A Functional Relationship between Specific NOS Isozymes and Beta-adrenergic Recep

描述（由申请人提供）：心脏收缩性的重要调节因子是-肾上腺素能受体（¿AR）信号。1和2受体是在心脏中表达的主要亚型。1受体通过调节l型Ca2+通道、磷蛋白（PLB）和肌钙蛋白I （Tnl）产生正性肌力和弹性效应。2受体的独特之处在于它们仅作为正性肌力发挥作用；这些受体通过调节l型Ca2+通道促进收缩性。AR信号本身可由一氧化氮合成酶（NOS）产生的一氧化氮（NO）调节。在肌细胞内，两种NOS亚型组成性表达：NOS1和NOS3。NOS1信号增强了对非特异性AR刺激的功能性反应，而NOS3信号则降低了这种反应。然而，具体的NOS异构体在调节个体AR受体中的作用仍有待确定。有趣的是，NOS1和¿1受体已被证明通过相似的蛋白质靶点（即plb）调节收缩性，而NOS3和¿2受体通过l型Ca2+通道调节收缩性。最近，NOS3和¿2受体已被证明在肌细胞内划分到相似的位置。因此，我们假设NOS1会增加由于¿1受体激活而不调节¿2受体信号（Aim1）的功能反应。此外，我们假设NOS3会由于¿2受体激活而不调节¿1受体信号（Aim2）而降低功能反应。功能实验（即测量Ca2+瞬态，缩短振幅和l型Ca2+通道电流）和western blotting （plb -磷酸化）来研究这种拟议的功能相互作用，将在暴露于各种AR刺激的野生型（WT，对照），NOS1敲除（KO）和NOS3 KO小鼠的分离肌细胞中进行。我们的研究表明，当AR信号被改变时，这种信号通路会促进收缩功能障碍和心脏重塑，从而与健康相关。因此，确定如何调节AR信号是至关重要的。随着对这一现象的深入了解，我们的研究将使新的治疗方法的发展成为可能。

DESCRIPTION (provided by applicant): An essential regulator of cardiac contractility is ¿-adrenergic receptor (¿AR) signaling. (¿1 and ¿2 receptors are the primary subtypes expressed in the heart. ¿1 receptors produce positive inotropic and lusitropic effects through modulation of the L-type Ca2+ channel, phospholamban (PLB) and troponin I (Tnl). ¿2 receptors are unique in that they only function as positive inotropes; these receptors promote contractility through regulation of the L-type Ca2+ channel. ¿AR signaling itself can be regulated by nitric oxide (NO) produced via NO synthase (NOS). Within the myocyte, two NOS isoforms are constitutively expressed: NOS1 and NOS3. NOS1 signaling has been shown to augment the functional response to non-specific ¿AR stimulation while NOS3 signaling reduces it. However, it remains to be determined what effect specific NOS isoforms have in the regulation of individual ¿AR receptors. Interestingly, NOS1 and ¿1 receptors have been shown to regulate contractility via similar protein targets (i.e.PLB), while NOS3 and ¿2 receptors regulate contractility through the L-type Ca2+ channel. Recently, NOS3 and ¿2 receptors have been shown to compartmentalize to similar locations within the myocyte. Therefore, we hypothesize NOS1 will increase the functional response due to ¿1 receptor activation with no modulation of ¿2 receptor signaling (Aim1). In addition, we hypothesize NOS3 will decrease the functional response due to ¿2 receptor activation with no modulation of ¿1 receptor signaling (Aim2). Functional experiments (i.e. measurement of Ca2+ transient, shortening amplitude, and L-type Ca2+channel current) and western blotting (PLB-phosphorylation) to investigate this proposed functional interaction will be conducted in isolated myocytes from wild type (WT, control), NOS1 knockout (KO) and NOS3 KO mice exposed to various ¿AR stimuli. Our study holds health-relatedness due to the fact that when ¿AR signaling is altered, this signaling pathway promotes contractile dysfunction and cardiac remodeling. Thus, it is crucial to determine how ¿AR signaling is regulated. With a deeper understanding of this phenomena, our study will make possible the development of new therapeutic treatments.