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Maxi-K channels consist of a pore-forming alpha subunit and a regulatory beta subunit, which confers the channel with a higher Ca(2+) sensitivity. Estradiol bound to the beta subunit and activated the Maxi-K channel (hSlo) only when both alpha and beta subunits were present. This activation was independent of the generation of intracellular signals and could be triggered by estradiol conjugated to a membrane-impenetrable carrier protein. This study documents the direct interaction of a hormone with a voltage-gated channel subunit and provides the molecular mechanism for the modulation of vascular smooth muscle Maxi-K channels by estrogens.

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Addition of membrane vesicles prepared from transverse tubule (T-tubule) membranes of rabbit skeletal muscle to the aqueous phase of a planar lipid bilayer induces a stepwise increase in conductance. This conductance is both voltage and Ca2' dependent. At 1 mM Ca2+, the steady-state conductance is maximal at voltages higher than +20 mV and decreases for more negative voltages. (Voltages refer to the side to which the vesicles are added, cia.) Decreasing the Ca2+ concentration reversibly shifts the conductance-voltage curve toward the right along the voltage axis. Furthermore, Ca2+ can activate the conductance only if added to the cis compartment. Neither Mg2+, Ba2+, nor Cd2+ can activate the conductance induced by T-tubule vesicles. Addition of 5 mM tetraethylammonium ion to the trans, but not the cis, side abolishes the T-tubule-induced conductance. The Ca2+_ dependent conductance appears as a consequence ofionic channel formation. Single-channel activity appears in bursts followed by periods of time in which the channel remains "silent". The conductance of the oven channel averages 226 pS in 0.1 M KCl and is voltage and Ca + independent. However, the fraction of time that the channel remains in the open state is voltage and Ca2+ dependent in a manner that parallels the voltage and Ca2+ dependence ofthe multichannel membrane. The channel is 6.6 times more permeable to K+ than to Na+ and is impermeable to C1-.In striated muscle, the transverse tubular (T-tubule) system forms pathways for conducting the electrical activity of the surface membrane deep into the muscle fiber. Although the electrical properties ofT-tubules appear to be different from those of surface membrane (1-3), their study is made difficult by the geometry of the system. A possibility for direct study of the isolated T-tubule membrane has recently been opened by the isolation ofvesicular membrane preparations of T-tubule origin (4, 5). Rosemblatt et aL (5) have been able to obtain a highly purified preparation of T-tubule membranes and have shown that it is altogether different in lipid composition, protein composition, and enzymatic activities from either surface membranes or sarcoplasmic reticulum membranes.The objective ofthe present work was to study the interaction of T-tubule membrane vesicles with planar lipid bilayer membranes. This was done in the hope that we would be able to reconstitute in the planar bilayer one or more of the ion-conductive pathways present in the T-tubule membrane. In this way, the ion-conducting protein(s) could be studied in a membrane having a low background conductance and a well-defined and controllable composition. The former characteristic is important because it allows for the possibility of resolving singlechannel activity (6).Following the methods described by Miller and Racker (7) and by Miller (8), we have developed a reproducible procedure to incorporate a Ca2+-dependent K+ conductance present in Ttubule vesicles into bilayers. Ca2+-activated K+ conductances can be found in a great variety of...