Integrin linked kinase (ILK) is involved in the cell-extracellular matrix (ECM) adhesion. Since b1 integrins are relevant for cellular signaling and vasculo-angiogenesis, we determined the role of ILK in this context using embryoid bodies (EBs) derived from ILK deficient (ILK -/-) and wild type (wt) embryonic stem (ES) cells. ILK (-/-) EBs showed changes in the morphology and numbers of endothelial tubes. Major defects in BM formation were observed by analysis of laminin, collagen IV and fibronectin. Since VEGF signaling is critical for vasculogenesis, we analysed next its integrity using [Ca2+]i imaging experiments on magnet-associated cell sorting (MACS)-sorted endothelial cells from wt and ILK (-/-) EBs. Wt cells (n=24) responded to VEGF (20ng/ml) with a [Ca2+]i transient whereas this was absent in most of the ILK (-/-) cells (n=25); by contrast [Ca2+]i transients were evoked in both wt and ILK (-/-) cells by the G protein coupled agonists carbachol (CCh, 1 mM) and bradykinin (100nM). Application of m-3M3FBS, an activator of PLC led to similar changes in the [Ca2+]i signal. The expression and distribution of VEGF receptor 2 resulted unaltered in the ILK (-/-) cells compared to wt. Thus, ILK is important for embryonic vascular development by affecting ECM stability and by regulating morphogenesis. Moreover the compromised vascular development appears to be due to defects of tyrosine kinase receptor-mediated signaling.