Recently developed rat heart myocyte cell lines have afforded us the opportunity to evaluate the expression of several transcription factors associated with early cardiac development. These factors include, but are not limited to, Nkx-2.5/Csx, MEF-2C and MLP (Muscle LIM Protein). These factors have been shown to be temporally expressed in pre-cardiac mesenchyme coincident with the earliest stages of heart development. Using the BWEM and CLEM myocyte cell lines as models of the embryonic, committed cardiomyocyte, we have evaluated the basal expression levels of these three genes over multiple passages. Both cell lines express these genes, with MEF-2C being the most abundant based on Northern blot hybridization analyses. Interestingly, as these cells increased their passage number, there was a corresponding increase in their basal expression levels. To evaluate potential 'downstream' effectors of these genes, we examined the basal expression levels of two cardiac-specific genes cTNC and MLC-2v. Transcript levels for both of these contractile filament genes were elevated with passage, suggestive of a inductive process mediated by one or all these three transcription factors. Promoter analysis of MLC-2v expression in the CLEM line shows that this increase is transcriptionally-mediated and the lines retain the necessary regulatory factors to maintain and control the transcription of this gene. Analysis of the dynamics of the regulatory role(s) that these three transcription factors play in cardiac development can now be evaluated in a homogeneous, cell culture system.