OPIATE RECEPTOR DOWN-REGULATION AND DESENSITIZATION IN NEURO-BLASTOMA X GLIOMA NG108-15 HYBRID-CELLS ARE 2 SEPARATE CELLULAR ADAPTATION PROCESSES

Abstract

Chronic treatment of neuroblastoma .times. glioma NG108-15 hybrid cells with the opiate agonist etorphine resulted in a decrease in both opiate receptor density (receptor down-regulation) and opiate ability to inhibit prostaglandin E1 (PGE1)-stimulated increases in cAMP levels (receptor desensitization). Opiate receptor down-regulation and desensitization were homologous as indicated by the lack of apparent change in muscarinic, .alpha.2-adrenergic and PGE1 receptor binding and also retention albeit modulation of the ability of carbachol and norepinephrine to inhibit PGE1-stimulated increases in cAMP levels after 24 h of etorphine treatment. PGE1-stimulated increases in cAMP levels remained identical in etorphine-treated and control cells. Receptor desensitization and receptor down-regulation in NG108-15 cells may be 2 separate cellular adaptation processes because: with an agonist which appears to be efficiently coupled, i.e., an agonist whose apparent Kd value is much larger than its apparent IC50 [median inhibitory concentration] value for regulation of cAMP levels (Ki), the concentration of ligand required to produce half-maximal down-regulation is analogous to its Ki value, while the concentration of ligand required to produce half-maximal desensitization is related to its Kd value; receptor desensitization precedes receptors down-regulation; only opiate agonists could produce receptor down-regulation, while both opiate agonists and partial agonists could desensitize post-receptor occupancy events. Still further evidence for dissociability of these processes was obtained by incubating NG108-15 cells with etorphine at 30.degree. C for 2 h which produced a decrease in etorphine''s ability to regulate adenylate cyclase while [3H]diprenorphine binding remained unaltered. IC50 values of D-Ala2-D-Leu5-enkephalin''s competition for [3H]diprenorphine binding to intact cells increased 19.6-fold after etorphine treatment for 90 min, while naloxone IC50 values remained unalatered. This apparent increase in IC50 values was much lower, .apprx. 2-fold, when receptor binding was carried out in membranes isolated from cells treated with etorphine chronically. Analysis of [3H]etorphine binding to such membranes in the presence of 10 mM Mg2+ indicated a loss of receptor binding sites with no change in apparent affinity, while [3H]diprenorphine binding revealed no significant alteration in either Bmax [maximal binding capacity] or Kd values. During opiate receptor desensitization, a reduction of agonist high-affinity sites occurs with no apparent alteration in total receptor number.