Cocultures of neurons and astrocytes as a model for examining hypoxia-induced neuronal death.

Astrocytes perform critical functions necessary for neuronal survival. Thus, examining the influence of astrocyte function on neuronal cell death during disease, including hypoxia/ischemia, has become an important avenue of investigation. In this chapter we detail the methodology and potential pitfalls for establishing cocultures of astrocytes and cortical neurons for studying hypoxia-induced neuronal death. In brief, astrocyte cultures are first established until they reach confluence. The medium is exchanged from a medium that supports astrocyte growth to a medium that supports neuronal viability 24 h before adding neurons to the astrocyte monolayer. After the neurons mature, the cultures are exposed to severe hypoxia and neuronal death is quantified 1-2 days later.