Laboratory of Cardiovascular Science, Gerontology Research Center, NIA, NIH, 5600 Nathan Shock Dr, Baltimore, MD 21224-6825, USA.
Circ Res. 2010 Sep 17;107(6):767-75. doi: 10.1161/CIRCRESAHA.110.220517. Epub 2010 Jul 22.
Sinoatrial node cells (SANCs) generate local, subsarcolemmal Ca(2+) releases (LCRs) from sarcoplasmic reticulum (SR) during late diastolic depolarization. LCRs activate an inward Na(+)-Ca(2+) exchange current (I(NCX)), which accelerates diastolic depolarization rate, prompting the next action potential (AP). The LCR period, ie, a delay between AP-induced Ca(2+) transient and LCR appearance, defines the time of late diastolic depolarization I(NCX) activation. Mechanisms that control the LCR period, however, are still unidentified.
To determine dependence of the LCR period on SR Ca(2+) refilling kinetics and establish links between regulation of SR Ca(2+) replenishment, LCR period, and spontaneous cycle length.
Spontaneous APs and SR luminal or cytosolic Ca(2+) were recorded using perforated patch and confocal microscopy, respectively. Time to 90% replenishment of SR Ca(2+) following AP-induced Ca(2+) transient was highly correlated with the time to 90% decay of cytosolic Ca(2+) transient (T-90(C)). Local SR Ca(2+) depletions mirror their cytosolic counterparts, LCRs, and occur following SR Ca(2+) refilling. Inhibition of SR Ca(2+) pump by cyclopiazonic acid dose-dependently suppressed spontaneous SANCs firing up to ≈50%. Cyclopiazonic acid and graded changes in phospholamban phosphorylation produced by β-adrenergic receptor stimulation, phosphodiesterase or protein kinase A inhibition shifted T-90(C) and proportionally shifted the LCR period and spontaneous cycle length (R(2)=0.98).
The LCR period, a critical determinant of the spontaneous SANC cycle length, is defined by the rate of SR Ca(2+) replenishment, which is critically dependent on SR pumping rate, Ca(2+) available for pumping, supplied by L-type Ca(2+) channel, and ryanodine receptor Ca(2+) release flux, each of which is modulated by cAMP-mediated protein kinase A-dependent phosphorylation.
窦房结细胞（SANCs）在舒张晚期去极化时从肌浆网（SR）产生局部的、亚肌小节 Ca(2+)释放（LCRs）。LCRs 激活内向 Na(+)-Ca(2+)交换电流（I(NCX)），这加速了舒张期去极化率，促使下一个动作电位（AP）。LCR 期间，即在 AP 诱导的 Ca(2+)瞬变与 LCR 出现之间的延迟，定义了晚期舒张期去极化 I(NCX)激活的时间。然而，控制 LCR 期间的机制仍然未知。
确定 LCR 期间对 SR Ca(2+)再填充动力学的依赖性，并建立 SR Ca(2+)再填充、LCR 期间和自发循环长度之间的调节关系。
使用穿孔贴片和共聚焦显微镜分别记录自发性 AP 和 SR 内腔或胞浆 Ca(2+)。AP 诱导的 Ca(2+)瞬变后 SR Ca(2+)再填充至 90%的时间与胞浆 Ca(2+)瞬变 90%衰减的时间（T-90(C)）高度相关。局部 SR Ca(2+)耗竭与胞浆 Ca(2+)耗竭相对应，发生在 SR Ca(2+)再填充之后。肌浆网 Ca(2+)泵的抑制剂环匹阿尼酸剂量依赖性地抑制自发性 SANCs 放电，最多可达 ≈50%。环匹阿尼酸和β-肾上腺素能受体刺激、磷酸二酯酶或蛋白激酶 A 抑制引起的磷酸化酶蛋白磷酸化的分级变化，改变 T-90(C)，并相应地改变 LCR 期间和自发循环长度（R(2)=0.98）。
LCR 期间是自发性 SANC 循环长度的关键决定因素，由 SR Ca(2+)再填充的速率定义，这与 SR 泵速率、泵入的 Ca(2+)、由 L 型 Ca(2+)通道供应的 Ca(2+)和肌浆网 Ca(2+)释放通量密切相关，这些都受 cAMP 介导的蛋白激酶 A 依赖性磷酸化调节。
