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Glutamate is the most abundant excitatory amino acid in the central nervous system. It has also been described as a potent toxin when present in high concentrations because excessive stimulation of its receptors leads to neuronal death. Glial influence on neuronal survival has already been shown in the central nervous system, but the mechanisms underlying glial neuroprotection are only partly known. When cells isolated from newborn rat retina were maintained in culture as enriched neuronal populations, 80% of the cells were destroyed by application of excitotoxic concentrations of glutamate. Massive neuronal death was also observed in newborn retinal cultures containing large numbers of glia, or when neurons were seeded onto feeder layers of purified cells prepared from immature (postnatal 8 day) rat retina. When newborn retinal neurons were seeded onto feeder layers of purified glial cells prepared from adult retinas, application of excitotoxic amino acids no longer led to neuronal death. Furthermore, neuronal death was not observed in mixed neuron/glial cultures prepared from adult retina. However, in all cases (newborn and adult) application of kainate led to amacrine cell-specific death. Activity of glutamine synthetase, a key glial enzyme involved in glutamate detoxification, was assayed in these cultures in the presence or absence of exogenous glutamate. Whereas pure glial cultures alone (from young or adult retina) showed low activity that was not stimulated by glutamate addition, mixed or co-cultured neurons and adult glia exhibited up to threefold higher levels of activity following glutamate treatment. These data indicate that two conditions must be satisfied to observe glial neuroprotection: maturation of glutamine synthetase expression, and neuron-glial signalling through glutamate-elicited responses.

Glutamate, the principal retinal neurotransmitter, can also act as a toxin when present in excessive concentrations as may occur in pathologies such as retinal ischemia or more generally in cerebral neuronal degenerative disease. As glial cells play pivotal roles in transfer of blood-borne molecules and in glutamate clearance, we investigated the effects of the excitatory amino acids glutamic and kainic acid on different in vitro preparations of retinal Müller glial cells. Glial viability or morphology were not influenced by excitatory amino acid exposure in either pure glial cultures or in monolayer cultures of mixed neonatal neurons and glia, whereas kainic acid specifically lysed amacrine cells in mixed or pure neuronal cultures. When retinal fragments were pre-incubated in excitatory amino acids prior to dissociation and seeding into culture, under these conditions Müller glial cells exhibited a dramatic loss of their normal epithelioid form to a retracted morphology. However, glial cell viability was not compromised, and rapid restoration of epithelioid in vitro glial morphology could be achieved by addition of exogenous epidermal and basic fibroblast growth factor to the culture medium. This study demonstrates that glial cells are structurally perturbed by excitotoxic conditions and that such effects are dependent on normal glial-neuronal interactions.