AbstractAstrocytes constitute the parenchymal border of the blood–brain barrier (BBB), modulate the exchange of soluble and cellular elements, and are essential for neuronal metabolic support. Thus, astrocytes critically influence neuronal network integrity. In hypoxia, astrocytes upregulate a transcriptional program that has been shown to boost neuroprotection in several models of neurological diseases. We investigated transgenic mice with astrocyte‐specific activation of the hypoxia‐response program by deleting the oxygen sensors, HIF prolyl‐hydroxylase domains 2 and 3 (Phd2/3). We induced astrocytic Phd2/3 deletion after onset of clinical signs in experimental autoimmune encephalomyelitis (EAE) that led to an exacerbation of the disease mediated by massive immune cell infiltration. We found that Phd2/3‐ko astrocytes, though expressing a neuroprotective signature, exhibited a gradual loss of gap‐junctional Connexin‐43 (Cx43), which was induced by vascular endothelial growth factor‐alpha (Vegf‐a) expression. These results provide mechanistic insights into astrocyte biology, their critical role in hypoxic states, and in chronic inflammatory CNS diseases.