Prevention of dopaminergic neuronal death by cyclic AMP in mixed neuronal/glial mesencephalic cultures requires the repression of presumptive astrocytes

Cyclic AMP-elevating agents are highly effective in preventing the loss of dopaminergic neurons that occurs spontaneously in neuronal-glial mesencephalic cultures. We demonstrate here that cAMP causes a concomitant decline in the number of dividing non-neuronal cells, suggesting that inhibition of proliferation contributes to neuroprotection. Consistent with this hypothesis, a transient treatment with the antimitotic cytosine arabinoside, at concentrations that induce long-term repression of glial cell proliferation, mimicked the neuroprotective action of cAMP and also obviated the need for the cyclic nucleotide. Treatment with cAMP-elevating agents reduced the population of OX-42-positive microglial cells and the number of immature astrocytes expressing vimentin and low levels of the astrocytic marker glial fibrillary acidic protein. The effect on the immature astrocytes, however, seemed essential for neuroprotection. Ciliary neurotrophic factor and leukemia inhibitory factor, which stimulate astrocyte differentiation without reducing cell proliferation, failed to reproduce the protective effects of the cyclic nucleotide. Cyclic AMP did not operate by counteracting the action of the astrocyte mitogen epidermal growth factor or by reducing activation of the mitogen-activated protein kinase signaling pathway. The neuroprotective and antiproliferative actions of cAMP, however, were closely mimicked by olomoucine and roscovitine, potent inhibitors of the cyclin-dependent kinase CDK1 that are structurally related to cAMP. Measurement of CDK1 activity confirmed that neuroprotection was closely correlated with inhibition of this kinase by cAMP. In summary, neuroprotection of mesencephalic dopaminergic neurons by cAMP most probably requires the repression of presumptive astrocytes through inhibition of CDK1.