Electrophysiological properties vary in different regions of the heart, which is essential for effective cardiac function. Increasing evidence suggests that cardiac electrical heterogeneity is attributed to regional variability in distribution of ion channel and related genes. Dramatic progress in genomics has led to the discovery of a large number of ion channel genes, but how these genes are expressed in specific regions of the heart and function in relation to its physiological function are largely unknown. We have recently profiled ~500 channel and related genes in sino-atrial node (SAN), atrio-ventricular node (AVN), atrium and Ventricle by using Mouse Known Gene Array. To validate cDNA array results, we have compared relative expression levels of 96 channel genes in these regions with Real-time PCR.Hearts were excised from 20-25 g adult C57BL mice after schedule 1 killing by cervical dislocation. Total RNA was prepared from SAN, AVN, atrium and ventricles as described by Wittwer et al (2002). Real-time PCR arrays, called Micro Fluidic Cards (Applied Biosystems), were used. Ninety-six pre-designed TaqMan probe and primer sets were pre-loaded in the card. Two ng per well of cDNA were thermal cycled and quantitative ion channel expression results were analysed using the 7900HT Sequence Detection System (Applied Biosystems). Results obtained from 2 pools of 10 mice for each tissue, showed that there are regional differences in channel gene expression in murine heart. Channel genes Cacna1d (Cav1.3), Cacna1g (Cav3.1), Cacna1h (Cav3.2), Cacna2d2, Cacnb3, Scn3b, Kcna1, Kcna6, Kcnab1, Kcne3, Gja5 (Cx40) and HCN1 are more highly expressed in nodal tissue (SAN/AVN) than in atrium and ventricle, while channel genes Gja1(Cx43), Cacn2a1c (Cav1.2), Scn5a, Kcnd2, Kcnj11 and Kcnj2 are preferentially expressed in muscle regions. Channel gene expression between SAN and AVN is similar; this could due to the functional and structural similarities between these two nodal regions. However, Pln and Cacn1c are preferentially expressed in ventricle as compared to atria. Inversely, Gja5, Kcna4, Kcnj3 and Kcnq1 are preferentially expressed in atria versus ventricle. The results suggest that regional differential expression of various channel genes underlies electrophysiological heterogeneity of the heart. Regional pattern expression of ion channels genes in heart could provide new targets for designing drugs with high specificity of action.