Miller MB, Wilson RS, Lam TT, Nairn AC, Picciotto MR

Activation of nicotinic acetylcholine receptors containing ?4 and ?2 subunits (?4/?2* nAChRs) in the mammalian brain is necessary for nicotine reinforcement and addiction. We previously identified interactions between ?4/?2* nAChRs and calcium/calmodulin-dependent protein kinase II (CaMKII) in mouse and human brain tissue. Following co-expression of ?4/?2 nAChR subunits with CaMKII ...

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in HEK cells, mass spectrometry identified 8 phosphorylation sites in the ?4 subunit. One of these sites and an additional site were identified when isolated ?4/?2* nAChRs were dephosphorylated and subsequently incubated with CaMKII in vitro, while 3 phosphorylation sites were identified following incubation with protein kinase A (PKA) in vitro. We then isolated native ?4/?2* nAChRs from mouse brain following acute or chronic exposure to nicotine. Two CaMKII sites identified in HEK cells were phosphorylated, and 1 PKA site was dephosphorylated following acute nicotine administration in vivo, whereas phosphorylation of the PKA site was increased back to baseline levels following repeated nicotine exposure. Significant changes in ?2 nAChR subunit phosphorylation were not observed under these conditions, but 2 novel sites were identified on this subunit, 1 in HEK cells and 1 in vitro. These experiments identified putative CaMKII and PKA sites on ?4/?2* nAChRs and novel nicotine-induced phosphorylation sites in mouse brain that can be explored for their consequences on receptor function.

Date: Oct. 15, 2018

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