Dopamine modulation of neuronal Na(+) channels requires binding of A kinase-anchoring protein 15 and PKA by a modified leucine zipper motif.
In hippocampal pyramidal cells, dopamine acts at D1 receptors to reduce peak Na(+) currents by activation of phosphorylation by PKA anchored via an A kinase-anchoring protein (AKAP15). However, the mechanism by which AKAP15 anchors PKA to neuronal Na(+) channels is not known. By using a strategy of coimmunoprecipitation from transfected tsA-201 cells, we have found that AKAP15 directly interacts with Na(v)1.2a channels via the intracellular loop between domains I and II. This loop contains key functional phosphorylation sites. Mutagenesis indicated that this interaction occurs through a modified leucine zipper motif near the N terminus of the loop. Whole-cell patch clamp recordings of acutely dissociated hippocampal pyramidal cells revealed that the D1 dopamine receptor agonist SKF 81297 reduces peak Na(+) current amplitude by 20.5%, as reported previously. Disruption of the leucine zipper interaction between Na(v)1.2a and AKAP15 through the inclusion of a small competing peptide in the patch pipette inhibited the SKF 81297-induced reduction in peak Na(+) current, whereas a control peptide with mutations in amino acids important for the leucine zipper interaction did not. Our results define the molecular mechanism by which G protein-coupled signaling pathways can rapidly and efficiently modulate neuronal excitability through local protein phosphorylation of Na(+) channels by specifically anchored PKA.
Few
W Preston
WP
Department of Pharmacology, University of Washington, Seattle, WA 98195-7280, USA.
Scheuer
Todd
T
Catterall
William A
WA
D048868:Adaptor Proteins, Signal Transducing / Q000737:chemistry / Q000378:metabolism*; D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D000818:Animals; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism*; D004298:Dopamine / Q000494:pharmacology*; D006624:Hippocampus / Q000187:drug effects; D016350:Leucine Zippers; D051379:Mice; D008969:Molecular Sequence Data; D050505:Mutant Proteins / Q000737:chemistry; D062551:NAV1.2 Voltage-Gated Sodium Channel; D009419:Nerve Tissue Proteins; D009474:Neurons / Q000187:drug effects* / Q000378:metabolism*; D011485:Protein Binding / Q000187:drug effects; D017434:Protein Structure, Tertiary / Q000187:drug effects; D051381:Rats; D015222:Sodium Channels / Q000737:chemistry / Q000378:metabolism*
D048868:Adaptor Proteins, Signal Transducing; D050505:Mutant Proteins; D062551:NAV1.2 Voltage-Gated Sodium Channel; D009419:Nerve Tissue Proteins; C568253:Scn2A protein, rat; C000654500:Scn2a protein, mouse; D015222:Sodium Channels; D017868:Cyclic AMP-Dependent Protein Kinases; D004298:Dopamine