Beta3-Adrenoceptor in the eel (Anguilla anguilla) heart:negative inotropy and NO-cGMP-dependent mechanism

Abstract

SUMMARY Neuroendocrine regulation of cardiac function involves a population of three types of β-adrenoceptors (ARs). In various mammalian species,β1- and β2-AR stimulation produces an increase in contractility;whereas β3-AR activation mediates negative inotropic effects. At the moment, nothing is known about the physiological role of β3-AR in fish. Using an isolated working heart preparation, we show that a β3-AR selective agonist BRL37344 (0.1-100 nmol l-1) elicits a dose-dependent negative inotropism in the freshwater eel Anguilla anguilla. This effect was insensitive to the β1/β2-AR inhibitor nadolol (10 μmol l-1), but was blocked by theβ3-AR-specific antagonist SR59230 (10 nmol l-1). The analysis of the percentage of stroke work (SW) variations, in terms of EC50 values, induced by BRL37344 alone (10 nmol l-1), and in presence of SR59230 (10 nmol l-1), indicated a competitive antagonism of SR59230. In addition to the classic positive inotropism, the non-specific β agonist isoproterenol (100 nmol l-1) induced, in 30% of the preparations, a negative inotropic effect that was abrogated by pre-treatment with SR59230, pointing to a β3-mediated pathway. The BRL37344-induced negative inotropic effect was abolished by exposure to a Gi/o proteins inhibitor pertussis toxin (PTx; 0.01 nmol l-1), suggesting a Gi/o-dependent mechanism. Using L-N5(l-imino-ethyl)ornithine (L-NIO; 10 μmol l-1), as a nitric oxide (NO) synthase (NOS) blocker and haemoglobin (Hb; 1 μmol l-1), as a NO scavenger, we demonstrated that NO signalling is involved in the BRL37344-induced response. Pre-treatment with either an inhibitor of soluble guanylate cyclase (GC) 1H-(1,2,4)oxadiazolo-(4,3-a)quinoxalin-1-one (ODQ; 10 μmol l-1), or an inhibitor of the cGMP-activated protein kinase (PKG) KT5823 (100 nmol l-1), abolished the β3-dependent negative inotropism,indicating the cGMP-PKG component as a crucial target of NO signalling. Taken together, our findings provide functional evidence for the presence ofβ3-like adrenoceptors in the eel Anguilla anguilla heart identifying, for the first time in a working fish heart, theβ3-AR-dependent negative inotropy discovered in mammals.