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Biomedical subjects

J L Balligand

Publications and source records attributed to J L Balligand.

At least 19 recordsLinked to original sources
PubMed2001

Hsp90 and caveolin are key targets for the proangiogenic nitric oxide-mediated effects of statins.

3-Hydroxy-3-methylglutaryl (HMG)-coenzyme A reductase inhibitors or statins exert direct beneficial effects on the endothelium in part through an increase in nitric oxide (NO) production. Here, we examined whether posttranslational modifications of the endothelial NO synthase (eNOS) could account for the proangiogenic effects of statins. We used endothelial cells (ECs) isolated from cardiac microvasculature, aorta, and umbilical veins, as well as dissected microvessels and aortic rings, that were cultured on reconstituted basement membrane matrix (Matrigel). Tube or precapillary formation was evaluated after statin treatment, in parallel with immunoblotting and immunoprecipitation experiments. Atorvastatin stimulated NO-dependent angiogenesis from both isolated and outgrowing (vessel-derived) ECs, independently of changes in eNOS expression. We found that in macro- but not microvascular ECs, atorvastatin stabilized tube formation through a decrease in caveolin abundance and its inhibitory interaction with eNOS. We also identified the chaperone protein hsp90 as a key target for the proangiogenic effects of statins. Using geldanamycin, an inhibitor of hsp90 function, and overexpression of recombinant hsp90, we documented that the statin-induced phosphorylation of eNOS on Ser1177 was directly dependent on the ability of hsp90 to recruit Akt in the eNOS complex. Finally, we showed that statin promoted the tyrosine phosphorylation of hsp90 and the direct interaction of hsp90 with Akt, which further potentiated the NO-dependent angiogenic processes. Our study provides new mechanistic insights into the NO-mediated angiogenic effects of statins and underscores the potential of these drugs and other modulators of hsp90 and caveolin abundance to promote neovascularization in disease states associated or not with atherosclerosis.

Animals↗
PubMed2001

Hsp90 ensures the transition from the early Ca2+-dependent to the late phosphorylation-dependent activation of the endothelial nitric-oxide synthase in vascular endothelial growth factor-exposed endothelial cells.

Vascular endothelial growth factor (VEGF) exerts its angiogenic effects partly through the activation of endothelial nitric-oxide synthase (eNOS). Association with heat shock protein 90 (hsp90) and phosphorylation by Akt were recently shown to separately activate eNOS upon VEGF stimulation in endothelial cells. Here, we examined the interplay between these different mechanisms in VEGF-exposed endothelial cells. We documented that hsp90 binding to eNOS is, in fact, the crucial event triggering the transition from the Ca(2+)-dependent activation of eNOS to the phosphorylation-mediated potentiation of its activity by VEGF. Accordingly, we showed that early VEGF stimulation first leads to the Ca(2+)/calmodulin disruption of the caveolin-eNOS complex and promotes the association between eNOS and hsp90. eNOS-bound hsp90 can then recruit VEGF-activated (phosphorylated) Akt to the complex, which in turn can phosphorylate eNOS. Further experiments in transfected COS cells expressing either wild-type or S1177A mutant eNOS led us to identify the serine 1177 as the critical residue for the hsp90-dependent Akt-mediated activation of eNOS. Finally, we documented that although the VEGF-induced phosphorylation of eNOS leads to a sustained production of NO independently of a maintained increase in [Ca(2+)](i), this late stage of eNOS activation is strictly conditional on the initial VEGF-induced Ca(2+)-dependent stimulation of the enzyme. These data establish the critical temporal sequence of events leading to the sustained activation of eNOS by VEGF and suggest new ways of regulating the production of NO in response to this cytokine through the ubiquitous chaperone protein, hsp90.

Animals↗
PubMed2001

Upregulation of beta(3)-adrenoceptors and altered contractile response to inotropic amines in human failing myocardium.

