Vol. 302, Issue 3, 1002-1012, September 2002

Differential Effects of µ-Opioid Receptor Ligands on Ca²⁺ Signaling

J. Mark Quillan, Kurt W. Carlson, Chunyan Song, Danxin Wang and Wolfgang Sadée

Departments of Biopharmaceutical Sciences and Pharmaceutical Chemistry, University of California, San Francisco, San Francisco, California (J.M.Q., K.W.C., C.S., D.W., W.S.); and Department of Pharmacology, College of Medicine and Public Health, The Ohio State University, Columbus Ohio (D.W., W.S.)

Activation of µ-opioid receptors (MORs) transfected into human embryonic kidney 293 cells, caused a multiphasic increase in cytosolic free Ca²⁺ levels (Ca²⁺i). The first Ca²⁺i maximum (peak 1) between 5 and 7 s depended on the presence of extracellular Ca²⁺ (Ca²⁺e). The second phase peaking at ~15 s (peak 2) was independent of Ca²⁺e and thus represents Ca²⁺ release from intracellular stores. A decrease in temperature from 37 to 25°C also caused reduction of peak 1 but not peak 2, suggesting that the two responses arise from mechanistically distinct pathways. A delayed Ca²⁺e-dependent third response phase is thought to represent capacitative Ca²⁺e influx evoked after release of Ca²⁺ from internal stores. Agonists and antagonists of two major classes of opioid ligands, oxymorphinans (morphine and naloxone) and oripavines (etorphine and diprenorphine), had differential effects on Ca²⁺ currents. Although morphine activated both phases with equal potency, etorphine was 20-fold less potent at stimulating peak 1 over peak 2. Similarly, the antagonists, naloxone and diprenorphine, blocked the Ca²⁺ response to each agonist with greatly varying potencies. Specifically, concomitant injection of diprenorphine failed to affect peak 1 (thought to represent rapid Ca²⁺e influx) stimulated by morphine while fully blocking peak 2 (intracellular Ca²⁺ release). However, diprenorphine potently inhibited peak 1 as well when added to the cells before morphine, indicating limited or slow access of diprenorphine to these morphine binding sites. The existence of multiple, functionally distinct binding site conformations could account for these findings. In conclusion, different opioid ligands can differentially affect Ca²⁺ response patterns resulting from MOR activation.