Investigate the regional heterogeneity of electrophysiological properties and intracellular calcium in the heart

Cardiovascular diseases, and the progression to heart failure, are one of the leading causes of death. Cardiac tissue slices are becoming increasingly popular as a model system for cardiac electrophysiology and pharmacology research and development. The regional heterogeneity of intracellular calcium and trans-membrane potential, in particular transmural difference in the ventricle and its effect on the electrophysiological substrate is largely unexplored despite the well-established importance of the electrophysiological heterogeneity of the heart. Such information however, can not currently be obtained from either isolated single cell or Langendorff perfused whole heart preparations that are widely used in cardiac research. Here, we described in detail preparation, handling and optical mapping recording using voltage and calcium fluorescent dyes in ventricular tissue slices from mice. We also tried to look at alternans and arrhythmia in different regions of heart by applying field stimulation at different frequencies. We investigated the possible role of Pak1 in autonomic modulation of sino-atrial node pace making activity. The ventricle was cut into transverse and longitudinal direction for optical mapping recording. APD and CaTD were longer in mid-myocardium region compare to epicardium and endocardium region of transverse section of the ventricle. The mid-myocardium region contains M-cells which had a prominent spike and dome action potential configuration. We also observed longer CaTD and APD in apical region compare to mid and basal region of left ventricle (longitudinal slices). The rapid increase in voltage and calcium responses in ventricular slices was mainly caused by applying cumulative pacing frequencies (from 2 Hz to 16 Hz). At higher frequency, alternans and arrhythmia were seen in different regions of ventricle. Pak1 played significant role in the response of murine SAN to autonomic signaling, acting to oppose adrenergic and augment cholinergic responses in the intact system. Pak1 acted as an anti-hypertrophic and anti-arrhythmic in β-adrenergic stress condition. In conclusion, the methodology describe here, thus provides a novel model system for the study of regional heterogeneity of several electrophysiological properties in the heart. Our study indicates that there are transmural and regional differences in calcium transient and transmembrane potential in ventricular slices (transverse and longitudinal section). Pak1 displays cardio-protective effects by de-phosphorylation of several cardiac proteins (cTnI, cTnC and MyBP-C) and activation of protein phosphatase 2A (PP2A).