BACKGROUND: Contrary to beta(1)- and beta(2)-adrenoceptors, beta(3)-adrenoceptors mediate a negative inotropic effect in human ventricular muscle. To assess their functional role in heart failure, our purpose was to compare the expression and contractile effect of beta(3)-adrenoceptors in nonfailing and failing human hearts. METHODS AND RESULTS: We analyzed left ventricular samples from 29 failing (16 ischemic and 13 dilated cardiomyopathic) hearts (ejection fraction 18.6+/-2%) and 25 nonfailing (including 12 innervated) explanted hearts (ejection fraction 64.2+/-3%). beta(3)-Adrenoceptor proteins were identified by immunohistochemistry in ventricular cardiomyocytes from nonfailing and failing hearts. Contrary to beta(1)-adrenoceptor mRNA, Western blot analysis of beta(3)-adrenoceptor proteins showed a 2- to 3-fold increase in failing compared with nonfailing hearts. A similar increase was observed for Galpha(i-2) proteins that couple beta(3)-adrenoceptors to their negative inotropic effect. Contractile tension was measured in electrically stimulated myocardial samples ex vivo. In failing hearts, the positive inotropic effect of the nonspecific amine isoprenaline was reduced by 75% compared with that observed in nonfailing hearts. By contrast, the negative inotropic effect of beta(3)-preferential agonists was only mildly reduced. CONCLUSIONS: Opposite changes occur in beta(1)- and beta(3)-adrenoceptor abundance in the failing left ventricle, with an imbalance between their inotropic influences that may underlie the functional degradation of the human failing heart.

Adrenergic beta-3 Receptor Agonists↗
PubMed2001

Hydroxy-methylglutaryl-coenzyme A reductase inhibition promotes endothelial nitric oxide synthase activation through a decrease in caveolin abundance.

BACKGROUND: Hypercholesterolemia is causally associated with defects of endothelial nitric oxide (NO)-dependent vasodilation. Increased uptake of cholesterol by endothelial cells (ECs) upregulates the abundance of the structural protein caveolin-1 and impairs NO release through the stabilization of the inhibitory heterocomplex between caveolin-1 and endothelial NO synthase (eNOS). Therefore, we examined whether the hydroxy-methylglutaryl-coenzyme A reductase inhibitor atorvastatin modulates caveolin abundance, eNOS activity, and NO release through a reduction in endogenous cholesterol levels. METHODS AND RESULTS: ECs were incubated with increasing doses of atorvastatin in the absence or in the presence of human LDL cholesterol (LDL-Chol) fractions in the presence of antioxidants. Our results show that atorvastatin (10 nmol/L to 1 micromol/L) reduced caveolin-1 abundance in the absence (-75%) and in the presence (-20% to 70%) of LDL-Chol. This was paralleled by a decreased inhibitory interaction between caveolin-1 and eNOS and a restoration and/or potentiation of the basal (+45%) and agonist-stimulated (+107%) eNOS activity. These effects were observed in the absence of changes in eNOS abundance and were reversed with mevalonate. In the presence of LDL-Chol, atorvastatin also promoted the agonist-induced association of eNOS and the chaperone Hsp90, resulting in the potentiation of eNOS activation. CONCLUSIONS: We provide biochemical and functional evidence that atorvastatin promotes NO production by decreasing caveolin-1 expression in ECs, regardless of the level of extracellular LDL-Chol. These findings highlight the therapeutic potential of inhibiting cholesterol synthesis in peripheral cells to correct NO-dependent endothelial dysfunction associated with hypercholesterolemia and possibly other diseases.

Animals↗
PubMed2001

Real-time RT-PCR for the detection of beta-adrenoceptor messenger RNAs in small human endomyocardial biopsies.

