Flow-dependent activation of maxi K+ channels in apical membrane of rabbit connecting tubule

The Ca2+-activated maxi K+ channel was found in the apical membrane of everted rabbit connecting tubule (CNT) with a patch-clamp technique. The mean number of open channels (NPo) was markedly increased from 0.007 +/- 0.004 to 0.189 +/- 0.039 (n = 7) by stretching the patch membrane in a cell-attached configuration. This activation was suggested to be coupled with the stretch-activation of Ca2+-permeable cation channels, because the maxi K+ channel was not stretch-activated in both the cell-attached configuration using Ca2+-free pipette and in the inside-out one in the presence of 10 mm EGTA in the cytoplasmic side. The maxi K+ channel was completely blocked by extracellular 1 microM charybdotoxin (CTX), but was not by cytoplasmic 33 microM arachidonic acid (AA). On the other hand, the low-conductance K+ channel, which was also found in the same membrane, was completely inhibited by 11 microM AA, but not by 1 microM CTX. The apical K+ conductance in the CNT was estimated by the deflection of transepithelial voltage (DeltaVt) when luminal K+ concentration was increased from 5 to 15 mEq. When the tubule was perfused with hydraulic pressure of 0.5 KPa, the DeltaVt was only -0. 7 +/- 0.4 mV. However, an increase in luminal fluid flow by increasing perfusion pressure to 1.5 KPa markedly enhanced DeltaVt to -9.4 +/- 0.9 mV. Luminal application of 1 microM CTX reduced the DeltaVt to -1.3 +/- 0.6 mV significantly in 6 tubules, whereas no significant change of DeltaVt was recorded by applying 33 microM AA into the lumen of 5 tubules (DeltaVt = -7.2 +/- 0.5 mV in control vs. DeltaVt = -6.7 +/- 0.6 mV in AA). These results suggest that the Ca2+-activated maxi K+ channel is responsible for flow-dependent K+ secretion by coupling with the stretch-activated Ca2+-permeable cation channel in the rabbit CNT.