Fading of 5-HT₄ receptor-mediated inotropic responses to 5-hydroxytryptamine is caused by phosphodiesterase activity in porcine atrium

Abstract

Inotropic responses to 5-hydroxytryptamine (5-HT) in human and porcine atrium can fade, suggesting 5-HT₄ receptor desensitization. De Maeyer et al., however, show in this issue that inhibition of phosphodiesterases with isobutyl-methyl-xanthine prevents fading of 5-HT₄ receptor-mediated responses to 5-HT and the partial agonist prucalopride in porcine atrium.

5-Hydroxytryptamine (5-HT) is a cardiostimulant in man. 5-HT increases contractility and hastens relaxation of isolated myocardium from right and left atrium. These effects are mediated through 5-HT₄ receptors, and are associated with increases in cyclic AMP and stimulation of cyclic AMP-dependent protein kinase (Kaumann et al., 1990, Sanders & Kaumann, 1992). 5-HT also elicits rate-dependent arrhythmias through 5-HT₄ receptors in atrial trabeculae and myocytes (Kaumann & Sanders, 1994; Sanders et al., 1995). Chronic treatment with β₁-adrenoceptor-selective blockers enhances both the inotropic and arrhythmic effects of 5-HT (Kaumann & Sanders, 1994; Sanders et al., 1995). A remarkably low density of 5-HT₄ receptors (∼4 fmol mg⁻¹) mediates the atrial effects of 5-HT (Kaumann et al., 1996). Initial searches for ventricular 5-HT₄ receptors were negative (Jahnel et al., 1992; Schoemaker et al., 1993). However, ventricular mRNA for 5-HT_4(a) and 5-HT_4(b) was detected (Bach et al., 2001), making the existence of functional 5-HT₄ receptors plausible. The failure to detect ventricular effects of 5-HT could be due to a very low 5-HT₄ receptor density combined with avid hydrolysis of cyclic AMP by phosphodiesterases (PDE). Indeed, in the presence of the nonselective PDE inhibitor isobutyl-methyl-xanthine (IBMX), 5-HT does increase contractile force, hasten relaxation and cause arrhythmic contractions in ventricular trabeculae. These effects are antagonized by the 5-HT₄-selective blocker GR113808, consistent with functional ventricular 5-HT₄ receptors (Brattelid et al., 2004). The 5-HT₄ mRNA is increased four-fold in ventricles from failing hearts compared to nonfailing hearts (Brattelid et al., 2004).

Porcine atrial and sinoatrial 5-HT₄ receptors have been used as experimental models for human atrial 5-HT₄ receptors. 5-HT produces tachycardia in healthy volunteers (Le Messurier et al., 1959), but the nature of the human sinoatrial 5-HT receptors remains unknown. To investigate the properties of sinoatrial 5-HT receptors, spontaneously beating right atrium of new-born piglets were used (Kaumann, 1990). The chronotropic potency and intrinsic activity of 5-HT and 5-carboxamidotryptamine, as well as of the partial agonists renzapride and cisapride, corresponded to a 5-HT₄ receptor profile and agreed quantitatively with the corresponding positive inotropic effects of these ligands on human atrium (Kaumann, 1990, Kaumann et al., 1991b). The results make it plausible that human sinoatrial 5-HT receptors are 5-HT₄ receptors, but experiments to prove this (Kaumann & Sanders, 1998) are still pending. 5-HT₄ receptors also mediate the tachycardia evoked by 5-HT and partial agonists in adult pigs (Villalon et al., 1991; Parker et al., 1995). In addition, functional 5-HT₄ receptors have been identified in paced left atria of new-born piglets (Kaumann et al., 1991a; Parker et al., 1995) but not in porcine ventricle (Lorrain et al., 1992; Schoemaker et al., 1992). However, as found in human ventricle, the PDE inhibitor IBMX also uncovers functional 5-HT₄ receptors in porcine ventricle. Interestingly, 5-HT was twice as efficacious and a 15-fold more potent inotropic agonist on ventricular trabeculae from adult pigs than from new-born piglets (Brattelid et al., 2004). The age-related difference was mirrored by a greater PKA activation in trabeculae from adult pigs than from new-born piglets (Brattelid et al., 2004).

