Cardiovascular diseases are the leading causes of morbidity and mortality in developed countries. One such disease is left ventricular noncompaction (LVNC, OMIM604169). The prevalence of LVNC reportedly ranged from 0.01% to 0.3%, and is higher in patients with heart failure, reportedly 3% to 4%. Genetic inheritance occurs in at least 30?50% of patients and genes that cause LVNC include the genes that encode sarcomeric or cytoskeletal proteins and Notch signaling pathway genes. In LVNC, trabeculae fail to undergo compaction. Trabeculae are sheet-like structures extending from the myocardium to the heart lumen that function to increase surface area when the coronary circulation system is not developed yet. A lack of trabeculation causes embryonic lethality, and excess trabeculation causes LVNC and heart failure in humans. Our studies demonstrated that the orientated cell division and directional migration of cardiomyocytes in the single-cell- thick myocardium contribute to trabecular initiation by forming a multiple-layer myocardium. Then endocardial cells (EndCs) invade the multiple layered myocardium and allocate cardiomyocytes to form trabeculae before E9.5. Many mutations of contractile protein genes correlate with LVNC, and it has been speculated that the contractile function of the myocardium is required for ventricular compaction; however, no genetic models have been made to test this hypothesis. The subsequent questions in this field will be what are the mechanisms that regulate the trabecular formation, de novo trabeculation, and ventricular compaction. Our preliminary data show that early the Itgb1 gene causes defects in trabecular formation, growth, Notch1 activation and deposition of Fibronectin (Fn), and late deletion causes defects in compaction and Notch1 activation. Furthermore, global deletion of the Itga5 gene or Fn1 gene causes defects in cardiovascular morphogenesis, but early lethality prevented the study of their functions in trabecular morphogenesis. Our heart-specific deletion of preliminary data show that heart-specific Itga6 gene (encoding integrin ?6 subunit or ?6), which is expressed specifically in the trabecular zone during compaction, reduces cardiac contractile function. Therefore, our central hypothesis is that integrin ?5?1 and ?6?1 have distinct functions in cardiac morphogenesis with ?5?1 regulating trabecular formation and growth, and integrin ?6?1 regulating ventricular contraction and the subsequent compaction and Notch1 activation. There are two aims to test the hypothesis. Genetic tools such as trabecular specific and compact zone specific Cre lines will be used to test these hypotheses. We have developed some unique expertise, e.g., single cell lineage tracing assay to study the mechanisms of trabeculation; ECHO to measure the contractility of embryonic hearts treated with drug or vehicle; rAAV9 system to rescue the LVNC defect. Completion of the studies will determine whether ?5?1 axis regulates the trabecular formation and ?6?1 regulates compaction in vivo and determine whether enhancement of contraction and activation of Notch1 can rescue the LVNC cardiomyopathy. deletion of the