Myocardial Nitric Oxide Content Research Articles

Inhibition of Rho-kinase by fasudil restores the cardioprotection of ischemic postconditioninng in hypercholesterolemic rat heart.

Ischemic postconditioning (IPoC) reduces lethal reperfusion injury under normal conditions, but its effectiveness is blocked by hypercholesterolemia. The present study aimed to determine whether the inhibition of Rho‑kinase by fasudil restores the cardioprotection of IPoC in the hypercholesterolemic rat heart, and to elucidate the potential mechanisms underlying this process. The isolated rat hearts underwent 30min global ischemia and 120min reperfusion. IPoC was induced by six cycles of 10sec ischemia and 10sec reperfusion at the onset of the reperfusion. Fasudil was administered 15min prior to ischemia, and wortmannin and L‑NAME were administered following IPoC. The myocardial infarct size, apoptosis, myocardial nitric oxide (NO) content and Rho‑kinase activity, as well as the activation of the phosphatidylinositol 3‑kinase/Akt/endothelial nitric oxide synthase (PI3K/Akt/eNOS) pathway, were examined. The results revealed that IPoC and 1µM fasudil treatment alone failed to reduce the infarct size and apoptosis rate. However, IPoC combined with 1µM fasudil treatment or 10µM fasudil treatment alone restored the cardioprotection as evidenced by the decreasing in infarct size and apoptosis rate, whereas it was blocked by the administration of wortmannin or L‑NAME. Furthermore, IPoC combined with 1µM fasudil treatment also enhanced the phosphorylation of Akt and eNOS and conferred a significant increase in the content of NO. By contrast, no significant improvements were demonstrated in the phosphorylation of Akt and eNOS, as well as myocardial NO content when treated with 1µM fasudil and IPoC alone. The inhibition of Rho‑kinase by fasudil was able to restore the cardioprotection of IPoC in the hypercholesterolemic rat heart. The underlying mechanisms involved in this process appear to be mediated by the activation of the PI3K/Akt/eNOS signal pathway and an increase in the myocardial NO content.

Protective effect of picroside II on myocardial ischemia reperfusion injury in rats

The aim of this study was to determine the effect of picroside II on myocardial ischemia reperfusion injury in rats and to explore its underlying mechanism. Isolated rat hearts underwent 30 minutes of global ischemia followed by 120 minutes of reperfusion. Different doses of picroside II (1 μM, 10 μM, and 100 μM) were given 20 minutes before ischemia. Phosphoinositide 3-kinase inhibitor (wortmannin) and nitric oxide synthase (NOS) inhibitor (L-NG-nitroarginine methyl ester) were given 10 minutes before picroside II treatment. The cardiac function, myocardial infarct size, apoptosis, myocardial nitric oxide content, the expressions of Bcl-2 and Bax, and the activation of the phosphoinositide 3-kinase/Akt/endothelial NOS pathway were evaluated. Treatment with 10 μM and 100 μM picroside II significantly improved postischemic myocardial function, reduced myocardial infarct size, inhibited apoptosis, increased myocardial NO content, upregulated Bcl-2, downregulated Bax, and increased the phosphorylation of Akt and endothelial NOS, but cardioprotection was not shown in the 1 μM picroside II treatment group and was abrogated by wortmannin and L-NG-nitroarginine methyl ester. Furthermore, cardioprotection in the 100 μM picroside II treatment group was superior to that in the 10 μM picroside II treatment group. In conclusion, the data reveals that picroside II has a significant protective effect on myocardial ischemia reperfusion injury in a dose-dependent manner, which was mediated by upregulating the phosphoinositide 3-kinase/Akt/endothelial NOS pathway to increase nitric oxide production and regulating the expressions of Bcl-2 and Bax to inhibit apoptosis.

NITRIC OXIDE TRIGGERS DELAYED ANESTHETIC PRECONDITIONING-INDUCED CARDIAC PROTECTION VIA ACTIVATION OF NUCLEAR FACTOR-κB AND UPREGULATION OF INDUCIBLE NITRIC OXIDE SYNTHASE

Nitric oxide (NO) plays a pivotal role both in triggering and mediating delayed protection against myocardial I/R injury during anesthetic-induced preconditioning (APC). However, the signaling mechanisms that underlie this phenomenon remain unclear. Using isoflurane as a representative anesthetic, the present study tested the hypothesis that NO released after anesthetic-induced preconditioning initiates delayed cardioprotection via activation of nuclear transcription factor-kappaB (NF-kappaB), leading to myocardial adaptation by upregulation of iNOS and increase in production of NO. Sprague-Dawley rats that received open-chest surgery under pentobarbital anesthesia were subject to 30 min of left coronary artery occlusion, followed by 120 min of reperfusion. Exposure to 60 min of 2.1% isoflurane inhalation with oxygen 24 h before ischemia significantly reduced I/R-induced myocardial infarct size that was associated with overexpression of iNOS protein and increased NO content in the heart. These protective effects were abolished by pretreatment with a NOS inhibitor, N-nitro-L-arginine methyl ester, an NF-kappaB blocker, diethyldithiocarbamate, before isoflurane, or a selective iNOS inhibitor, S-methylisothiourea, before left coronary artery occlusion. Isoflurane exposure also evoked a robust increase in myocardial NO content, followed by nucleus-bound translocation of p65 or p50 subunit of NF-kappaB and increase in NF-kappaB DNA-binding activity in heart tissues. These molecular events after isoflurane exposure were blocked by pretreatment with N-nitro-L-arginine methyl ester. We conclude that NO generated immediately after isoflurane exposure triggers downstream activation of NF-kappaB, resulting in subsequent upregulation of iNOS expression and NO synthesis that mediate APC-induced delayed cardioprotection.

Nitric oxide modifies the sarcoplasmic reticular calcium release channel in endotoxemia by both guanosine-3′,5′ (cyclic) phosphate-dependent and independent pathways*

a) To determine whether decreased sarcoplasmic calcium release channel (CRC) activity is a mechanism by which myocardial contractility is reduced in endotoxemia; b) to determine whether nitric oxide modulates CRC activity in endotoxemia; and c) to examine two nitric oxide signaling pathways in relation to CRC function in endotoxemia. Randomized, prospective using a rat model of endotoxemia. : Research laboratory. Sprague-Dawley rats. Endotoxemia was induced by lipopolysaccharide administration. The effects of nitric oxide were studied using the highly selective inducible nitric oxide synthase inhibitor N-(3-(aminomethyl)benzyl)acetamidine dihydrochloride (1400W) and the specific guanylyl cyclase inhibitor 1-H (1, 2, 4)oxadiazolo[4,3-a]quinoxalin-1-one (ODQ). We assessed myocardial contractility, myocardial nitric oxide content, and guanosine-3',5' (cyclic) phosphate (cGMP) content. We determined CRC activity by calcium release and ryanodine binding assays. We followed these variables at four time points through the course of endotoxemia. We found that myocardial contractility and CRC activity were decreased in late but not in early endotoxemia. Furthermore, inducible nitric oxide synthase inhibition with 1400W restored contractility and CRC activity in late endotoxemia but paradoxically worsened these variables in early endotoxemia. Through the use of the guanylyl cyclase inhibitor ODQ, we demonstrate that nitric oxide acts through cGMP-mediated mechanisms in early and late endotoxemia. We investigated cGMP-independent pathways by assessing the oxidative status of the CRC. We found that in late endotoxemia, nitric oxide decreased the number of free thiols, demonstrating that nitric oxide also acts through cGMP-independent pathways. Nitric oxide has a dual effect on the CRC in endotoxemia. At low concentrations, as measured in early endotoxemia, nitric oxide stabilizes the CRC through cGMP-mediated mechanisms. In late endotoxemia, high nitric oxide concentrations decrease channel activity through both cGMP-dependent and cGMP-independent mechanisms.

Capsaicin-sensitive sensory neurons regulate myocardial nitric oxide and cGMP signaling

We studied whether tissue levels of nitric oxide (NO) and cGMP are regulated by sensory nerves in normoxic and ischemic hearts. Wistar rats were treated with capsaicin to deplete neurotransmitters from capsaicin-sensitive sensory nerves. In separate experiments, capsaicin was applied perineurally to both vagus nerves for selective chemodenervation of vagal cardiac afferent nerves. Systemic capsaicin administration significantly decreased basal myocardial NO content assessed by electron spin resonance (ESR) spectroscopy, whereas, local treatment of vagus nerves did not change it. Both systemic and local capsaicin treatment decreased cardiac cGMP content measured by radioimmunoassay. In separate experiments, isolated hearts from control and systemic capsaicin-treated rats were subjected to 30-min global ischemia. NO signal intensity increased 10-fold after ischemia, whereas, cardiac cGMP decreased. Capsaicin pretreatment did not influence ischemic NO or cGMP content. These results suggest a major role for capsaicin-sensitive sensory neurons in the maintenance of basal but not ischemic myocardial NO and cGMP content. Vagal sensory nerves may be involved in the regulation of basal myocardial cGMP but not basal NO level. Consequently, basal NO content in the heart is regulated primarily by spinal afferent nerves.

Time course of nitric oxide, peroxynitrite, and antioxidants in the endotoxemic heart

To determine the time course for myocardial production of nitric oxide, peroxynitrite, and glutathione, to determine the activities of the myocardial antioxidant enzymes glutathione peroxidase, superoxide dismutase, and glutathione reductase throughout endotoxemia and into recovery, and to correlate the levels of these variables to left ventricular contractility in endotoxemia. Rats were treated with lipopolysaccharide. Endotoxemic hearts were examined at baseline, 4, 16, 24, and 48 hrs after lipopolysaccharide. Saline time-control groups were treated identically. A pulmonary research laboratory of a university teaching hospital. Lipopolysaccharide administration resulted in decreased contractility at 16 hrs as assessed by the isolated papillary muscle technique. Contractility recovered by 24 hrs. Myocardial glutathione content initially increased, but it was decreased from baseline by 16 hrs, as was glutathione peroxidase activity. Both superoxide dismutase and glutathione reductase activities were increased early (4 hrs) and remained elevated throughout the course of the experiment. Myocardial nitric oxide content (assessed by the chemiluminescence technique) was increased by 4 hrs and was markedly elevated by 16 hrs. Nitric oxide levels remained elevated despite recovery of contractility at 24 hrs. Similarly, peroxynitrite (assessed by measurement of 3-nitrotyrosine by high-pressure liquid chromatography) was elevated at 16 hrs and remained elevated despite normalization of contractility at 24 and 48 hrs. Myocardial dysfunction in endotoxemia correlates mainly with decreased glutathione content and glutathione peroxidase activity rather than nitric oxide or peroxynitrite formation. These data indicate that lipopolysaccharide-induced myocardial dysfunction is not solely caused by elevated myocardial nitric oxide levels but rather caused by the sum of complex interactions between various oxygen- and nitrogen-derived radicals.

Nitric Oxide and Cardiac Contractility in Human Heart Failure

The free radical gas nitric oxide (NO) is produced by 3 isoforms of nitric oxide synthase (NOS). All of them are present in the heart: NOS1 (nNOS, “neuronal” NOS) has been detected in cardiac conduction tissue and intracardiac neurons; NOS2 (iNOS, “cytokine-inducible” NOS) can be expressed by virtually all cells in the heart, often in conjunction with the expression of inflammatory cytokines; and finally, NOS3 (eNOS, “endothelial-constitutive” NOS) is expressed in coronary endothelium, endocardium, and cardiac myocytes. NOS3 regulates the tone of vascular smooth muscle cells; the permeability and platelet adhesion of endothelial cells; and the receptor-effector coupling, energetics, contractility, and apoptosis of cardiomyocytes.1,2 Since the original demonstration of myocardial NOS2 activity in idiopathic dilated cardiomyopathy,3 a negative inotropic effect of NO frequently was hypothesized to contribute to the depressed contractile function of failing myocardium. This hypothesis was inspired by the simultaneous publication of experimental results that showed a depressed contractile response of isolated cardiomyocytes to β-adrenergic agonists after NOS2 induction by lipopolysaccharides.4 See p 2318 To further explore this depressant action of NO on myocardial contractility, numerous experimental and clinical studies were performed, but they yielded apparently conflicting results on the inotropic effect of NO from either endogenous or pharmacological sources.2 As reported in this issue of Circulation , Cotton et al5 tackled the question of the inotropic action of NO by measuring left ventricular (LV) performance in normal control subjects and in patients with dilated cardiomyopathy during intracoronary infusion of the NOS inhibitor NG-monomethyl-l-arginine (L-NMMA). During L-NMMA infusion, Cotton et al5 observed a modest drop (14%) in LV dP/dtmax in the control group and no change in LV dP/dtmax in the cardiomyopathy group, despite myocardial expression of NOS2 in their patients. They concluded that the small …

Selective NOS Inhibition Restores Myocardial Contractility in Endotoxemic Rats; However, Myocardial NO Content Does Not Correlate with Myocardial Dysfunction

The role of nitric oxide (NO) in lipopolysaccharide (LPS)-induced myocardial dysfunction remains controversial as some investigators concluded that inhibition of NO synthesis improves left ventricular (LV) contractility, whereas others did not. We investigated the relationship between LPS-induced LV dysfunction and LV NO production. We postulated that high myocardial NO concentrations would correspond to decreased contractility and low NO concentrations would correspond to recovery. In a rat model of endotoxemia, we used the isolated papillary preparation to assess inotropic dysfunction. We measured LV NO content and hemodynamics at baseline, 4, 16, and 48 h after LPS administration. LPS caused a decrease in LV contractility at 16 h with recovery at 48 h. Myocardial NO levels were elevated at all time periods. However, at 48 h in spite of normalization of LV contractility, myocardial NO content remained elevated. Pretreatment of LPS animals with the nonselective nitric oxide synthase (NOS) inhibitor N (G)-nitro-L-arginine methyl ester (L-NAME) worsened LV contractility, decreased LV NO content, and increased mortality. However, pretreatment with the relatively selective inducible NOS (iNOS) inhibitor S-methylisothiourea sulfate (SMT) restored LV contractility. Myocardial NO content in the SMT was lower than that of the LPS only group, but higher than the L-NAME group. We conclude that SMT is beneficial to myocardial contractility in this model of endotoxemia, whereas pretreatment with L-NAME is associated with further deterioration of contractility and increased mortality. Moreover, our data indicate that high myocardial NO concentrations do not necessarily correlate with decreased contractility.