Quantification of mRNAs from extremely small human samples remains a challenge. Requiring minimal amounts of tissue and no post-reaction manipulation, real-time reverse transcriptase-polymerase chain reaction (RT-PCR) is an attractive method to quantitatively assess the expression of rare mRNAs. We evaluated the applicability of the technique on RNA extracted from human endomyocardial biopsies and isolated cardiomyocytes, and compared the technique to the RT-competitive PCR approach. Primers and probes were designed to amplify the three subtypes of human beta -adrenoceptors (beta1-, beta2- and beta3 AR), as well as reference genes such as glyceraldehyde-3-phosphate dehydrogenase (GAPDH), Hypoxanthine-guanine phosphoribosyltransferase (HPRT), and the oncogene ABL by real-time RT-PCR. Specific primers and a deleted competitor were synthetized to compare the quantitation of the beta 3 AR mRNA expression by RT-competitive PCR. We validated the technique on human cardiomyocytes either freshly isolated or selectively excised from fixed sections of human myocardium by Laser Capture Microdissection. The standard curves obtained for the cDNA's analysed showed mean slopes comprised between -3.3 and -3.7. Inter- and intra-assay variability of gene quantitation was reflected by mean values of the variance coefficients of Ct of 4.84+/-1.13% and 2.73+/-0.39% or 3.32+/-1.03% and 2.21+/-0.24% (corresponding to percent variances of copy numbers of 83.07+/-12.72% and 34.45+/-9.03% or 47.40+/-8.59% and 23.83+/-3.16%) for human beta3 AR and GAPDH genes, respectively. The expression of GAPDH, HPRT and ABL mRNA was characterized by a very low dispersion of individual values across cardiac pathologies, suggesting that these genes may be used as reference genes in quantitative PCR studies. Finally, we applied the technique to detect rare mRNAs, such as beta -AR mRNAs, from small human endomyocardial biopsies and even isolated cardiomyocytes. Real-time RT-PCR is appropriate to quantitate rare messenger RNAs, including in extremely small human tissue samples. This method appears very promising for futures studies of gene expression in several pathophysiological conditions, including heart failure.

Adult↗
PubMed2001

Endogenous nitric oxide mechanisms mediate the stretch dependence of Ca2+ release in cardiomyocytes.

Stretching of cardiac muscle modulates contraction through the enhancement of the Ca2+ transient, but how this occurs is still not known. We found that stretching of myocytes modulates the elementary Ca2+ release process from ryanodine-receptor Ca2+-release channels (RyRCs), Ca2+ sparks and the electrically stimulated Ca2+ transient. Stretching induces PtdIns-3-OH kinase (PI(3)K)-dependent phosphorylation of both Akt and the endothelial isoform of nitric oxide synthase (NOS), nitric oxide (NO) production, and a proportionate increase in Ca2+-spark frequency that is abolished by inhibiting NOS and PI(3)K. Exogenously generated NO reversibly increases Ca2+-spark frequency without cell stretching. We propose that myocyte NO produced by activation of the PI(3)K-Akt-endothelial NOS axis acts as a second messenger of stretch by enhancing RyRC activity, contributing to myocardial contractile activation.

Animals↗
PubMed2001

Cardiovascular effects of beta 3-adrenoceptor stimulation in perinephritic hypertension.

BACKGROUND: A new beta 3-adrenoceptor (beta3-AR) has been shown to mediate peripheral vasodilation. This study was conducted to evaluate effects of the beta3-AR agonist, SR58611 in normal and hypertensive dogs. MATERIALS AND METHODS: In protocol 1, SR58611 was infused in normal dogs after placebo, after beta1/beta2 blockade with nadolol, after beta1/beta2/beta3 blockade with bupranolol and after combined autonomic blockade (CAB). In protocol 2, perinephritic hypertension was produced in dogs, which received SR58611 at 3 and 6 weeks of hypertension. Effects of SR58611 were evaluated at 7 weeks of hypertension after CAB. RESULTS: In normal dogs, SR58611 produced a dose-dependent decrease in mean aortic pressure (AOP) (from 116 +/- 19 to 100 +/- 19 mmHg, - 14%; P < 0.05) that was accompanied by baroreflex activation (heart rate increased by 70%; P < 0.01). This hypotensive effect resulting from peripheral vasodilation persisted after nadolol or CAB while baroreflex activation was blunted or abolished. A biphasic response of cardiac output, characterized by a rise and a decline (P < 0.05) reflected a reduction in after- and pre-load. After CAB, SR58611 did not modify cardiac contractility. SR58611 stimulated lipolysis as reflected by a 4-fold increase in blood free fatty acids (FFA) (P < 0.0005). Under CAB, the rise of FFA was reduced (P < 0.01). In hypertensive dogs, SR58611 produced a dose-dependent decrease in mean AOP (from 168 +/- 32 to 125 +/- 35 mmHg; - 26%, P < 0.0001), that was greater than in normal dogs (P < 0.05). Reflex-mediated tachycardia also occurred but at higher blood pressure values. Blood FFA rose similarly (P < 0.0001). Under CAB, heart rate remained unchanged but SR58611 still induced a decrease (P < 0.0001) in mean AOP concomitantly with a rise of (dP/dt)/DP40 (P < 0.005), an effect not observed in normal dogs. CONCLUSIONS: Beta3-AR stimulation exerts hypotensive effects, increases cardiac contractility and stimulates lipolysis in hypertensive dogs.

