ONTOGENY OF NITRIC-OXIDE SYNTHASES IN THE VENTILATORY MUSCLES

Nitric oxide (NO) acts as an endogenous mediator in mature skeletal mu scle. In this study, we investigated the regulation of the endothelial (eNOS) and neuronal (nNOS) isoforms of nitric oxide synthase (NOS) in skeletal-muscle development (rat diaphragm). Muscle NOS activity, nNO S and eNOS protein, and mRNA expressions were markedly increased durin g the late gestational and early postnatal periods. Expression of both isoforms, however, declined progressively thereafter. Similarly, argi ninosuccinate lyase and argininosuccinate synthetase, both involved in the recycling of L-citrulline to L-arginine, were expressed at high l evels in rat embryonic and neonatal diaphragms, with gradual reduction in their expression during late postnatal development. Immunostaining revealed extensive nNOS expression at the sarcolemma in neonatal and mature diaphragms, whereas eNOS expression was limited to the endothel ium. Both neonatal and adult diaphragms expressed an alternatively spl iced nNOS isoform with an insert of 34 amino acids between exons 16 an d 17. In vitro-generated muscle force rose significantly after NOS inh ibition in both neonatal and adult diaphragms, but the magnitude of fo rce augmentation was larger in adult than in neonatal diaphragm. These results indicate that constitutive NOS isoforms are developmentally r egulated in skeletal muscles, suggesting multiple roles for NO in deve loping and mature skeletal-muscle fibers.