We have used in situ hybridization to analyze Na,K-ATPase alpha and beta subunit gene expression during zebrafish embryogenesis. The most striking finding is that each of the 14 Na,K-ATPase genes exhibits a distinct expression profile. All alpha and beta subunit genes are expressed in the nervous system, although the pattern of expression in different regions varies dramatically. In peripheral tissues, three of the five alpha1-like genes are expressed in pronephros and mucous cells, one is expressed in heart, and one is predominant in skeletal muscle. The alpha2 gene is expressed in brain and heart but is most prominent in skeletal muscle, while the two alpha3 genes are restricted in their expression to the nervous system. Of the six beta subunit genes, beta1a is expressed at highest abundance in lens, pronephros, and heart, while beta1b transcripts are abundant in mucous cells. The two beta2-like genes are differentially expressed in the nervous system. One beta3 gene is expressed exclusively in brain while the other is abundantly expressed in skeletal muscle. Based on these expression patterns, we predict that at least 14 alpha/beta subunit pairs are likely to be formed in different tissues.