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- Characterization of the cyclic nucleotide phosphodiesterase subtypes involved in the regulation of the L-type Ca2+ current in rat ventricular myocytesPublication . Verde, Ignacio; Vandecasteele, Grégoire; Lezoualc'h, Frank; Fischmeister, RodolphePhosphorylation of cardiac L-type Ca2+ channels by cyclic AMP-dependent protein kinase (PKA) plays a determinant role in the hormonal regulation of myocardial contraction. PKA increases the mean open probability of individual Ca2+ channels which results in an increase in the macroscopic L-type calcium current (ICa) (McDonald et al., 1994). Activation of PKA usually results from an increased production of cyclic AMP by activation of membrane receptors positively coupled to adenylyl cyclase via stimulatory G proteins (Gs). The best documented of such a regulation is the positive inotropic effect of sympathomimetic amines, such as isoprenaline (Hartzell et al., 1991; Hove-Madsen et al., 1996). However, cardiac myocytes, as most other cell types, also possess a negative feedback mechanism to adenylyl cyclase activation which is constituted of the cyclic nucleotide phosphodiesterases (PDEs), a family of enzymes that break down cyclic AMP into 5'-AMP (Beavo, 1995). Cyclic nucleotide PDE activity, at any given location within the cell, will counterbalance the synthesis of cyclic AMP and determine the extent of PKA activation and, hence, of protein phosphorylation. In particular, at the sarcolemmal membrane, this balance between adenylyl cyclase and PDE activities will control the degree of ICa stimulation upon hormonal activation (Fischmeister & Hartzell, 1991; Hove-Madsen et al., 1996). Other factors are involved, such as cyclic AMP compartmentation (Jurevicius & Fischmeister, 1996), PKA tethering to the membrane (Gao et al., 1997), or phosphatase activity (Wiechen et al., 1995). [...]
- Cyclic GMP regulation of the L-type Ca2+ channel current in human atrial myocytesPublication . Verde, Ignacio; Vandecasteele, Grégoire; Rucker-Martin, C.; Donzeau-Gouge, P.; Fischmeister, RodolpheThe cardiac L-type Ca2+ channel current (ICa) is an important determinant of myocardial contractility. Its regulation by neurotransmitters, hormones, and paracrine factors contributes to the control of cardiac output to meet the demands of the body. A large number of these extracellular first messengers, acting on specific membrane receptors in cardiac myocytes, regulate the activity of adenylyl cyclase which in turn controls the intracellular concentration of cAMP, the activity of the cAMPdependent protein kinase (PKA), and the degree of phosphorylation and stimulation of L-type Ca2+ channels (Hartzell, 1988; McDonald et al. 1994; Hove-Madsen et al. 1996; Striessnig, 1999). A typical example of such regulation is the control of heart function by the sympathetic and parasympathetic nervous systems, which act via adrenoceptors and muscarinic receptors (Brodde & Michel, 1999). In addition to the cAMP cascade, other factors regulate heart function by acting primarily on the cGMP cascade; these include atrial and brain natriuretic peptides (de Bold et al.1996) and nitric oxide (NO) (Paulus & Shah, 1999; Shah & MacCarthy, 2000).
- cGMP-stimulated cyclic nucleotide phosphodiesterase regulates the basal calcium current in human atrial myocytesPublication . Verde, Ignacio; Rivet-Bastide, Michèle; Vandecasteele, Grégoire; Hatem, Stéphane; Bénardeau, Agnès; Mercadier, Jean-Jacques; Fischmeister, RodolpheEHNA (Erythro-9-[2-hydroxy-3-nonyl]adenine) is a wellknown inhibitor of adenosine deaminase. Recently, EHNA was shown to block the activity of purified soluble cGMPstimulated phosphodiesterase (PDE2) from frog, human, and porcine heart with an apparent Ki value of approximately 1 microM and with negligible effects on Ca2+/calmodulin PDE (PDE1), cGMP-inhibited PDE (PDE3), and low Km cAMP-specific PDE (PDE4) (Méry, P.F., C. Pavoine, F. Pecker, and R. Fischmeister. 1995. Mol. Pharmacol. 48:121-130; Podzuweit, T., P. Nennstiel, and A. Muller. 1995. Cell. Signalling. 7:733- 738). To investigate the role of PDE2 in the regulation of cardiac L-type Ca2+ current (ICa), we have examined the effect of EHNA on ICa in freshly isolated human atrial myocytes. Extracellular application of 0.1-10 microM EHNA induced an increase in the amplitude of basal ICa ( approximately 80% at 1 microM) without modification of the current-voltage or inactivation curves. The maximal stimulatory effect of EHNA on ICa was comparable in amplitude with the maximal effect of isoprenaline (1 microM), and the two effects were not additive. The effect of EHNA was not a result of adenosine deaminase inhibition, since 2'-deoxycoformycin (1-30 microM), another adenosine deaminase inhibitor with no effect on PDE2, or adenosine (1-10 microM) did not increase ICa. In the absence of intracellular GTP, the substrate of guanylyl cyclase, EHNA did not increase ICa. However, under similar conditions, intracellular perfusion with 0.5 microM cGMP produced an 80% increase in ICa. As opposed to human cardiomyocytes, EHNA (1-10 microM) did not modify ICa in isolated rat ventricular and atrial myocytes. We conclude that basal ICa is controlled by PDE2 activity in human atrial myocytes. Both PDE2 and PDE3 may contribute to keep the cyclic nucleotides concentrations at minimum in the absence of adenylyl and/or guanylyl cyclase stimulation.
- Muscarinic and beta-adrenergic regulation of heart rate, force of contraction and calcium current is preserved in mice lacking endothelial nitric oxide synthasePublication . Vandecasteele, Grégoire; Eschenhagen, Thomas; Scholz, Hasso; Stein, Birgitt; Verde, Ignacio; Fischmeister, RodolpheNitric oxide (NO) is an ubiquitous signaling molecule produced from L-arginine by NO synthase (NOS). In the vasculature, NO mediates parasympathetic endothelium-dependent vasodilation. NO may also mediate the parasympathetic control of myocardial function1. This is supported by the observations that NOS3, the endothelial constitutive NOS, is expressed in normal cardiac myocytes from rodents2 and human3, and NOS and/or guanylyl cyclase inhibitors antagonize the effect of muscarinic agonists on heart rate4,5, atrio–ventricular conduction6, contractility2,4,7 and L-type calcium current1,2,5,6. Here we examine the autonomic regulation of the heart in genetically engineered mice deficient in NOS3 (NOS3-KO)(ref. 8). We show that the chronotropic and inotropic responses to both β-adrenergic and muscarinic agonists were unaltered in isolated cardiac tissue preparations from NOS3-KO mice, although these mice have a defective parasympathetic regulation of vascular tone8,9. Similarly, β-adrenergic stimulation and muscarinic inhibition of the calcium current did not differ in cardiac myocytes from NOS3-KO mice and those from wild-type mice. RT–PCR did not demonstrate upregulation of other NOS isoforms. Similarly, Gi/Go proteins and muscarinic receptor density were unaltered. These data refute the idea that NOS3 is obligatory for the normal autonomic control of cardiac muscle function10.