Abstract 1237: Delta-Notch Signaling Determines Atrioventricular Conduction System Cell Fate
Introduction: There is growing evidence from several model organisms that even at the ‘heart tube’ stage of development, atrioventricular (AV) ring myocytes are beginning to differentiate into conduction tissue, with slowing of the activation wavefront and increased refractory periods. The signals responsible for the specification of central conduction system are relatively inaccessible in most models, but using functional assays in the developing zebrafish it is possible to explore the earliest steps in myocardial patterning and differentiation. Based on the expression of zebrafish notch1b in the AV endocardium, we hypothesized that notch and delta were determinants of the conduction system cell fate.
Methods: Antisense morpholino oligos were used to knockdown target gene expression in zebrafish embryos. In vivo calcium imaging with calcium green-1 was used to characterize myocardial activation patterns during the first 72 hours post fertilization. Standard whole mount immunohistochemistry (IHC) was performed.
Results: Morpholino knockdown of notch1b expression leads to the failure of AV conduction tissue development in 28% of fish tested (n=108). In contrast, knockdown of delta expression results in a dramatic expansion of the atrioventricular ring in 96% of injected embryos (n=111). The expanded AV ring has AV conduction tissue properties as evidenced by spontaneous 2:1 AV Block observed in 6% of delta knockdown embryos (n=68) and by calcium activation patterns in delta knockdown embryos that demonstrate slowing of the activation wavefront throughout the elongated ring. IHC analyses suggest that this expansion results from recruitment of ventricular myocytes to a conduction cell fate.
Conclusions: These results directly confirm previous evidence that central slow AV conduction system is derived from primary cardiomyocytes and define, for the first time, a critical role for canonical Notch-Delta signaling in this cell fate decision.