Impaired sinoatrial node function and increased susceptibility to atrial fibrillation in mice lacking natriuretic peptide receptor C

摘要

Natriuretic peptides (NPs) are critical regulators of the cardiovascular system that are currently viewed as possible therapeutic targets for the treatment of heart disease. Recent work demonstrates potent NP effects on cardiac electrophysiology, including in the sinoatrial node (SAN) and atria. NPs elicit their effects via three NP receptors (NPR-A, NPR-B and NPR-C). Among these receptors, NPR-C is poorly understood. Accordingly, the goal of this study was to determine the effects of NPR-C ablation on cardiac structure and arrhythmogenesis. Cardiac structure and function were assessed in wild-type (NPR-C^+/+) and NPR-C knockout (NPR-C^−/−) mice using echocardiography, intracardiac programmed stimulation, patch clamping, high-resolution optical mapping, quantitative polymerase chain reaction and histology. These studies demonstrate that NPR-C^−/− mice display SAN dysfunction, as indicated by a prolongation (30%) of corrected SAN recovery time, as well as an increased susceptibility to atrial fibrillation (6% in NPR-C^+/+ vs. 47% in NPR-C^−/−). There were no differences in SAN or atrial action potential morphology in NPR-C^−/− mice; however, increased atrial arrhythmogenesis in NPR-C^−/− mice was associated with reductions in SAN (20%) and atrial (15%) conduction velocity, as well as increases in expression and deposition of collagen in the atrial myocardium. No differences were seen in ventricular arrhythmogenesis or fibrosis in NPR-C^−/− mice. This study demonstrates that loss of NPR-C results in SAN dysfunction and increased susceptibility to atrial arrhythmias in association with structural remodelling and fibrosis in the atrial myocardium. These findings indicate a critical protective role for NPR-C in the heart.Key points class="unordered" style="list-style-type:disc"> Natriuretic peptides (NPs) elicit their effects via multiple NP receptors (including NPR-A, NPR-B and NPR-C, with NPR-C being relatively poorly understood). We have studied the effects of NPR-C ablation on cardiac structure, function and arrhythmogenesis using NPR-C knockout (NPR-C^−/−) mice. NPR-C^−/− mice are characterized by sinoatrial node (SAN) dysfunction and a profound increase in susceptibility to atrial fibrillation. Increased susceptibility to arrhythmias in NPR-C^−/− mice was associated with slowed electrical conduction in the SAN as well as the right and left atria due to enhanced collagen expression and deposition in the atria (structural remodelling), but without changes in action potential morphology (electrical remodelling) in isolated SAN or atrial myocytes. This study demonstrates a critical protective role for NPR-C in the heart. class="head no_bottom_margin" id="__sec2title">IntroductionNatriuretic peptides (NPs), including atrial (ANP), B-type (BNP) and C-type (CNP) NPs, are a group of powerful cardioprotective hormones that play a critical role in the maintenance of cardiovascular homeostasis in normal conditions and in cardiovascular disease (Levin et al. ; Potter et al. ). In fact, due to their potent effects in the cardiovascular system, NPs are currently in use (nesiritide) (Cataliotti & Burnett, ; Lee & Burnett, ) or in development (CD-NP) (Lee et al. ; Rose, ) for the treatment of heart failure. Despite this, current understanding of the distinct roles of specific NP receptors (NPRs) in the heart is incomplete, which may partially explain the lack of clinical efficacy of some NPs currently in use (Burnett & Korinek, ; O'Connor et al. ).There are three known NPRs denoted NPR-A, NPR-B and NPR-C (Lucas et al. ; Potter et al. ; Rose & Giles, ). Most attention has been given to NPR-A and NPR-B, which are well known particulate guanylyl cyclase (GC) receptors that mediate increases in cGMP upon receptor activation (Potter et al. ). Most physiological effects of NPs have been attributed to these