Decreased pacemaker activity in aged sinoatrial node

心脏变时性和变力性的一般理论如何应用于衰老？ 我们早期和最近的研究都为理解人类和动物模型中与年龄相关的心脏收缩力和心率恶化提供了开创性的观点。我们对巴尔的摩老龄化纵向研究（BLSA）中的健康参与者进行的研究表明，在对分级运动应激做出反应时，心率增加和收缩末期容积减少的能力发生了实质性的年龄相关变化。(Of注意，只有后者，而不是前者，可以通过物理调节来改善。 我们随后证明，人类收缩力和心率的缺陷部分是由于cAMP-PKA的AR刺激反应降低。 1.收缩力 在离体大鼠心室肌中，我们直接证明了对AR刺激的Ca 2+循环和收缩反应的减少。在没有-AR刺激的情况下，Ca 2+瞬变和收缩的松弛时间延长，我们追踪到SERCA 2的表达减少。AP也显着延长，由于年龄相关的变化，L-型Ca 2+和K+电流。在单个VM中，除了表面膜离子通道的许多特性之外，我们还记录了VM Ca 2+时钟中与年龄相关的减少，表现为兴奋-Ca 2+释放-收缩耦合过程恢复的时间延长，这是由于响应于L型Ca 2+电流激活的SR Ca 2+释放通过RyRs的恢复时间延长。我们还表明，SR的Ca 2+负荷和Ca 2+瞬变的幅度保存在心肌细胞从旧的心脏，凭借延长AP。 2.心率 我们在兔SANC中发现了通过耦合时钟系统进行心率调节，最终（25年后）在阐明与年龄相关的变时储备减少的潜在细胞机制方面取得了进展。具体来说，我们的概念突破，即基础起搏细胞自动性的调节需要由PDE和磷酸酶活性调节的Ca-PKA-CaMK II信号传导，而受体的刺激只是延长了这种调节，这使我们假设，参与调节固有心率的机制随着年龄的增长而恶化，而调节心率储备的机制可能涉及减慢心率。在某些方面，至少与VM中调节Ca 2+循环和收缩性的机制的年龄相关性恶化有关。 磷蛋白（PLB）磷酸化Ser 16（PLB/总PLB）的免疫标记在控制没有不同的年龄，增加2.2 ± 0.2倍后，孵育与10 M IBMX在年轻的，但没有增加在老SANC。 内在SR Ca 2+循环及其对PDE抑制的响应随年龄的增长而下降，并且是至少部分地解释IHR年龄相关性降低的候选机制。五年来，我们一直试图分离出表现出稳定正常自律性的单个小鼠窦房结细胞，但都失败了。最近，我们取得了一些成功。我们的初步研究结果表明：自发AP放电率的单一SANC下降，随着年龄的增长，最大RyR钙释放通量响应AP（索引为d Ca/dt最大）下降，随着年龄的增长和动力学的Ca 2+瞬态松弛变得减少。 提示老年小鼠内源性cAMP-PKA-Ca ~（2+）信号通路存在缺陷。这种缺陷可能是由于：1）。Ca 2+循环蛋白质例如SR Ca 2+泵、受磷蛋白（PLB）、RyR的量或功能降低。和/或2.）响应cAMP介导的PKA依赖性磷酸化的增加，降低Ca 2+循环蛋白例如PLB的磷酸化。3.）第三章这些缺陷可以解释为什么老年人的心脏不能像年轻人的心脏跳动得那么快。今后的计划是：1）在来自年轻和老年SAN的细胞中采用蛋白质印迹法来测量Ca 2+循环蛋白密度; 2）直接测量cAMP-PKA-Ca 2+途径的下游信号传导，即PKA依赖的蛋白磷酸化和磷酸酶活性。

How does the General Theory of Cardiac Chronotropy and Inotropy Apply to Aging? Both our earlier and very recent studies have contributed seminal perspectives toward understanding age-associated deterioration of both cardiac contractility and heart rate in both humans, and in animal models. Our studies of healthy participants in the Baltimore Longitudinal Study of Aging (BLSA) demonstrated substantial age-associated changes in the ability to increase heart rate and reduce end systolic volume in response to graded exercise stress. (Of note, only the latter, but not the former, can be improved by physical conditioning.) We subsequently demonstrated that deficits both in contractility and heart rate in humans are due, in part, to reduced -AR stimulation response of cAMP-PKA. 1. Contractility In isolated rat cardiac ventricular muscle, we had directly demonstrated a reduction in the Ca2+ cycling and contractile response to -AR stimulation. The relaxation time of the Ca2+ transient and contraction in the absence of -AR stimulation were prolonged, that we traced to a reduced expression of SERCA2. The AP was also markedly prolonged, due to age-associated changes in L-type Ca2+ and K+ currents. In single VM, in addition to numerous properties of surface membrane ion channels, we documented an age-associated reduction in the VM Ca2+ clock, manifest as a prolonged time for restitution of the excitation- Ca2+ release-contraction coupling process, was due to a prolonged restitution time for SR Ca2+ release via RyRs in response to activation by an L-type Ca2+ current. We also showed that SR Ca2+ loading and the amplitude of the Ca2+ transient were preserved in myocytes from the old heart, by virtue of the prolonged AP. 2. Heart rate Our discovery of heart rate regulation by a coupled-clock system in rabbit SANC enabled progress, finally (after 25 years), on the elucidation of potential cellular mechanisms of the age-associated reduction in chronotropic reserve. Specifically, our conceptual breakthrough that regulation of basal pacemaker cell automaticity requires Ca-PKA-CaMKII signaling regulated by PDE and phosphatase activities, and that stimulation of receptors merely extended this regulation, led us to hypothesize that mechanisms involved in the deterioration of mechanisms that regulate of intrinsic heart rate that accompanies advancing age and that those regulating heart rate reserve may involve a slowing of the Ca2+ clock of SANC and linked, in some respects, at least, to the age-associated deterioration of mechanisms that regulate Ca2+ cycling and contractility in VM. Phospholamban (PLB) phosphorylation at Ser16 (PLB/total PLB) immunolabeling in control did not differ with age, increased 2.20.2 fold after incubation with 10 M IBMX in young, but did not increase in old SANC. The intrinsic SR Ca2+ cycling and its response to PDE inhibition decline with aging, and are candidate mechanisms to explain, in part at least, the age-associated decrease in IHR. We had failed for five years in our attempts to isolate single mouse sinoatrial node cells that exhibited stable normal automaticity. Recently, we have met with some success. Our initial results indicate that: the spontaneous AP firing rate of single SANC declines with age; the maximum RyR Ca release flux in response to an AP (indexed as d Ca/dt max) declines with age; and the kinetics of relaxation of the Ca2+ transient become reduced with age. Our results suggest that the intrinsic cAMP-PKA-Ca2+ signaling is deficient in the aged mice. This deficiency may result from: 1.)reduced amount or function of Ca2+ cycling proteins e.g. SR Ca2+ pump, phospholamban (PLB), RyRs. and/or 2.) reduced phosphorylation of Ca2+ cycling proteins e.g.PLB, in response to an increase in cAMP-mediated-PKA dependent phosphorylation. 3.) These deficiencies may explain why the aged heart cannot beat as fast as the young heart. Future plans are: 1) to employ western blotting in cells from young and old SANs to measure Ca2+ cycling protein density; 2) to directly measure the downstream signaling of cAMP-PKA- Ca2+ pathway i.e. PKA-dependent protein phosphorylation and phosphatase activity.