Studies of the unitary properties of adenosine-5'-triphosphate-regulated potassium channels of frog skeletal muscle

Abstract

1. Patch-clamp techniques were used to study adenosine-5'-triphosphate (ATP)-dependent K+ channels in sarcolemmal vesicles from frog skeletal muscle. In addition to its ATP dependence, opening of these channels was voltage dependent, the open-state probability (P open) increasing with depolarization. 2. The reversal potential of unitary currents changed with external K+ concentration, [K+]o, as expected if the Na-K permeability ration (pNa/pK) equals 0.015. Unitary conductance increased with increasing [K+]o from 14.8 +/- 0.5 pS (n = 5) in 2.5 mM-K+ to 42.3 +/- 1.0 pS (n = 8) in 60 mM-K+. This increase was less than that expected from independence. 3. Replacement of 60 mM-external K+ by 60 mM-external Rb+ shifted the reversal potential of unitary currents by -6.7 mV, suggesting that Rb+ enters channels nearly as easily as does K+ (Rb-K permeability ration, pRb/pK = 0.76). Unitary currents were much smaller in Rb+, consistent with Rb+ binding within the channel. 4. The ATP-regulated K+ channel was blocked by both internal and external tetraethylammonium ions (TEA+). 2 mM-TEA+, applied to the cytoplasmic face of membrane patches, interrupted channel openings. Higher concentrations reduced unitary current amplitude, suggesting an increase in the rapidity of TEA+ block. 5. The reduction in P open by ATP was consistent with 1:1 binding and a dissociation constant of 0.135 mM. ATP appeared not to be hydrolysed to close channels. Adenosine 5'-diphosphate (ADP) and adenosine 5'-monophosphate (AMP) were less effective than ATP, but retained channel closing properties. Substitution of adenine with other purines or with pyrimidine bases substantially reduced activity, as did substitution of ribose by 2'-deoxyribose or by ribose 2',3'-dialdehyde. 6. Sarcoplasmic Ca2+ did not influence P open. 7. Myotubes, grown from thigh muscles of new-born rats, appeared to lack ATP-dependent K+ channels. Adult frog muscle appeared to lack high-conductance Ca2+-dependent K+ channels, at least in the surface membrane. Such channels were found in myotube membranes. 8. Open- and closed-time histograms were constructed and were consistent with at least two open and at least three closed states. Channel openings were grouped in bursts. Open times, burst lengths and the number of openings per burst were reduced by ATP. 9. The effects of [K+]o on unitary conductance and of K+ replacement with Rb+ are discussed in terms of a simple Eyring rate theory formulation.(ABSTRACT TRUNCATED AT 400 WORDS)