Mu and delta opioid receptor desensitization in undifferentiated human neuroblastoma SHSY5Y cells

Abstract

Both mu and delta opioid receptors are expressed in undifferentiated human neuroblastoma SHSY5Y cells and are negatively coupled to adenylate cyclase. The ability of various mu opioid, delta opioid and alpha-2 adrenergic agonists to inhibit acutely forskolin-stimulated adenylate cyclase activity in undifferentiated SHSY5Y cells after chronic administration with the selective mu opioid agonist [N-MePhe3,D-Pro4]morphiceptin (PLO17) or delta opioid agonist, [D-Pen2,D-Pen5]enkephalin (DPDPE) was assessed. In control cells, both PLO17 and DPDPE inhibited cyclic AMP (cAMP) formation with equal maximal inhibition, i.e., 60 +/- 3 and 66 +/- 2%, having IC50 values of 51.1 +/- 1.3 and 3.7 +/- 1.0 nM, respectively. The inhibition of intracellular cAMP formation by both agonists could be blocked by pertussis toxin pretreatment. After 24 hr of chronic administration of PLO17 (50 nM to 10 microM), a concentration-dependent loss of the ability of mu opioid agonists PLO17 and DAMGO, but not the delta opioid agonists DPDPE, nor alpha-2 adrenergic agonist UK-14304 (5-Bromo-N-(4,5,-dihydro-1H-imidazol-2-yl)-6-quinoxalinamine) to inhibit adenylate cyclase activity was observed. In contrast, chronic administration of DPDPE (0.1 nM to 0.3 microM) resulted in a concentration-dependent reduction in the inhibition of cAMP formation produced by delta opioid agonists DPDPE and DSLET, but not mu opioid, nor alpha-2 adrenergic agonists tested. The observed homologous desensitization was also time-dependent. In addition, antagonist-induced increases in adenylate cyclase activity were observed only after chronic PLO17 administration.2+ Finally, chronic pretreatment of cells with PLO17 (10 microM) resulted in a significant decrease in mu opioid, but not delta opioid receptor, binding, whereas treatment with DPDPE (0.3 microM) resulted in a significant decrease in delta opioid, but not mu opioid receptor binding. Therefore, undifferentiated SHSY5Y cells may provide an excellent model system to study not only the signal transduction mechanisms of mu and/or delta opioid receptors, but also the cellular adaptations of specific opioid receptors.