receptors. Conversely, much less is known about NPR-C, which is not directly coupled to GC signalling. NPR-C was originally classified as a ‘clearance receptor’ with no signalling function (Maack et al. ); however, it is now known that NPR-C is able to activate inhibitory G proteins (Gi) and modulate adenylyl cyclase (AC) and phospholipase C signalling (Anand-Srivastava & Trachte, ; Anand-Srivastava, ). This coupling to Gi occurs via specific Gi-activator domains located within the 37 amino acid intracellular portion of NPR-C (Pagano & Anand-Srivastava, ; Zhou & Murthy, href="#b67" rid="b67" class=" bibr popnode tag_hotlink tag_tooltip" id="__tag_434897154">2003). NPR-C is able to bind all NPs with comparable affinity (Anand-Srivastava & Trachte, href="#b2" rid="b2" class=" bibr popnode tag_hotlink tag_tooltip" id="__tag_434897157">1993); thus, a more complete understanding of the role(s) of this receptor in the heart is of critical importance and may impact the future therapeutic use of NPs in heart disease.We have shown that NPs have robust effects on cardiac electrophysiology and provided the first clear evidence that NPR-C can mediate some of these effects (Rose et al. href="#b54" rid="b54" class=" bibr popnode tag_hotlink tag_tooltip" id="__tag_434897038">2003, href="#b55" rid="b55" class=" bibr popnode tag_hotlink tag_tooltip" id="__tag_434897106">2004; Rose & Giles, href="#b53" rid="b53" class=" bibr popnode tag_hotlink tag_tooltip" id="__tag_434897140">2008; Azer et al. href="#b3" rid="b3" class=" bibr popnode tag_hotlink tag_tooltip" id="__tag_434897159">2012, href="#b4" rid="b4" class=" bibr popnode tag_hotlink tag_tooltip" id="__tag_434897115">2014; Springer et al. href="#b60" rid="b60" class=" bibr popnode tag_hotlink tag_tooltip" id="__tag_434897142">2012). Specifically, we have found that in the presence of acute β-adrenergic receptor activation, BNP and CNP can potently reduce heart rate (HR) by decreasing spontaneous action potential (AP) firing in the sinoatrial node (SAN) (Rose et al. href="#b55" rid="b55" class=" bibr popnode tag_hotlink tag_tooltip" id="__tag_434897151">2004; Azer et al. href="#b3" rid="b3" class=" bibr popnode tag_hotlink tag_tooltip" id="__tag_434897053">2012). This occurs in conjunction with a reduction in diastolic depolarization slope (DD slope) and L-type Ca²⁺ current in SAN myocytes. We have also demonstrated, using high-resolution optical mapping, that NPs can slow conduction in the SAN and atrial myocardium by activating NPR-C (Azer et al. href="#b4" rid="b4" class=" bibr popnode tag_hotlink tag_tooltip" id="__tag_434897044">2014). These inhibitory effects of NPs on HR, SAN AP firing and SAN/atrial conduction are completely absent in NPR-C knockout mice (NPR-C^−/−) (Azer et al. href="#b3" rid="b3" class=" bibr popnode tag_hotlink tag_tooltip" id="__tag_434897035">2012, href="#b4" rid="b4" class=" bibr popnode tag_hotlink tag_tooltip" id="__tag_434897125">2014). These observations conclusively demonstrate that NPR-C is an important mediator of electrophysiological effects of NPs in the heart, including in the SAN and atria.Although our previous work demonstrates the involvement of NPR-C in mediating acute electrophysiological effects of NPs in the heart, the consequences of NPR-C ablation on cardiac structure and arrhythmogenesis have not been investigated. This constitutes a clear gap in our understanding of this receptor and of the physiology of NPs in the heart. Accordingly, the goal of the present study was to determine the consequences of the loss of NPR-C in the heart using NPR-C^−/− mice. Our novel data demonstrate that NPR-C^−/− mice are characterized by SAN dysfunction and increased susceptibility to atrial fibrillation (AF) in association with impaired electrical conduction and enhanced atrial fibrosis (structural remodelling). These findings indicate that NPR-C plays a critical protective role in the heart by preventing adverse structural remodelling due to fibrosis.