Adrenergic beta-3 Receptor Agonists↗
PubMed2001

Nitric oxide: does it play a role in the heart of the critically ill?

Nitric oxide regulates many aspects of myocardial function, not only in the normal heart but also in ischemic and nonischemic heart failure, septic cardiomyopathy, cardiac allograft rejection, and myocarditis. Accumulating evidence implicates the endogenous production of nitric oxide in the regulation of myocardial contractility, distensibility, heart rate, coronary vasodilation, myocardial oxygen consumption, mitochondrial respiration, and apoptosis. The effects of nitric oxide promote left ventricular mechanical efficiency, ie, appropriate matching between cardiac work and myocardial oxygen consumption. Most of these beneficial effects are attributed to the low physiologic concentrations generated by the constitutive endothelial or neuronal nitric oxide synthase. By contrast, inducible nitric oxide synthase generates larger concentrations of nitric oxide over longer periods of time, leading to mostly detrimental effects. In addition, the recently identified beta3-adrenoceptor mediates a negative inotropic effect through coupling to endothelial nitric oxide synthase and is overexpressed in heart failure. An imbalance between beta 1 and beta2-adrenoceptor and beta3-adrenoceptor, with a prevailing influence of beta3-adrenoceptor, may play a causal role in the pathogenesis of cardiac diseases such as terminal heart failure. Likewise, changes in the expression of endothelial nitric oxide synthase or inducible nitric oxide synthase within the myocardium may alter the delicate balance between the effects of nitric oxide produced by either of these isoforms. New treatments such as selective inducible nitric oxide synthase blockade, endothelial nitric oxide synthase promoting therapies, and selective beta3-adrenoceptor modulators may offer promising new therapeutic approaches to optimize the care of critically ill patients according to their stage and specific underlying disease process.

Critical Illness↗
PubMed2000

Dynamin mediates caveolar sequestration of muscarinic cholinergic receptors and alteration in NO signaling.

In cardiac myocytes, agonist binding to muscarinic acetylcholine receptors (mAchRs) leads to the targeting of stimulated receptors to plasmalemmal microdomains termed caveolae. Here, we examined whether this translocation leads to mAchR internalization and alteration in downstream NO signaling. Differential binding of membrane-permeant and -impermeant mAchR radioligands on caveolae-enriched membranes revealed that carbachol stimulation of cardiac myocytes induces sequestration of mAchRs through caveolae fission. GTP but not its non-hydrolyzable analog GTP gamma S drove the further detachment of caveolae from myocyte sarcolemma. Also, incubation of extracts of carbachol-stimulated myocytes with recombinant GTPase dynamin induced mAchR sequestration in budded caveolae, while dominant-negative K44A dynamin inhibited it. These data were confirmed by immunofluorescence microscopy on m2 mAchR-expressing COS cells. Finally, repeated carbachol stimulations of mAchRs co-expressed in COS cells with endothelial nitric oxide synthase (eNOS) and wild-type, but not mutant, dynamin led to a progressive increase in mAchR sequestration and a concurrent stabilization of the inhibitory eNOS-caveolin complex. These findings emphasize the role of caveolae in mAchR trafficking and NO signaling, and suggest that caveolae fission may contribute to G-protein-coupled receptor desensitization.

