Functional acetylcholine receptor--electroplax membrane microsacs (vesicles): purification and characterization.

Abstract

Kinetic analysis of the flux of Na+ in a heterogeneous population of acetylcholine receptor-rich microsacs (vesicles) formed by membrane fragments of [Electrophorus electricus] electroplax indicated that functional microsacs, which on average comprise only 15% of the preparation, can be filled with 190 mM NaCl while nonfunctional microsacs are filled by 190 mM CsCl. The functional microsacs were successfully separated from nonfunctional microsacs on the basis of their density differences with a continuous sucrose-190 mM CsCl density gradient. In the presence of acetylcholine analogs, all the internal Na+ in these microsacs rapidly exchanged with external ions. The efflux of Na+ followed a single exponential decay. The isolation of functional microsacs opened up at least 2 new avenues of investigation of the molecular mechanism of receptor-mediated processes. The 1st deals with the efficiency of the process, and the 2nd with the characterization of membrane components important in this process. The efficiency of the receptor-mediated process that allows inorganic ions to equilibrate across the membranes of the microsacs can adequately account for electrophysiological results obtained with muscle and nerve cells. In the receptor-rich heterogeneous population of microsacs the concentration of receptor sites in functional and nonfunctional microsacs is about the same and is therefore not the only factor determining functionality. Significant differences between functional and nonfunctional microsacs were found in the concentrations of acetylcholinesterase and Na+-K+ ATPase.