Ontogeny of the (Na+,K+)-ATPase during chick skeletal myogenesis.

The properties of the (Na+,K+)-ATPase were studied at different stages of the in vitro and in ouo development of embryonic chick skeletal muscle.Two approaches have been used: 1) direct titration of the ATPase with [3Hlouabain and 2) assay of the transport properties of (Na+,K+)-ATPase, using the ouabain-sensitive rates of *'Rb+ uptake.[3H10uabain binds to a single class of binding sites on both intact cells and crude homogenates from muscle cells.The equilibrium dissociation constant (KO) for the [3H]ouabain-receptor complex was 0.4 .+0.2 p~ at all stages of muscle development either in ouo or in vitro.The number of ouabain-binding sites increased from 0.3 pmol/mg of protein in 11-day-old chick embryo pectoralis muscle to 8.8 pmol/mg of protein at hatching.It increased from 0.3 pmol/mg of protein in undifferentiated myoblasts isolated from 11-day-old embryos to 2.1 pmol/mg of protein in myotubes that had differentiated in vitro.The time course of appearance of ouabain-binding sites during muscle differentiation closely follows the fusion of myoblasts into myotubes, but is not a consequence of fusion since fusionblocked myoblasts also acquire ouabain-binding sites.Flux studies using "Rb+ lead to essentially the same conclusions as the [3H]ouabain-binding experiments.A comparison of the amounts of the (Na+,K+)-ATPase and of the action potential Na+ channel in muscle cells at different stages of the in ouo and in vitro differentiation suggests that the density of these two membrane components may be coordinately regulated.