Organ culture studies as well as mammalian models have also shown that doxorubicin exerts harmful effects on vascular endothelium, leading to impaired vasodilatory response of arteries. It appears that administration of a single dose of doxorubicin in rabbits is associated with rapid deterioration of endothelium-dependent and independent vascular responses. Furthermore, administration of doxorubicin in humans is associated with acute reduction of flow-mediated dilatation in the brachial arteries and of nitric oxide level in the plasma. High-frequency ultrasound with enhanced contrast agents enables in vivo imaging and analysis of blood perfusion. Microbubbles are a contrast agent that enhances the acoustic signal of blood in the circulation; they are small enough to move freely through the bloodstream, and are used as markers for visualization and quantification of regional microvasculature. Fibred confocal fluorescence microscopy was designed for in vivo imaging of fluorescent signals in living animals. The FCFM with its optical mini-probes enables in vivo fluorescent visualization of microvasculature with a minimal invasive intervention. We have set up a platform of live, high-resolution molecular mice imaging, suitable for capturing vessels’ characteristics, arterial blood flow and organs blood volume. This imaging setup enables us to detect acute, real-time, treatment-induced effects within the same individuals and follow them over a period of time. Using both imaging tools, we could observe that doxorubicin had an effect on blood vessels already 3 minutes after administration. The acute reduction in gonadal and femoral blood flow and the impairment of the blood vessels wall may represent an acute universal doxorubicin-related vascular toxicity, an initial event in organ injury. Of all the chemotherapy-induced side effects, the direct vascular injury is the least characterized. The vascular endothelium is an essential barrier that protects the tissues integrity, regulates the homeostasis of water and solvents between the plasma and the tissues and plays a role in the regulation of arterial vasomotor tone. Impairment of the vascular endothelium may result in disintegration of the blood vessel wall and leakage of fluids from the blood into the extracellular matrix, compromising organ function. Studies addressing the vascular toxicity of certain chemotherapeutic agents indicated that they were more toxic to endothelial cells than to tumor cells.