Cardiac gene promoter analysis remains an integral method in molecular cardiology and continues to provide novel insights into the transcriptional mechanisms that regulate gene expression in the myocardium. Initial studies focused on the regulated expression of contractile genes, since their transcripts are abundant and their cDNAs were among the first to be cloned. More recent studies have focused on the promoters of genes expressed at much lower levels, including those that encode ion channels, signaling proteins, and the cardiac transcription factors. The standard approach to analyze myocardial gene promoters has been to transfert reporter plasmids into cultured neonatal rat cardiac myocytes. This approach has the unique advantage of allowing the exploration of different signaling mechanisms by supplementing culture media with different agonists and inhibitors. In addition, cis-elements that control gene expression under different physiological stresses have been further characterized in the context of heterologous promoters to demonstrate their “stand-alone” functional properties in the absence of confounding influences from other cis-elements and their cognate transcription factors. Here we illustrate the characterization of cardiac gene promoter activity using reporter constructs and heterologous promoter studies in cultured cardiac myocytes.