Animals↗
PubMed2000

Beta3-adrenoceptors in the cardiovascular system.

beta-Adrenoceptors of the beta1 and beta2 subtypes classically mediate the effects of catecholamines on the contractility of cardiac muscle and the relaxation of vascular smooth muscle. Since the molecular characterization of the beta3-adrenoceptor in 1989, most studies of this adrenoceptor subtype have focused on its control of lipolysis in adipose tissues. However, more recent studies have investigated the involvement of beta3-adrenoceptors in the physiological control of cardiac and vascular contractility. In this article, the pharmacological and molecular evidence that supports the functional role of beta3-adrenoceptors in cardiovasculartissues of various species, including humans, will be discussed. These data might provide new insights into our understanding of the abnormal responsiveness of the cardiovascular system to catecholamines in heart failure and its treatment with beta3-adrenoceptor antagonists.

Adrenergic beta-3 Receptor Antagonists↗
PubMed2000

A specific method for measurement of nitric oxide synthase enzymatic activity in peritoneal biopsies.

UNLABELLED: A specific method for measurement of nitric oxide synthase enzymatic activity in peritoneal biopsies. BACKGROUND: Nitric oxide (NO) is synthesized by NO synthase (NOS) isoforms that are expressed in the peritoneum. Thus far, NOS activity in the peritoneum has been assessed by nonspecific methods. We describe the application of a specific method for determination of NOS activity in rat and human peritoneal biopsies. METHODS: The L-citrulline assay is based on the stoechiometric production of NO and L-[3H]-citrulline from L-[3H]-arginine by NOS. The assay is technically difficult when applied on small samples with relatively low levels of NOS activity, which required specific procedures for extraction and samples processing. Reaction parameters ensuring assay linearity in the peritoneum were defined. Peritoneum lysates were also used for immunoblot analysis to identify the NOS isoforms involved. RESULTS: A significant NOS activity is detected in the normal peritoneum because of both Ca2+-dependent and Ca2+-independent NOS. The specificity of NOS activity has been demonstrated by various controls, including the NOS inhibitor L-NMMA. Competition experiments with L-valine and amino acid analyses have reasonably excluded the interference of endogenous arginase and L-arginine, which both might underestimate NOS activity. The procedure is sensitive; it detects a high range of NOS activities as well as the appropriate NOS isoforms in various tissues and conditions, as shown by correlations with immunoblot studies. CONCLUSIONS: We have adapted and characterized the L-citrulline assay to measure specific NOS activities within the peritoneum. The peritoneum lysate assayed for NOS activity can also be used for characterizing NOS isoform expression by immunoblot analysis.

Adult↗
PubMed2000

The negative inotropic action of catecholamines: role of beta3-adrenoceptors.

There is now evidence for the involvement of four beta-adrenoceptor populations in the regulation of cardiac function by catecholamines. Beta1- and beta2-adrenoceptor stimulation classically produces an increase in contractility. A fourth beta-adrenoceptor, as yet uncloned and designated provisionally as a beta4-adrenoceptor, also mediates a positive inotropic effect. Beta3-adrenoceptors, which had been cloned at the end of the eighties, has been extensively studied as a potential target for antiobesity and antidiabetic drugs. Its characterization in the heart has opened new fields of investigations for the understanding of the cardiac adrenergic regulation. This review describes the cardiac electrical and mechanical effects induced by Beta3-adrenoceptor stimulation in different species (including human), as well as the signaling pathway. It also analyzes the role of these receptors in the abnormal responsiveness of catecholamines in heart failure.

Adrenergic Antagonists↗
PubMed2000

[Beta 3 adrenergic receptor: physiologic role and potential therapeutic applications].

