AKAP79/150 Signal Complexes in G-Protein Modulation of Neuronal Ion Channels

Abstract

Voltage-gated M-type (KCNQ) K+ channels play a critical role in modulation of neuronal excitability and action potential firing. A-kinase-anchoring protein (AKAP)79/150 mediated PKC phosphorylation of M channels is involved in M current (IM) suppression by muscarinic M1, but not bradykinin B2 receptors. In this study, we first explored if purinergic and angiotensin suppression of IM in superior cervical ganglion (SCG) sympathetic neurons involves AKAP79/150. Transfection into rat SCG neurons of ΔA-AKAP79, which lacks the A-domain necessary for PKC binding, or the absence of AKAP150 in AKAP150-/- mice, did not affect IM suppression by the purinergic agonist UTP, nor by bradykinin, but did reduce IM suppression by muscarinic agonists and by angiotensin II. Transfection of AKAP79, but not ΔA-AKAP79 or AKAP15, “rescued” the muscarinic suppression of IM in AKAP150-/- neurons. We also tested association of AKAP79 or KCNQ channels with M1, B2, P2Y6 and AT1 receptors via fluorescence resonance energy transfer (FRET) experiments on CHO cells under total internal refection fluorescence microscopy, which revealed substantial FRET between AKAP79 with M1 and AT1 receptors, but only weak FRET with P2Y6 or B2 receptors. Similarly, we observed strong FRET between KCNQ2 with M1 and AT1, but not P2Y6 or B2, receptors. The involvement of AKAP79/150 in the regulation of N- and L-type Ca2+ channels in SCG neurons by Gq/11-coupled muscarinic receptors and by cAMP/PKA was also studied. We found AKAP79/150 to not play a role in the former, but to be necessary for forskolin-induced up-regulation of the L-type current. Our data suggest that AKAP79/150 orchestrates signal complexes that include PKC, KCNQ subunits and M1 or AT1 receptors, which correlates with the PIP2-depletion mode of neuronal M current suppression, but does not generalize to Gq/11-mediated inhibition of N- or L-type Ca2+ channels.