Effects of trypsin on large-conductance Ca(2+)-activated K(+) channels of guinea-pig outer hair cells

Abstract

High-conductance Ca(2+)-activated K(+) (BK(Ca)) channels from isolated adult guinea-pig outer hair cells were studied in inside-out membrane patches. They had a 300 pS unitary conductance and were inhibited by tetraethyl ammonium (1 mM), iberiotoxin (33 nM) and charybdotoxin (50 nM). In symmetrical 144 mM KCl their K(+) permeability (P(K)) was 5.4 x 10(-13) cm(3)/s; this was reduced to around 4.5 x 10(-13) cm(3)/s with 160 mM Na(+) in place of K(+) on either internal or external membrane surface. BK(Ca) channels from trypsin-isolated hair cells had a high open probability, that depended on both membrane voltage (16 mV/e-fold change) and the concentration of calcium ions at their intracellular surface ([Ca(2+)](i)). The Hill coefficient was 3-4. About 50% of BK(Ca) channels from mechanically isolated outer hair cells had similar characteristics; the remainder had the same high conductance but a low open probability. Trypsin (<0.5 mg/ml) applied to the intracellular face of these 'inactive' channels markedly increased their open probability. It is possible that exposure to trypsin during cell isolation removes an inactivating beta subunit. This would account for the absence of 'inactive' BK(Ca) channels in trypsin-isolated cells.