title: Sinoatrial node pacemaker activity requires Ca(2+)/calmodulin-dependent protein kinase II activation.
abstract: Cardiac beating arises from the spontaneous rhythmic excitation of sinoatrial (SA) node cells. Here we report that SA node pacemaker activity is critically dependent on Ca(2+)/calmodulin-dependent protein kinase II (CaMKII). In freshly dissociated rabbit single SA node cells, inhibition of CaMKII by a specific peptide inhibitor, autocamtide-2 inhibitory peptide (AIP, 10 micromol/L), or by KN-93 (0.1 to 3.0 micromol/L), but not its inactive analog, KN-92, depressed the rate and amplitude of spontaneous action potentials (APs) in a dose-dependent manner. Strikingly, 10 micromol/L AIP and 3 micromol/L KN-93 completely arrested SA node cells, which indicates that basal CaMKII activation is obligatory to the genesis of pacemaker AP. To understand the ionic mechanisms of the CaMKII effects, we measured L-type Ca(2+) current (I(Ca, L)), which contributes both to AP upstroke and to pacemaker depolarization. KN-93 (1 micromol/L), but not its inactive analog, KN-92, decreased I:(Ca, L) amplitude from 12+/-2 to 6+/-1 pA/pF without altering the shape of the current-voltage relationship. Both AIP and KN-93 shifted the midpoint of the steady-state inactivation curve leftward and markedly slowed the recovery of I(Ca, L) from inactivation. Similar results were observed using the fast Ca(2+) chelator BAPTA, whereas the slow Ca(2+) chelator EGTA had no significant effect, which suggests that CaMKII activity is preferentially regulated by local Ca(2+) transients. Indeed, confocal immunocytochemical imaging showed that active CaMKII is highly localized beneath the surface membrane in the vicinity of L-type channels and that AIP and KN-93 significantly reduced CaMKII activity. Thus, we conclude that CaMKII plays a vital role in regulating cardiac pacemaker activity mainly via modulating I(Ca, L) inactivation and reactivation, and local Ca(2+) is critically involved in these processes.
authors.affiliation: Laboratory of Cardiovascular Sciences, National Institute of Aging, Gerontology Research Center, Baltimore, Md, USA.
authors.forename: T M
authors.initials: TM
authors.lastname: Vinogradova
authors.forename: Y Y
authors.initials: YY
authors.lastname: Zhou
authors.forename: K Y
authors.initials: KY
authors.lastname: Bogdanov
authors.forename: D
authors.initials: D
authors.lastname: Yang
authors.forename: M
authors.initials: M
authors.lastname: Kuschel
authors.forename: H
authors.initials: H
authors.lastname: Cheng
authors.forename: R P
authors.initials: RP
authors.lastname: Xiao
mesh_terms: D000200:Action Potentials / Q000187:drug effects; D000818:Animals; D001596:Benzylamines / Q000494:pharmacology; D002118:Calcium / Q000737:chemistry / Q000502:physiology; D002121:Calcium Channel Blockers / Q000494:pharmacology; D015220:Calcium Channels / Q000187:drug effects / Q000502:physiology; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000037:antagonists & inhibitors / Q000502:physiology*; D004305:Dose-Response Relationship, Drug; D004533:Egtazic Acid / Q000031:analogs & derivatives* / Q000737:chemistry; D004789:Enzyme Activation; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006339:Heart Rate / Q000502:physiology*; D007150:Immunohistochemistry; D066298:In Vitro Techniques; D008239:Lysine / Q000494:pharmacology; D018613:Microscopy, Confocal; D009543:Nifedipine / Q000494:pharmacology; D009578:Nitrobenzenes / Q000494:pharmacology; D018408:Patch-Clamp Techniques; D011817:Rabbits; D012849:Sinoatrial Node / Q000166:cytology / Q000502:physiology*; D013449:Sulfonamides / Q000494:pharmacology; D000096926:Benzenesulfonamides
chemical_list: D001596:Benzylamines; D002118:Calcium; D002121:Calcium Channel Blockers; D015220:Calcium Channels; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D004533:Egtazic Acid; D004791:Enzyme Inhibitors; D008239:Lysine; D009543:Nifedipine; D009578:Nitrobenzenes; D013449:Sulfonamides; C120345:caged AIP; C072105:KN 93; D000096926:Benzenesulfonamides; C025603:1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid