Receptors In Ventricular Myocytes Research Articles

Ros and CaMKII Modulate Sodium Overload-Induced Calcium Leak from Ryanodine Receptors in Ventricular Myocytes

Introduction: Elevation of the diastolic free calcium concentration ([Ca2+]cyt) alters excitation-contraction coupling and contractility in cardiac myocytes. It is known that Na+ overload increases the levels of diastolic [Ca2+]cyt and induces arrhythmias via reverse mode of Na+/ Ca2+ exchanger. We hypothesized that Ca2+ leak induced by oxidation/phosphorylation of ryanodine receptors (RyRs) contributes to the rise in [Ca2+]cyt during Na+ overload. Methods: We used confocal fluorescence imaging of intact and membrane-permeabilized isolated rabbit and rat ventricular myocytes to study Ca2+ transients, cytosolic sodium ([Na+]cyt ) and intracellular reactive oxygen species (ROS) levels. Cellular Na+ overload was induced using 5 nM anemone toxin-II (ATX-II), a late Na+ current (INaL) enhancer. Results: ATX-II-induced [Na+]cyt overload caused abnormal Ca2+ transients in intact rabbit and rat myocytes. Additionally, high cytosolic Na+ increased ROS levels and activated CaMKII. These effects were abolished by the INa inhibition with ranolazine and TTX, CaMKII inhibition with KN93 and AIP or by the application of antioxidants such as DTT and coenzyme Q10. In the experiments with membrane-permeabilized myocytes, rapid elevation of [Na+]cyt from 5 mM to 20 mM resulted in an increase of ROS production in mitochondria and Ca2+ leak from RyRs by measuring the frequency of spontaneous Ca2+ waves. The CaMKII inhibitors, KN-93 and AIP, attenuated the effects of high [Na+]cyt on Ca2+ signaling. Conclusions: These data suggest that [Na+]cyt regulates RyRs activity. An increase in [Na+]cyt induces ROS production; ROS activates CaMKII leading to altered Ca2+ transients and abnormal Ca2+ handling.

Effects of anandamide on potassium channels in rat ventricular myocytes: a suppression of Ito and augmentation of KATP channels

Anandamide is an endocannabinoid that has antiarrhythmic effects through inhibition of L-type Ca(2+) channels in cardiomyocytes. In this study, we investigated the electrophysiological effects of anandamide on K(+) channels in rat ventricular myocytes. Whole cell patch-clamp technique was used to record K(+) currents, including transient outward potassium current (I(to)), steady-state outward potassium current (I(ss)), inward rectifier potassium current (I(K1)), and ATP-sensitive potassium current (I(KATP)) in isolated rat cardiac ventricular myocytes. Anandamide decreased I(to) while increasing I(KATP) in a concentration-dependent manner but had no effect on I(ss) and I(K1) in isolated ventricular myocytes. Furthermore, anandamide shifted steady-state inactivation curve of I(to) to the left and shifted the recovery curve of I(to) to the right. However, neither cannabinoid 1 (CB(1)) receptor antagonist AM251 nor CB(2) receptor antagonist AM630 eliminated the inhibitory effect of anandamide on I(to). In addition, blockade of CB(2) receptors, but not CB(1) receptors, eliminated the augmentation effect of anandamide on I(KATP). These data suggest that anandamide suppresses I(to) through a non-CB(1) and non-CB(2) receptor-mediated pathway while augmenting I(KATP) through CB(2) receptors in ventricular myocytes.

Pressure overload-induced hypertrophy in transgenic mice selectively overexpressing AT2 receptors in ventricular myocytes

The role of the angiotensin II type 2 (AT2) receptor in cardiac hypertrophy remains controversial. We studied the effects of AT2 receptors on chronic pressure overload-induced cardiac hypertrophy in transgenic mice selectively overexpressing AT2 receptors in ventricular myocytes. Left ventricular (LV) hypertrophy was induced by ascending aorta banding (AS). Transgenic mice overexpressing AT2 (AT2TG-AS) and nontransgenic mice (NTG-AS) were studied after 70 days of aortic banding. Nonbanded NTG mice were used as controls. LV function was determined by catheterization via LV puncture and cardiac magnetic resonance imaging. LV myocyte diameter and interstitial collagen were determined by confocal microscopy. Atrial natriuretic polypeptide (ANP) and brain natriuretic peptide (BNP) were analyzed by Northern blot. Sarco(endo)plasmic reticulum Ca2+-ATPase (SERCA)2, inducible nitric oxide synthase (iNOS), endothelial NOS, ERK1/2, p70S6K, Src-homology 2 domain-containing protein tyrosine phosphatase-1, and protein serine/threonine phosphatase 2A were analyzed by Western blot. LV myocyte diameter and collagen were significantly reduced in AT2TG-AS compared with NTG-AS mice. LV anterior and posterior wall thickness were not different between AT2TG-AS and NTG-AS mice. LV systolic and diastolic dimensions were significantly higher in AT2TG-AS than in NTG-AS mice. LV systolic pressure and end-diastolic pressure were lower in AT2TG-AS than in NTG-AS mice. ANP, BNP, and SERCA2 were not different between AT2TG-AS and NTG-AS mice. Phospholamban (PLB) and the PLB-to-SERCA2 ratio were significantly higher in AT2TG-AS than in NTG-AS mice. iNOS was higher in AT2TG-AS than in NTG-AS mice but not significantly different. Our results indicate that AT2 receptor overexpression modified the pathological hypertrophic response to aortic banding in transgenic mice.

UTP transactivates epidermal growth factor receptors and promotes cardiomyocyte hypertrophy despite inhibiting transcription of the hypertrophic marker gene, atrial natriuretic peptide.

In neonatal rat ventricular myocytes, activation of receptors that couple to the G(q) family of heterotrimeric G proteins causes hypertrophic growth, together with expression of "hypertrophic marker" genes, such as atrial natriuretic peptide (ANP) and myosin light chain 2 (MLC2). As reported previously for other G(q)-coupled receptors, stimulation of alpha(1)-adrenergic receptors with phenylephrine (50 microM) caused phosphorylation of epidermal growth factor (EGF) receptors as well as activation of ERK1/2, cellular growth, and ANP transcription. These responses depended on EGF receptor activation. In marked contrast, stimulation of G(q)-coupled purinergic receptors with UTP caused EGF receptor phosphorylation, ERK1/2 activation, and cellular growth but minimal increases in ANP transcription. UTP inhibited phenylephrine-dependent transcription from ANP and MLC2 promoters but not transcription from myoglobin promoters or from AP-1 elements. Myocardin is a muscle-specific transcription enhancer that activates transcription from ANP and MLC2 promoters but not myoglobin promoters or AP-1 elements. UTP inhibited ANP and MLC2 responses to overexpressed myocardin but did not inhibit responses to c-Jun, GATA4, or serum response factor, all of which are active in nonmuscle cells. Thus, UTP inhibits transcriptional responses to phenylephrine only at cardiac-specific promoters, and this may involve the muscle-specific transcription enhancer, myocardin. These studies show that EGF receptor activation is necessary but not sufficient for ANP and MLC2 responses to activation of G(q)-coupled receptors in ventricular myocytes, because inhibitory mechanisms can oppose such stimulation. ANP is a compensatory and protective factor in cardiac hypertrophy, and mechanisms that reduce its generation need to be defined.

Modulation of protein phosphatase 2a by adenosine A1 receptors in cardiomyocytes: role for p38 MAPK.

Adenosine A1 receptor activation causes protein phosphatase 2a (PP2a) activation in ventricular myocytes. This attenuates beta-adrenergic functional effects in the heart (Liu Q and Hofmann PA. Am J Physiol Heart Circ Physiol 283: H1314-H1321, 2002). The purpose of the present study was to identify the signaling pathway involved in the translocation/activation of PP2a by adenosine A1 receptors in ventricular myocytes. We found that N6-cyclopentyladenosine (CPA; an adenosine A1 receptor agonist)-induced PP2a translocation was blocked by p38 MAPK inhibition but not by JNK inhibition. CPA increased phosphorylation of p38 MAPK, and this effect was abolished by pertussis toxin and inhibitors of the cGMP pathway. Moreover, CPA-induced PP2a translocation was blocked by inhibition of the cGMP pathway. Guanylyl cyclase activation mimicked the effects of CPA and caused p38 MAPK phosphorylation and PP2a translocation. Finally, CPA-induced dephosphorylations of troponin I and phospholamban were blocked by pertussis toxin and attenuated by p38 MAPK inhibition. These results suggest that adenosine A1 receptor-mediated PP2a activation uses a pertussis toxin-sensitive Gi protein-guanylyl cyclase-p38 MAPK pathway. This proposed, novel pathway may play a role in acute modulation of cardiac function.

AT 2 , Judgment Day

As reviewed recently in these pages,1 the vasoconstrictor angiotensin II (Ang II) is one of the circulating peptide growth factors, and clinically the most relevant, that fulfill many of Koch’s postulates for having additional effects locally in the myocardium. These criteria include local production of Ang II within the heart, its upregulation there by hypertrophic signals, its release from cardiac cells after mechanical load, the presence of its receptors on the surface of cardiac cells, and the responsiveness of both cardiac myocytes and cardiac fibroblasts to provocation by this peptide.1,2 This local circuit of Ang II production, release, binding, and transduction within the myocardium itself is the most cogent explanation for the effectiveness of ACE inhibitors in heart failure, even at doses that do not alter loading conditions.1,2 See p 346 Three related receptors exist for Ang II: AT1a, AT1b, and AT2, which communicate with a complex cell-wide web of signaling cascades through GTP-binding (G) proteins as the proximal transducer. The operation of a local circuit for Ang II within the myocardium poses two questions, which have attracted much recent experimental effort, and an even larger share of controversy. First, given that multiple receptors exist for Ang II, which type or types are responsible for pathobiology provoked by the local circuit? Second, given that cardiac muscle and nonmuscle cells are both potential targets for Ang II, what is the respective role of each? Genetic models that altogether lack the receptors individually provide an invaluable experimental tool beyond what is achievable by use of pharmacological inhibitors alone. Surprisingly, the Ang II receptor …

Distinct cardioprotective effects of adenosine mediated by differential coupling of receptor subtypes to phospholipases C and D

Adenosine released during cardiac ischemia exerts a marked protective effect in the heart that is mediated by the A(1) and A(3) subtypes of adenosine receptors. The signaling pathways activated by these adenosine receptors have now been characterized in a chick embryo ventricular myocyte culture model of cardioprotection against ischemia. Selective A(1) and A(3) receptor agonists were shown to activate phospholipases C and D, respectively, to achieve their distinct cardioprotective effects. The specificity of the A(3) receptor-phospholipase D interaction was also demonstrated in chick embryo atrial myocytes (which do not express endogenous A(3) receptors) that had been transfected with a vector encoding the human A(3) receptor. Activation of both endogenous A(1) and A(3) receptors in ventricular myocytes resulted in a protective response greater than that induced by stimulation of either receptor alone. Agonists that activate both adenosine A(1) and A(3) receptors may thus prove beneficial for the treatment of myocardial ischemia.