Attenuation by PACAP of glutamate-induced neurotoxicity in cultured retinal neurons

The effects of pituitary adenylate cyclase activating polypeptides (PACAPs: PACAP27, PACAP38) on glutamate-induced neurotoxicity were examined using c ultured retinal neurons obtained from 3- to 5-day old Wistar rats. Cell via bility was evaluated by double staining with fluorescein diacetate and prop idium iodide. Effects of PACAPs on the increase in intracellular Ca2+ conce ntration ([Ca2+](i)) in retinal neurons was investigated using the Ca2+ ima ge analyzing system with fura-2. The cAMP contents and the mitogen-activate d protein (MAP) kinase activity in retinal cultures were measured by radioi mmunoassay. Concomitant application of PACAPs (10 nM(-1) mu M) with glutama te (1 mM) for 10 min inhibited the delayed death of retinal neurons, which was observed 24 h after glutamate (1 mM) treatment in a dose-dependent mann er. Protection by PACAPs (100 nM) against glutamate-induced neurotoxicity w as antagonized by PACAP6-38 (1 mu M), a PACAP antagonist, and H-89 (1 mu M) , a protein kinase A (PKA) inhibitor. However, PACAPs did not affect the gl utamate-induced increase in [Ca2+](i), but PACAPs (1-100 nM) increased the cAMP levels in a dose-dependent manner. In addition, activation of MAP kina se by PACAP38 (1 mu M) was inhibited by simultaneous application with H-89 (I mu M) These findings suggest that PACAPs attenuate glutamate-induced del ayed neurotoxicity in cultured retinal neurons by activating MAP kinase thr ough the activation of cAMP-stimulated PKA. (C) 1999 Published by Elsevier Science B.V. All rights reserved.