Abstract

The depolarization of neurons induced by impairment of Na⁺,K⁺-ATPase activity after long-term opiate treatment has been shown to involve the development of opioid dependence. However, the mechanisms underlying changes in Na⁺,K⁺-ATPase activity after opioid treatment are unclear. The best-established molecular adaptation to long-term opioid exposure is up-regulation of the cAMP/cAMP-dependent protein kinase (PKA) signaling pathway; this study, therefore, was undertaken to investigate the role of up-regulation of cAMP/PKA signaling pathway in alteration of the mouse hippocampal Na⁺,K⁺-ATPase activity. The results demonstrated that short-term morphine treatment dose dependently stimulated Na⁺,K⁺-ATPase activity. This action could be significantly suppressed by adenylyl cyclase activator 7β-acetoxy-8,13-epoxy-1α,6β,9α-trihydroxylabd-14-en-11-one (forskolin), or the cAMP analog dibutyryl-cAMP. Contrary to short-term morphine treatment, long-term treatment significantly inhibited Na⁺,K⁺-ATPase activity. Moreover, an additional decrease in Na⁺,K⁺-ATPase activity was observed by naloxone precipitation. The effects of both short- and long-term morphine treatment on Na⁺,K⁺-ATPase activity were naltrexone-reversible. The regulation of Na⁺,K⁺-ATPase activity by morphine was inversely correlated with intracellular cAMP accumulation. N-[2-(4-Bromocinnamylamino)ethyl]-5-isoquinoline (H89), a specific PKA inhibitor, mimicked the stimulatory effect of short-term morphine but antagonized the inhibitory effect of long-term morphine treatment on Na⁺,K⁺-ATPase activity. However, okadaic acid, a protein phosphatase inhibitor, suppressed short-term morphine stimulation but potentiated long-term morphine inhibition of Na⁺,K⁺-ATPase activity. The regulation of Na⁺,K⁺-ATPase activity by morphine treatment seemed to associate with the alteration in phosphorylation level but not to be relevant to the change in abundance of Na⁺,K⁺-ATPase. These findings strongly demonstrate that cAMP/PKA signaling pathway involves regulation of Na⁺,K⁺-ATPase activity after activation of opioid receptors.