Solubilization and Characterization of a [3H]Hemicholinium-3 Binding Site in Rat Brain

Abstract

A sodium-dependent high-affinity [3H]hemicholinium-3 ([3H]HCh-3) binding site was solubilized from rat striatal synaptic plasma membranes by 0.2% deoxycholate. Deoxycholate solubilization of the [3H]HCh-3 binding site was dependent upon both detergent concentration and ionic strength of the solubilization medium. Specific [3H]HCh-3 binding of the solubilized preparation was both sodium- and chloride-dependent and saturable, exhibiting an affinity of 14.2 nM and a capacity (Bmax) of 695 fmol/mg protein. Choline and other analogs inhibited specific [3H]HCh-3 binding to the solubilized preparation in a concentration-dependent manner with the similar rank order of potency observed in crude synpatic membranes. Treatments known to disrupt both protein and lipid moieties resulted in diminished specific [3H]HCh-3 binding binding. These results suggest that the characteristics of the solubilized [3H]HCh-3 binding site are similar to those of the membrane-bound site.