The effect of radiation therapy on tumour vasculature has been a subject of debate over the years. Increased oxygenation and perfusion have been documented during radiation treatments. Conversely, death of irradiated tumour endothelial cells by apoptosis, predicted to impair blood flow, has been proposed as a major determinant of tumour control. To approach these contradictions, we used video 2-photon microscopy to study the vascular response to irradiation in two different murine tumour models: MC38, a highly vascularized model with many non-perfused vessels and B16F10, a moderately vascularized model, both grown in transgenic mice with endothelium marked by the fluorescent protein tdTomato. These tumours received either a single dose of radiation (15 Gy) or a fractionated dose (5 × 3 Gy). Unexpectedly, even these high doses led to little structural change of the perfused vasculature. However, non-perfused vessels and blind ends were substantially impaired after radiation accompanied by apoptosis of their endothelium. These changes did not alter perfusion of the less vascularized B16F10 tumour, but led to increased perfusion in the more vascular MC38 tumour. After irradiation, the proliferation of endothelial cells was blocked resulting in diminished numbers of sprouts. Further, endothelial cells from control and irradiated tumours were subjected to RNA sequencing, which confirmed that irradiation upregulates the expression of genes in apoptotic pathways as well as cell cycle regulation. We now explain how apoptosis of tumour endothelial cells after radiation is not detrimental to function, because the apoptosis occurs primarily in the endothelium of non-functional small or blunt end non-perfused vessels leaving the functional vessels intact.