Guang Yang1, Meng-Hua Zhou, Zhen Ren, Jia-Jie Xu, Yan-Ai Mei. 1. Institutes of Brain Science, School of Life Sciences and State Key Laboratory of Medical Neurobiology, Fudan University, Shanghai, PR China.
Abstract
BACKGROUND: Although tricyclic antidepressants amoxapine is proposed to target 5-HT and D2 receptors, very few studies have addressed the effect of amoxapine on molecular and cellular mechanisms via receptor pathways. In this study, we test the effect of amoxapine on rat cerebellar granule neurons (CGNs) to address this possibility. METHODS: CGNs cell culture, whole-cell current recording using a patch-clamp technique, western blot and non-radioactive detection analysis of phosphorylated protein kinase A (PKA) were used. RESULTS: Amoxapine inhibits delayed rectifier potassium (I(K)) current in a dose-dependent manner and modulates inactivation properties in CGNs. Those effects were not eliminated by preincubation with 5-HT or 5-HT receptor antagonists, but abolished by dopamine and D1/D5 receptor antagonists. Application of GTPγ-S and inhibitor of the Gs signalling cascade abolished the amoxapine-induced effect on I(K). The application of forskolin or dibutyryl-cAMP mimicked the inhibitory effect of amoxapine on I(K). Western blotting for phosphorylated PKA revealed that amoxapine significantly increased the intracellular levels of phosphorylated PKA, a marker of PKA activation. CONCLUSION: Amoxapine inhibits I(K) currents in rat CGNs via cAMP/PKA-dependent pathways, as in mouse cortical neurons we reported earlier, but that involves D1-like receptors instead of 5-HT receptors.
BACKGROUND: Although tricyclic antidepressants amoxapine is proposed to target 5-HT and D2 receptors, very few studies have addressed the effect of amoxapine on molecular and cellular mechanisms via receptor pathways. In this study, we test the effect of amoxapine on rat cerebellar granule neurons (CGNs) to address this possibility. METHODS: CGNs cell culture, whole-cell current recording using a patch-clamp technique, western blot and non-radioactive detection analysis of phosphorylated protein kinase A (PKA) were used. RESULTS:Amoxapine inhibits delayed rectifier potassium (I(K)) current in a dose-dependent manner and modulates inactivation properties in CGNs. Those effects were not eliminated by preincubation with 5-HT or 5-HT receptor antagonists, but abolished by dopamine and D1/D5 receptor antagonists. Application of GTPγ-S and inhibitor of the Gs signalling cascade abolished the amoxapine-induced effect on I(K). The application of forskolin or dibutyryl-cAMP mimicked the inhibitory effect of amoxapine on I(K). Western blotting for phosphorylated PKA revealed that amoxapine significantly increased the intracellular levels of phosphorylated PKA, a marker of PKA activation. CONCLUSION:Amoxapine inhibits I(K) currents in rat CGNs via cAMP/PKA-dependent pathways, as in mouse cortical neurons we reported earlier, but that involves D1-like receptors instead of 5-HT receptors.