Despite extensive research on cardiovascular development, much remains to be learned about the molecular and cellular mechanisms that give rise to the fully formed 3D structure of the heart. Though genetic studies have identified genes necessary for proper cardiac morphogenesis, our understanding of how these genes impact myocardial cell biology is limited. In this thesis, I use 4D microscopy of zebrafish embryos coupled with computational pos-processing to study how individual cells contribute to the formation of the ventricular trabeculae, and I identify a subpopulation of cardiomyocytes that can be marked by their activation of Notch signaling which go on to form the cortical myocardial layer of the adult zebrafish heart.

The wall of the developing ventricle is covered by a network of sheet-like muscular structures termed the cardiac trabeculae. Though a number of mutants have been identified that lack trabeculae, little is known about how cardiomyocytes interact to form these sheets. Previous work from our lab had suggested that cardiomyocytes migrate from the outer, compact layer of the heart towards the lumen to form these sheets. However, how exactly they migrated was unknown. I found that cardiomyocytes form the trabecular layer first by extending long processes over their neighbors, then by constricting their abluminal surfaces to move their cell bodies into the forming sheets. I further found that two mutations that disrupt trabeculation affect the initial processes differently. While erbb2 mutant cells form normal, stable processes, cells in embryos with non-contractile hearts extend processes that collapse quickly. This finding sheds new light on how different mutations can differentially inhibit trabecular development.

While examining the formation of the trabecular network, we identified a subset of cells in the compact layer that activated the Notch signaling pathway. These cells were found in stripes along the outer curvature of the heart, mirroring the pattern of trabecular development. Through lineage tracing analysis, we determine that these migrate to the trabecular layer by juvenile stages, and proceed to contribute to the outermost cortical layer of cardiomyocytes in the adult heart. Thus, we identify an interesting subpopulation of cardiomyocytes with roles in later stages of development.