Electrophysiological basis of the first heart beats

Earlier studies on cardiac pacemaker activity were inconclusive regarding the electrophysiological mechanism(s) of first electrical activity generation during cardiac development. We therefore aimed to investigate the role of ion channels in action potential (AP) formation and pacemaker activity during very early murine heart development (E8.5).

The voltage clamp mode of the whole-cell patch clamp technique was used to study whole cell ion currents at the single cell level. Extracellular field potentials (FPs) of whole mount hearts were recorded by the use of multi-electrodes arrays (MEAs). RT-PCR was performed to study mRNA expression of the tested ion channels. E8.5 cardiomyocytes displayed membrane potential (MP) oscillations, primitive APs but also more mature-like APs. Cells with primitive APs expressed higher levels of Ca(v)1.2, HCN4, Ca(v)3.2 and NCX1 than cells with MP oscillations and mature-like cells expressed higher levels of K(ir)3.1 (P-like) or K(ir)2.1 (V- and A- like) than cells with primitive APs. Blockers of I(CaL) (verapamil+nifedipine), I(NCX) (KB-R7943 and SEA0400), and I(CaT) (Ni(2+)) led to a complete halt of cardiac APs and FPs even when applied alone, while blocker of I(f) (Cs(+)) only suppressed AP/FP frequency.

Our data imply that i) The high expression of Ca(v)1.2, HCN4, Ca(v)3.2, and NCX1 contributed to the shift from MP oscillations to primitive APs at E8.5. ii) Spontaneous AP generation may rely on the sum of transsarcolemmal Ca(2+) influx through I(CaL), I(NCX) and I(CaT) since blockage of each individual ion channel/exchanger led to an immediate halt of beating at E8.5.