Abstract
Ca2+-activated basal adenylate cyclase (AC) in rabbit sinoatrial node cells (SANC) guarantees, via basal cAMP/PKA-calmodulin/CaMKII-dependent protein phosphorylation, the occurrence of rhythmic, sarcoplasmic-reticulum generated, sub-membrane Ca2+ releases that prompt rhythmic, spontaneous action potentials (APs). This high-throughput signaling consumes ATP.AimsWe have previously demonstrated that basal AC-cAMP/PKA signaling directly, and Ca2+ indirectly, regulate mitochondrial ATP production. While, clearly, Ca2+-calmodulin-CaMKII activity regulates ATP consumption, whether it has a role in the control of ATP production is unknown.Methods and ResultsWe superfused single, isolated rabbit SANC at 37°C with physiological saline containing CaMKII inhibitors, (KN-93 or autocamtide-2 Related Inhibitory Peptide (AIP)), or a calmodulin inhibitor (W-7) and measured cytosolic Ca2+, flavoprotein fluorescence and spontaneous AP firing rate. We measured cAMP, ATP and O2 consumption in cell suspensions. Graded reductions in basal CaMKII activity by KN-93 (0.5–3 µmol/L) or AIP (2–10 µmol/L) markedly slow the kinetics of intracellular Ca2+ cycling, decrease the spontaneous AP firing rate, decrease cAMP, and reduce O2 consumption and flavoprotein fluorescence. In this context of graded reductions in ATP demand, however, ATP also becomes depleted, indicating reduced ATP production.ConclusionsCaMKII signaling, a crucial element of normal automaticity in rabbit SANC, is also involved in SANC bioenergetics.
Highlights
The rate at which the heart beats is governed by the rate at which sinoatrial node cells (SANC) fire spontaneous action potentials (APs)
CAMP-mediated, protein kinase A-dependent (PKA) protein phosphorylation and Ca2+/calmodulin-dependent protein kinase II (CaMKII) protein phosphorylation couple the function of proteins of both clocks to regulate SANC normal automaticity [1,2]. It has been demonstrated in sinoatrial node cells that Ca2+ activated adenylyl cyclase produces a high basal level of cAMP compared to ventricular myocytes [3,4]
To decrease CaMKII activity we chose two concentrations of CaMKII inhibitors that had been previously shown [7] to reduce the AP firing in rabbit SANC rate by,40%, and to eliminate AP firing
Summary
The rate at which the heart beats is governed by the rate at which sinoatrial node cells (SANC) fire spontaneous action potentials (APs). CAMP-mediated, protein kinase A-dependent (PKA) protein phosphorylation and Ca2+/calmodulin-dependent protein kinase II (CaMKII) protein phosphorylation (phospholamban, ryanodine-receptors, L-type channel and etc.) couple the function of proteins of both clocks to regulate SANC normal automaticity [1,2]. It has been demonstrated in sinoatrial node cells that Ca2+ activated adenylyl cyclase produces a high basal level of cAMP compared to ventricular myocytes [3,4]. The level of Ca2+ pumping by SR Ca2+-ATPase is regulated by phospholamban phosporylation of both Ser (PKA) and Thr
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