urinary bladder and is also related to the oxytocin- and divalent cation-sensitive apical condictivity observed in frog skin and urinary bladder.