Endothelial nitric oxide synthase in the amphibian, Xenopus tropicalis

Abstract

Nitric oxide (NO) is generated by NO synthase (NOS) of which there are three isoforms: neuronal NOS, inducible NOS, and endothelial NOS (eNOS or NOS III). The presence of eNOS and endothelial NO signalling in fish and amphibian blood vessels has been controversial. This study utilised the genome of Xenopus (Silurana tropicalis) to clone eNOS and determine its tissue expression, and then establish if eNOS is involved in vascular regulation. The open reading frame of Xenopus eNOS (XteNOS) cDNA encoded an 1177 amino acid protein that showed closest structural identity to mammalian eNOS. XteNOS mRNA expression was highest in lung and skeletal muscle, with lower expression in the liver, gut, kidney, heart and brain. No discernable XteNOS mRNA expression was observed in the lateral and dorsal aortae. Western analysis of lung protein using an affinity‐purified anti‐XteNOS produced a single band at approximately 140 kDa, which is similar to mammalian eNOS. Immunohistochemistry showed XteNOS immunoreactivity in the collecting duct of the kidney and the lung parenchyma, but not in the endothelium of blood vessels of the kidney or mesentery. Myography using endothelium‐denuded lateral aorta of X. tropicalis found an acetylcholine‐induced NO‐mediated vasodilation that is most likely attributed to perivascular nitrergic nerves. Thus, Xenopus has an eNOS gene that is not expressed in the vascular endothelium.