Solubilization and characterization of mu, delta, and kappa opioid binding sites from guinea pig brain: physical separation of kappa receptors.

Sucrose density gradient centrifugation of digitonin-solubilized opioid binding sites from guinea pig brain and cerebellum was carried out. Centrifugation of extracts of whole brain into a gradient devoid of sodium and low in digitonin revealed the presence of two well-separated peaks of opioid binding activity. Peak A was shown to have the binding characteristics of kappa sites, whereas peak B seems to be a mixture of mu and delta sites. When extracts of guinea pig cerebellum were treated in the same manner, a single peak of binding activity was obtained that coincided with peak A from guinea pig brain and exhibited the characteristics of kappa binding sites. All three sites closely resemble their membrane-bound counterparts, retaining good affinity and selectivity for their appropriate ligands. The apparent sedimentation coefficients (S20,w) of the digitonin-solubilized binding sites present in the two peaks are 19 s for peak A and 34-39 s for peak B, and the estimated apparent molecular weights are 400,000 for kappa sites and 750,000-875,000 for the mixture of mu and delta sites. Our results suggest that kappa sites constitute separate molecular entities from mu and delta sites.