CHRONIC EXPOSURE TO KAPPA-OPIOIDS ENHANCES THE SUSCEPTIBILITY OF IMMORTALIZED NEURONS (F-11-KAPPA-7) TO APOPTOSIS-INDUCING DRUGS BY A MECHANISM THAT MAY INVOLVE CERAMIDE

Chronic exposure of embryonic brain to opioids leads to microcephaly a nd developmental abnormalities. An immortalized mouse neuroblastoma x dorsal root ganglion hybrid cell line stably transfected to overexpres s kappa-opioid receptors (F-11 kappa 7) showed complete loss of kappa- receptor binding to [H-3]U69,593 after exposure to the kappa-agonist U 69,593 for 24 h. U69,593 had no measurable effect on cell viability as determined by either cell viability or DNA fragmentation assays. Howe ver, when cell death (apoptosis) was induced by either staurosporine o r the phosphatidylinositol 3-kinase inhibitors wortmannin and LY294002 , cells pretreated with U69,593 for 24 h showed increased apoptosis co mpared with un treated cells. Thus, staurosporine (50 nM), wortmannin (4 mu M), and LY294002 (30 mu M) treatment for 24 h induced a 50% loss of cell viability and DNA fragmentation in 24 h. U69,593 pretreatment produced the same killing at lower concentrations, namely, 20 nM stau rosporine, 2 mu M wortmannin, and 14 mu M LY294002, respectively. The effects of U69,593 were time-, dose-, and naloxone-reversible, suggest ing that they are receptor-mediated. However, coaddition of U69,593 at the same time as staurosporine, wortmannin, or LY294002 did not enhan ce apoptosis. All three drugs that induced apoptosis were found to inc rease the level of ceramide, and pretreatment with U69,593 further inc reased the rate of formation of ceramide, a lipid that induces apoptos is in cells. We propose that chronic exposure to kappa-receptor agonis ts promotes increased vulnerability of neurons to apoptosis.