Aim: 

Mouse embryonic stem cells (ESCs) are pluripotent stem cells able to differentiate into several cell types, including cardiac myocytes. This work aims to characterize the f-current and HCN isoforms expressed by ESC-derived pacemaker cells.

Methods: 

mESCs were differentiated through the "hanging drops" technique which leads to the formation of embryoid bodies (EBs), cellular aggregates that recapitulate early stage of embryo differentiation. The EBs were dissociated at day 7 + 8 1 and electrophysiological experiments were performed in order to record spontaneous activity and I_f current. HCN are the ion channel isoforms which carry I_f current; HCN expression was characterized by RT-PCR and immunofluorescence experiments on undifferentiated ESC and on contractile part of the EBs.

Results: 

Electrophysiological experiments conducted on autorhythmic cells derived from the EBs showed that they exhibited spontaneous action potentials and express I_f current; two populations with different current kinetics were identified, one exhibiting a slow-activating I_f and the other a fast-activating I_f. RT-PCR analysis pointed out that both undifferentiated mESCs and the contractile parts of EBs expressed all the four isoforms of pacemaker channels. Immunofluorescence experiments, instead, demonstrated that only HCN1 and HCN4 were expressed in ES-derived myocytes, while HCN2 was never detected and HCN3 was limited to non-cardiac cells.

Conclusion: 

Our data show that ESC-derived pacemaker myocytes express I_f currents with different kinetics, which can be, at least in part, due to the prevalent expression of either HCN1 or HCN4 channels, the most widely expressed isoforms in the sinus node.