Background-Heart failure is associated with neurological deficits, including cognitive dysfunction. However, the molecular mechanisms underlying reduced cerebral blood flow in the early stages of heart failure, particularly when blood pressure is minimally affected, are not known. Methods and Results-Using a myocardial infarction model in mice, we demonstrate a tumor necrosis factor-␣ (TNF␣)-dependent enhancement of posterior cerebral artery tone that reduces cerebral blood flow before any overt changes in brain structure and function. TNF␣ expression is increased in mouse posterior cerebral artery smooth muscle cells at 6 weeks after myocardial infarction. Coordinately, isolated posterior cerebral arteries display augmented myogenic tone, which can be fully reversed in vitro by the competitive TNF␣ antagonist etanercept. TNF␣ mediates its effect via a sphingosine-1-phosphate (S1P)-dependent mechanism, requiring sphingosine kinase 1 and the S1P 2 receptor. In vivo, sphingosine kinase 1 deletion prevents and etanercept (2-week treatment initiated 6 weeks after myocardial infarction) reverses the reduction of cerebral blood flow, without improving cardiac function. Conclusions-Cerebral artery vasoconstriction and decreased cerebral blood flow occur early in an animal model of heart failure; these perturbations are reversed by interrupting TNF␣/S1P signaling. This signaling pathway may represent a potential therapeutic target to improve cognitive function in heart failure.

Abstract-Heart rate is controlled by the opposing activities of sympathetic and parasympathetic inputs to pacemaker myocytes in the sinoatrial node (SAN). Parasympathetic activity on nodal myocytes is mediated by acetylcholinedependent stimulation of M 2 muscarinic receptors and activation of G␣ i/o signaling. Although regulators of G protein signaling (RGS) proteins are potent inhibitors of G␣ i/o signaling in many tissues, the RGS protein(s) that regulate parasympathetic tone in the SAN are unknown. Our results demonstrate that RGS4 mRNA levels are higher in the SAN compared to right atrium. Conscious freely moving RGS4-null mice showed increased bradycardic responses to parasympathetic agonists compared to wild-type animals. Moreover, anesthetized RGS4-null mice had lower baseline heart rates and greater heart rate increases following atropine administration. Retrograde-perfused hearts from RGS4-null mice showed enhanced negative chronotropic responses to carbachol, whereas SAN myocytes showed greater sensitivity to carbachol-mediated reduction in the action potential firing rate. Finally, RGS4-null SAN cells showed decreased levels of G protein-coupled inward rectifying potassium (GIRK) channel desensitization and altered modulation of acetylcholine-sensitive potassium current (I KACh ) kinetics following carbachol stimulation. Taken together, our studies establish that RGS4 plays an important role in regulating sinus rhythm by inhibiting parasympathetic signaling and I KACh activity. (Circ Res. 2008;103:527-535.)Key Words: RGS proteins Ⅲ sinoatrial node Ⅲ parasympathetic Ⅲ GIRK channels H eart rate (HR) regulation by the autonomic nervous system is integrated by specialized autorhythmic (pacemaker) cells located within the sinoatrial node (SAN). Sympathetic neurotransmitters work via G s -coupled ␤-adrenergic receptors to increase adenylyl cyclase activity, intracellular cAMP concentration and protein kinase A activity. As a result, cAMP-regulated effectors such as hyperpolarizationactivated cyclic nucleotide-gated cation (HCN) channels, delayed rectifier, and voltage-gated Ca 2ϩ channels are enlisted by sympathetic activity to increase pacemaker cell firing rate. 1,2 By contrast, vagal parasympathetic activity decreases HR via G␣ i/o -coupled cholinergic M 2 muscarinic receptors (M 2 Rs). Several effects, mediated by both G␣ i/o and G␤␥ subunits, may contribute to this reduction in HR. G␤␥ heterodimers directly activate G protein-coupled inward rectifying potassium (GIRK) channels, resulting in membrane hyperpolarization. By contrast, G␣ i/o can both modulate phosphodiesterase activity and inhibit adenylyl cyclase activity, reduce both intracellular cAMP levels and protein kinase A activity, leading to decreased depolarizing currents carried by HCN and L-type calcium channels. [2][3][4][5] Because dysregulation of parasympathetic activity occurs in heart failure, sick sinus syndrome, and selected cardiac arrhythmias, 6 it is of clinical interest to identify key molecular regulators of parasympathetic signa...

