Lineage and development of the parasympathetic nervous system of the embryonic chick heart.

We were interested in the contribution of the cardiac neural crest to the complete anterior and posterior nerve plexus of the chick heart. This includes the pathways by which these cardiac neural crest-derived neuronal precursors enter the heart. As lineage techniques we used the traditional quail-chick chimera in combination with the newly introduced technique of retroviral reporter gene transfer to premigratory cardiac neural crest cells. Retrovirally infected embryos (n=23) and quail-chick chimeras (n=19) between stages HH27 and 40, were immunohistochemically evaluated, using the lineage markers LacZ (retroviral reporter) and QCPN (anti-quail nuclear marker), respectively and the neuronal differentiation markers HNK-1, RMO-270 and DO-170. Between stages HH27 and 33, quail-derived and LacZ positive cells were situated around the arterial cardiac vagal branches at the arterial pole, and vagal branches along the anterior cardinal veins and the sinal vagal branch at the venous pole. From stage HH35 onward, QCPN/LacZ-positive cardiac ganglia were observed throughout the anterior and posterior plexus and were mainly concentrated in the subepicardium near the distal ends of the arterial cardiac vagal branches and the sinal cardiac vagal branch respectively. From stage HH36 both the anterior and posterior plexus contained a population of large cardiac ganglion cells and a population of smaller cells along nerve branches as well as in the cardiac ganglia, which means that differentiation starts in both plexus at the same time. Furthermore only nerve fiber connections between the anterior and posterior plexus were observed. These results show that the cardiac neural crest contributes to the cardiac ganglion cells from both the entire anterior and posterior plexus. Furthermore these results suggest that these precursor cells enter the arterial pole via the arterial cardiac vagal branches and the venous pole via the sinal cardiac vagal branch without intermixing. Finally we show that in addition to the cardiac ganglia, the cardiac neural crest contributes to small myocardial glia or undifferentiated cells along nerve fibers, and some myocardial nerve fibers as well as nerve tissue in the adventitia of the large veins at the venous pole and in the adventitia of the coronary arteries.