Cyclothiazide and GYKI 52466 modulate AMPA receptor-mediated apoptosis in cortical neuronal cultures

In neocortical neuronal cultures, (S)-AMPA caused neurotoxicity which was concentration-dependent, receptor-mediated, slow and apoptotic in nature. (S)-AMPA (3-600 microM) failed to produce rapid neuronal swelling, but morphological observations and monitoring of viability at 24-72 h revealed 50% cell death consistent with apoptosis. (S)-AMPA induced cell shrinkage, neurite blebbing and nuclear condensation. Cyclothiazide (50 and 100 microM), which blocks AMPA receptor desensitization potentiated excitotoxicity with 75% of neurones undergoing slow death. The AMPA-selective antagonist GYKI 52466 (10-50 microM), attenuated (S)-AMPA-mediated neurotoxicity. DNA condensation, a hallmark of apoptosis, was found by labelling neurones with the DNA binding dye 4,6-diamidino-2-phenylindole HCl (DAPI). Gel electrophoresis revealed DNA fragmentation, which was increased by cyclothiazide and reduced by GYKI 52466 and cycloheximide. Overstimulation of the AMPA receptor produces a novel form of neuronal death, which is apoptotic, very slow in nature, and which could contribute to various neuropathologies.