Abstract A method has been adapted to produce membrane vesicles suitable for routine membrane patch clamping from neonate rat skeletal muscle. Single K + channel activity was recorded from cell-free inside-out patches. Most Ca 2+-activated voltage sensitive channels had large conductances of up to 300 pS, as determined from their current/voltage relationship, and an open probability (P o) approaching unity at positive membrane potentials. A lower conductance K + channel, probably responsible for inward rectification, had a lower conductance of about 100 pS. Outward rectifying K + channels were also observed with the lowest conductance, about 40 pS. 0.1 mM ATP when applied to the inner membrane surface reduced or blocked activity, drastically reducing P o without altering single channel conductance. Such an effect has been reported in other preparations but was different in the neonate preparation in that it blocked channels with conductances as high as 300 pS. The simple preparation described, which we have also used successfully on mature rat and mouse skeletal muscle, has potential in the analysis of channel activities in various conditions and pathologies without the need for tissue culture to produce suitable membrane preparations.