科研之友
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摘要
According to the lipid raft theory, the plasma membrane contains small domains enriched in cholesterol and sphingolipid, which may serve as platforms to organize membrane proteins. Using methyl-beta-cyclodextrin (M beta CD) to deplete membrane cholesterol, many G protein-coupled receptors have been shown to depend on putative lipid rafts for proper signaling. Here we examine the hypothesis that treatment of HEK293 cells stably expressing FLAG-tagged mu-opioid receptors (HEK FLAG-mu) or delta-opioid receptors (HEK FLAG-delta) with M beta CD will reduce opioid receptor signaling to adenylyl cyclase. The ability of the mu-opioid agonist [D-Ala(2),N-Me-Phe(4),Gly(5)-ol] enkephalin to acutely inhibit adenylyl cyclase or to cause sensitization of adenylyl cyclase following chronic treatment was attenuated with M beta CD. These effects were due to removal of cholesterol, because replenishment of cholesterol restored [D-Ala(2),N-Me-Phe(4),Gly(5)-ol] enkephalin responses back to control values, and were confirmed in SH-SY5Y cells endogenously expressing mu-opioid receptors. The effects of M beta CD may be due to uncoupling of the mu receptor from G proteins but were not because of decreases in receptor number and were not mimicked by cytoskeleton disruption. In contrast to the results in HEK FLAG-mu cells, M beta CD treatment of HEK FLAG-delta cells had no effect on acute inhibition or sensitization of adenylyl cyclase by delta-opioid agonists. The differential responses of mu- and delta-opioid agonists to cholesterol depletion suggest that mu-opioid receptors are more dependent on cholesterol for efficient signaling than delta receptors and can be partly explained by localization of mu- but not delta-opioid receptors in cholesterol- and caveolin-enriched membrane domains.
- 出版日期2009-8-14
