Functional characterization of cardiac progenitor cells and their derivatives in the embryonic heart post‐chamber formation

There is scant information on the fate of cardiac progenitor cells (CPC) in the embryonic heart after chamber specification. Here we simultaneously tracked three lineage‐specific markers (Nkx2.5, MLC2v, and ANF) and confirmed that CPCs with an Nkx2.5⁺MLC2v^?ANF^? phenotype are present in the embryonic (E) day 11.5 mouse ventricular myocardium. We demonstrated that these CPCs could give rise to working cardiomyocytes and conduction system cells. Using a two‐photon imaging analysis, we found that the majority of CPCs are not capable of developing Ca²⁺ transients in response to β‐adrenergic receptor stimulation. In contrast, Nkx2.5⁺ cells expressing MLC2v but not ANF are capable of developing functional Ca²⁺ transients. We showed that Ca²⁺ transients could be invoked in Nkx2.5⁺MLC2v⁺ANF⁺ cells only upon inhibition of Gi, muscarinic receptors, or nitric oxide synthase (NOS) signaling pathways. Our data suggest that these inhibitory pathways may delay functional specification in a subset of developing ventricular cells. Developmental Dynamics 238:2787–2799, 2009. © 2009 Wiley‐Liss, Inc.