Visual Abstract

ObjectivesGlucagon-like peptide-1 (GLP-1) is secreted from enteroendocrine cells and exerts a broad number of metabolic actions through activation of a single GLP-1 receptor (GLP-1R). The cardiovascular actions of GLP-1 have garnered increasing attention as GLP-1R agonists are used to treat human subjects with diabetes and obesity that may be at increased risk for development of heart disease. Here we studied mechanisms linking GLP-1R activation to control of heart rate (HR) in mice.MethodsThe actions of GLP-1R agonists were examined on the control of HR in wild type mice (WT) and in mice with cardiomyocyte-selective disruption of the GLP-1R (Glp1rCM−/−). Complimentary studies examined the effects of GLP-1R agonists in mice co-administered propranolol or atropine. The direct effects of GLP-1R agonism on HR and ventricular developed pressure were examined in isolated perfused mouse hearts ex vivo, and atrial depolarization was quantified in mouse hearts following direct application of liraglutide to perfused atrial preparations ex vivo.ResultsDoses of liraglutide and lixisenatide that were equipotent for acute glucose control rapidly increased HR in WT and Glp1rCM−/− mice in vivo. The actions of liraglutide to increase HR were more sustained relative to lixisenatide, and diminished in Glp1rCM−/− mice. The acute chronotropic actions of GLP-1R agonists were attenuated by propranolol but not atropine. Neither native GLP-1 nor lixisenatide increased HR or developed pressure in perfused hearts ex vivo. Moreover, liraglutide had no direct effect on sinoatrial node firing rate in mouse atrial preparations ex vivo. Despite co-localization of HCN4 and GLP-1R in primate hearts, HCN4-directed Cre expression did not attenuate levels of Glp1r mRNA transcripts, but did reduce atrial Gcgr expression in the mouse heart.ConclusionsGLP-1R agonists increase HR through multiple mechanisms, including regulation of autonomic nervous system function, and activation of the atrial GLP-1R. Surprisingly, the isolated atrial GLP-1R does not transduce a direct chronotropic effect following exposure to GLP-1R agonists in the intact heart, or isolated atrium, ex vivo. Hence, cardiac GLP-1R circuits controlling HR require neural inputs and do not function in a heart-autonomous manner.

Tumour necrosis factor (TNF) is a ubiquitously expressed cytokine with functions beyond the immune system. In several diseases, the induction of TNF expression in resistance artery smooth muscle cells enhances microvascular myogenic vasoconstriction and perturbs blood flow. This pathological role prompted our hypothesis that constitutively expressed TNF regulates myogenic signalling and systemic haemodynamics under non-pathological settings. Here we show that acutely deleting the TNF gene in smooth muscle cells or pharmacologically scavenging TNF with etanercept (ETN) reduces blood pressure and resistance artery myogenic responsiveness; the latter effect is conserved across five species, including humans. Changes in transmural pressure are transduced into intracellular signals by membrane-bound TNF (mTNF) that connect to a canonical myogenic signalling pathway. Our data positions mTNF ‘reverse signalling' as an integral element of a microvascular mechanosensor; pathologic or therapeutic perturbations of TNF signalling, therefore, necessarily affect microvascular tone and systemic haemodynamics.

No abstract

Acute β-blockade with metoprolol has been associated with increased mortality by undefined mechanisms. Since metoprolol is a relatively high affinity blocker of β(2)-adrenoreceptors, we hypothesized that some of the increased mortality associated with its use may be due to its abrogation of β(2)-adrenoreceptor-mediated vasodilation of microvessels in different vascular beds. Cardiac output (CO; pressure volume loops), mean arterial pressure (MAP), relative cerebral blood flow (rCBF; laser Doppler), and microvascular brain tissue Po(2) (G2 oxyphor) were measured in anesthetized mice before and after acute treatment with metoprolol (3 mg/kg iv). The vasodilatory dose responses to β-adrenergic agonists (isoproterenol and clenbuterol), and the myogenic response, were assessed in isolated mesenteric resistance arteries (MRAs; ∼200-μm diameter) and posterior cerebral arteries (PCAs ∼150-μm diameter). Data are presented as means ± SE with statistical significance applied at P < 0.05. Metoprolol treatment did not effect MAP but reduced heart rate and stroke volume, CO, rCBF, and brain microvascular Po(2), while concurrently increasing systemic vascular resistance (P < 0.05 for all). In isolated MRAs, metoprolol did not affect basal artery tone or the myogenic response, but it did cause a dose-dependent impairment of isoproterenol- and clenbuterol-induced vasodilation. In isolated PCAs, metoprolol (50 μM) impaired maximal vasodilation in response to isoproterenol. These data support the hypothesis that acute administration of metoprolol can reduce tissue oxygen delivery by impairing the vasodilatory response to β(2)-adrenergic agonists. This mechanism may contribute to the observed increase in mortality associated with acute administration of metoprolol in perioperative patients.

Our data establish CNP/NPR2 signaling as a novel regulator of aortic valve development and disease and elucidate the therapeutic potential of targeting this pathway to arrest disease progression.