The Neurovascular Unit (NVU), comprised of glia (astrocytes, oligodendrocytes, microglia), neurons, pericytes and endothelial cells, is a dynamic interface ensuring physiological functioning of the central nervous system (CNS), that gets affected and contributes to the pathology of several neurodegenerative diseases. Neuroinflammation is a common feature of neurodegenerative diseases and is primarily related to the activation state of perivascular microglia and astrocytes, which constitute two of its major cellular components. Our studies focus on monitoring in real time the morphological changes of perivascular astrocytes and microglia, as well as their dynamic interactions with the brain vasculature, under physiological conditions and following systemic neuroinflammation triggering both microgliosis and astrogliosis. To this end, we performed 2-photon laser scanning microscopy (2P-LSM) for intravital imaging of the cortex of transgenic mice visualizing the dynamics of microglia and astroglia following neuroinflammation induced by systemic administration of the endotoxin lipopolysaccharide (LPS). Our results indicate that following neuroinflammation the end-feet of activated perivascular astrocytes lose their close proximity and physiological crosstalk with vasculature, an event that most possibly contributes to a loss of blood-brain-barrier (BBB) integrity. At the same time, microglial cells get activated with more frequent contacts of the blood vessels. These dynamic responses of perivascular astrocytes and microglial are peaking at 4 days following LPS administration, however they still persist at a lower level 8 days after LPS, revealing incomplete reversal of inflammation affecting the glial properties and interactions with the NVU.