The characterization in 1989 of the gene encoding the beta 3-adrenoceptor helped to interpret the results of pharmacological experiments on atypical effects of catecholamines distinct from the classical activation of beta 1 and beta 2 adrenoceptors. In rodents, the beta 3 adrenoceptor is abundantly expressed in white adipose tissue where energy is stored in the form of triglycerides and in brown adipose tissue that is specialized for thermogenesis. Treatment of rodents with beta 3 adrenoceptor agonists induces a weight loss related to the stimulation of lipolysis in the two types of tissues. These results led to propose the use of these agonists for the treatment of human obesity and NIDDM. However, the poor lipolytic effect of these agonists in human adipose tissue and the recent discovery of functional beta 3 adrenoceptors in the human heart raise new questions on the therapeutic use of beta 3-adrenoceptor agonists in man. In the human ventricle, these agonists induce a negative inotropic effect. In vessels, stimulation of beta 3-adrenoceptors produces a vasodilation. If these effects are conserved in the failing heart, they could shed a new light on the pathogenic role of the hyperadrenergism associated with cardiac failure, as well as on its treatment with beta-adrenoceptor blockers.

Adipose Tissue↗
PubMed1999

Hypercholesterolemia decreases nitric oxide production by promoting the interaction of caveolin and endothelial nitric oxide synthase.

Hypercholesterolemia is a central pathogenic factor of endothelial dysfunction caused in part by an impairment of endothelial nitric oxide (NO) production through mechanisms that remain poorly characterized. The activity of the endothelial isoform of NO synthase (eNOS) was recently shown to be modulated by its reciprocal interactions with the stimulatory Ca2+-calmodulin complex and the inhibitory protein caveolin. We examined whether hypercholesterolemia may reduce NO production through alteration of this regulatory equilibrium. Bovine aortic endothelial cells were cultured in the presence of serum obtained from normocholesterolemic (NC) or hypercholesterolemic (HC) human volunteers. Exposure of endothelial cells to the HC serum upregulated caveolin abundance without any measurable effect on eNOS protein levels. This effect of HC serum was associated with an impairment of basal NO release paralleled by an increase in inhibitory caveolin-eNOS complex formation. Similar treatment with HC serum significantly attenuated the NO production stimulated by the calcium ionophore A23187. Accordingly, higher calmodulin levels were required to disrupt the enhanced caveolin-eNOS heterocomplex from HC serum-treated cells. Finally, cell exposure to the low-density lipoprotein (LDL) fraction alone dose-dependently reproduced the inhibition of basal and stimulated NO release, as well as the upregulation of caveolin expression and its heterocomplex formation with eNOS, which were unaffected by cotreatment with antioxidants. Together, our data establish a new mechanism for the cholesterol-induced impairment of NO production through the modulation of caveolin abundance in endothelial cells, a mechanism that may participate in the pathogenesis of endothelial dysfunction and the proatherogenic effects of hypercholesterolemia.

Allosteric Regulation↗
PubMed1999

Regulation of aquaporin-1 and nitric oxide synthase isoforms in a rat model of acute peritonitis.

The loss of ultrafiltration (UF) that accompanies acute peritonitis is a common problem in peritoneal dialysis (PD). It has been suggested that changes in nitric oxide (NO)-mediated vascular tone and permeability might be involved in the loss of UF, whereas channel-mediated water permeability should not be affected. This study used a model of acute peritonitis in rats to characterize changes in PD parameters, in correlation with: (1) expression studies of water channel aquaporin-1 and NO synthase (NOS) isoforms and (2) enzymatic assays for NOS in the peritoneum. Compared with controls, rats with peritonitis had a higher removal of plasma urea, a faster glucose absorption, and a loss of UF. Additional changes, including high protein loss, elevated leukocyte counts in dialysate, positive bacterial cultures, edema, and mononuclear infiltrates, were similar to those observed in PD patients with acute peritonitis. Acute peritonitis in rats induced a major increase in total NOS activity, which was inversely correlated with free-water permeability. The increased NOS activity was mediated by both inducible (Ca2+-independent) and endothelial (Ca2+-dependent) NOS isoforms and was reflected by increased peritoneal staining for nitrotyrosine. In contrast, aquaporin-1 expression was unchanged in rats with peritonitis. These findings cast light on the pathophysiology of permeability changes and loss of UF that characterize acute peritonitis. In particular, these data suggest that a local production of NO, mediated by different NOS isoforms, might play a key role in these changes.