The inotropic effects of 5-HT are more pronounced on atria from adult humans than on atria from new-born piglets (Kaumann & Sanders, 1998). This is in part related to an even lower atrial 5-HT₄ receptor density in newborn piglet (∼0.3 fmol mg protein⁻¹, Kaumann et al., 1995) than in adult man. Several other mammalian species, including the rat, do not express at all functional cardiac 5-HT₄ receptors (Kaumann, 1991). However, recent work has demonstrated the appearance of functional 5-HT₄ receptors that mediate increases in contractile force, cyclic AMP levels and relaxation in ventricular papillary muscles from rats with heart failure 6 weeks after coronary artery ligation (Qvigstad et al., 2005).

Inotropic responses to 5-HT and 5-HT₄ receptor partial agonists can fade in human and porcine atrial myocardium (Kaumann et al., 1991b; Sanders & Kaumann, 1992; Parker et al., 1995). Fading is usually attributed to desensitization of the receptor system but, in the case of receptors coupled to G_s protein, hydrolysis of cyclic AMP by PDEs can also contribute. For example, fading of the positive inotropic responses to glucagon is reduced by the PDE4-selective inhibitor rolipram in rat ventricle, suggesting involvement of PDE4 (Juan-Fita et al., 2004). In a carefully designed study, De Maeyer et al. (2005, this issue) report in paced left atria of young pigs that PDE inhibition with IBMX abolishes fading of the positive inotropic responses to 5-HT. The noncumulative administration of the partial agonist prucalopride increased contractile force that faded and cumulatively administered concentrations failed altogether to elicit effects. IBMX potentiated the responses to 5-HT and allowed the determination of cumulative concentration–effect curves for partial agonists without fade. Unlike inotropic responses, chronotropic responses to 5-HT and partial agonists do not fade in spontaneously beating right atria, suggesting that PDEs may not limit the responses through sinoatrial 5-HT₄ receptors. De Maeyer et al. found that atrial inotropic responses, in the presence of IBMX, were less marked in new-born piglets than adolescent pigs, as previously reported for ventricle by Brattelid et al. (2004).

The work of De Maeyer et al. (2005) clearly highlights PDE-evoked reduction of contractile responses, mediated through porcine 5-HT₄ receptors. The increase of the effects of 5-HT by IBMX is, however, not only due to prevention of cyclic AMP hydrolysis upon 5-HT₄ receptor activation but also to the considerable positive and lusitropic effects caused by IBMX. The IBMX-induced cardiostimulation adds to the 5-HT-evoked cardiostimulation and should be subtracted from the combined effects of 5-HT and IBMX, so that the 5-HT₄ receptor component of PDE activity can be assessed. De Maeyer et al. (2005) did not address the role of selective PDE isoenzymes. Experiments with inhibitors selective for the two major cardiac isoenzymes, PDE3 and PDE4, should be carried out to unravel which isoenzyme is mainly responsible for the fade. The sinoatrial tachycardia of 5-HT does not appear to fade but PDE activity could still contribute to reduce this response, although probably with a faster time-course than on left atrial myocardium. De Maeyer et al. (2005) confirmed the work of Krobert et al. (2005) that prucalopride elicits sinoatrial tachycardia in new-born piglets in the absence of IBMX. Krobert et al. (2005) found that the cumulative concentration–effect curve of prucalopride for the sinoatrial tachycardia is bell shaped, raising the possibility of a role for PDEs.

Endogenous 5-HT has been proposed to trigger cardiac arrhythmias (Kaumann, 1994; Brattelid et al., 2004) and a PDE inhibitor can facilitate arrhythmias and enhance mortality in heart failure (Packer et al., 1991). Therefore, the clinical combination of a PDE inhibitor with a 5-HT₄ receptor partial agonist should be used with caution.