Astrocyte-neuron communication following an ischaemic insult
Ischaemia results from the cessation of blood flow to all or part of the brain, causing a rapid depletion of O2 and impairment of oxidative phosphorylation. Bioenergetic failure follows within minutes, which precipitates directly immediate neuronal death. Upon reperfusion, despite the return of energy substrates, further neuronal death can occur up to several days later, depending on the severity of the initial insult. Astrocytes have been shown to be considerably more resistant to ischaemia/reperfusion injury than neurones and may play either a neuroprotective role or indeed exacerbate the neuronal injury. We have explored the possible interactions between astrocytes and neurones following ischaemia using an in vitro model of ischaemia/reperfusion injury, as a controlled environment that lends itself more easily to manipulation of the numerous variables involved in such an insult. As such we have produced an oxygen glucose deprivation model. We have constructed a chamber in which O2 can be lowered to a concentration of 1 M. We have further developed a primary cortical neuronal culture that is 99% pure and which can survive to at least 10 days in vitro. We have additionally established a novel system for the co-culture of astrocytes and neurones in order to study the communication between these cells in a manner that can allow the complete separation of one cell-type from another. Astrocytes cultured alone do not exhibit signs of cell death during reperfusion of 24hrs duration following ischaemia of up to 2hrs whereas neuron cultures show profound cell death following an ischaemic period of only 15mins. We have co-cultured neurones, which have been subjected to a 15min ischemic insult, with either non-insulted astrocytes or astrocyte conditioned medium during the reperfusion stage. Results show that both astrocytes and astrocyte-conditioned medium enhance neuronal survival. We have finally investigated possible mechanisms to account for this.