Tamoxifen-inducible gene targeting in mice with estrogen receptor-dependent Cre (CreER) recombinase has enormously advanced vascular biology research. However, CreER activation under the control of vascular endothelial promoters causes off-target effects that impair the vascularization of the retina, a widely used model to study the molecular and cellular mechanisms of angiogenesis. Although ubiquitously expressed CreER is also used to study retinal angiogenesis, it remains unknown whether it causes similar or more severe off-target effects compared to endothelial-selective CreER activation. Moreover, the molecular and cellular processes disrupted by CreER activation in endothelial cells remain to be identified. Here, we demonstrate that ubiquitous CreER activation in postnatal mice decreases body growth and impairs retinal angiogenesis. We further show that CreER activation slows endothelial cell proliferation and promotes apoptosis concomitant with p21/CDKN1A upregulation in the angiogenic retina, although retinal vasculature progressively normalizes. By contrast, CreER activation in quiescent adult retinal vasculature did not induce endothelial cell apoptosis or p21 activation. Altogether, our findings indicate that ubiquitous promoters should be avoided to drive CreER expression when studying gene function during retinal angiogenesis, and that CreER activation controls, although essential to account for endothelial apoptosis and proliferation defects in short-term studies, may be less critical for adult vascular studies.