Acute Disease↗
PubMed1998

The negative inotropic effect of beta3-adrenoceptor stimulation is mediated by activation of a nitric oxide synthase pathway in human ventricle.

Beta1- and beta2-adrenoceptors in heart muscle cells mediate the catecholamine-induced increase in the force and frequency of cardiac contraction. Recently, in addition, we demonstrated the functional expression of beta3-adrenoceptors in the human heart. Their stimulation, in marked contrast with that of beta1- and beta2-adrenoceptors, induces a decrease in contractility through presently unknown mechanisms. In the present study, we examined the role of a nitric oxide (NO) synthase pathway in mediating the beta3-adrenoceptor effect on the contractility of human endomyocardial biopsies. The negative inotropic effects of a beta3-adrenoceptor agonist, BRL 37344, and also of norepinephrine in the presence of alpha- and beta1-2-blockade were inhibited both by a nonspecific blocker of NO, methylene blue, and two NO synthase (NOS) inhibitors, L-N-monomethyl-arginine and L-nitroarginine-methyl ester. The effect of the NOS inhibitors was reversed by an excess of L-arginine, the natural substrate of NOS, but not by D-arginine. Moreover, the effects of the beta3-adrenoceptor agonist on contractility were associated with parallel increases in the production of NO and intracellular cGMP, which were also inhibited by NOS inhibitors. Immunohistochemical staining of human ventricular biopsies showed the expression of the endothelial constitutive (eNOS), but not the inducible (iNOS) isoform of NOS in both ventricular myocytes and endothelial cells. These results demonstrate that beta3-adrenoceptor stimulation decreases cardiac contractility through activation of an NOS pathway. Changes in the expression of this pathway may alter the balance between positive and negative inotropic effects of catecholamines on the heart potentially leading to myocardial dysfunction.

Adrenergic beta-Agonists↗
PubMed1998

Regulation of cardiac myocyte contractile function by inducible nitric oxide synthase (iNOS): mechanisms of contractile depression by nitric oxide.

Inflammatory cytokines have been implicated in the reversible depression of cardiac contractile function accompanying local or systemic immune stimulation. Incubation of cardiac myocytes with soluble components in the supernatant from cultured rat lung macrophages activated with endotoxin decreases their contractile response to beta-adrenergic stimulation through the induction of iNOS and the subsequent production of nitric oxide by these cells. In the present study, we characterize the mechanisms underlying NO's attenuation of adrenergic responsiveness in cardiac myocytes. iNOS was induced in cultured ventricular myocytes from adult rats by incubation for 20 h with conditioned medium from lipopolysaccharide (LPS)-activated macrophages. iNOS induction did not induce any alteration in beta-adrenergic receptor density or affinity, Galphai protein abundance, or adenylyl cyclase activity in cultured myocytes. Myocyte exposure to activated macrophage-conditioned medium markedly attenuated the elevation of cAMP in response to isoproterenol (Iso, 2 nM). Induction of iNOS with the macrophage-conditioned medium also potentiated the Iso-induced increase in myocyte cGMP. This cGMP increase was totally abolished by NOS inhibitors. NOS inhibition also returned the attenuated cAMP response to 2 nM Iso to levels observed in control cells. Pre-incubation of the cells in isobutylmethylxanthine (IBMX), a phosphodiesterase inhibitor, also partly reversed the attenuation of cAMP increase with 2 nM Iso in cells expressing iNOS. Brief (15 min) exposure of myocytes to the NO donor, S-nitrosoacetylcysteine (SNAC, 100 micro M) which produced a three-fold increase in intracellular cGMP, also decreased by half the contractile response of cardiac myocytes to Iso (2 nM). We conclude that NO endogenously produced by iNOS decreases the intracellular levels of cAMP in response to beta-adrenergic stimulation in isolated cardiac myocytes, in part through a cGMP-mediated mechanism. This effect may participate in the NO-dependent depression of cardiac function following cytokine exposure.

1-Methyl-3-isobutylxanthine↗