DEATH OF SEPTAL CHOLINERGIC NEURONS PRODUCED BY CHRONIC EXPOSURE TO GLUTAMATE IS PREVENTED BY THE NONCOMPETITIVE NMDA RECEPTOR-CHANNEL ANTAGONIST, MK-801 - ROLE OF NERVE GROWTH-FACTOR AND NITRIC-OXIDE

To study the sequence of degenerative events possibly associated with cholinergic cell death in Alzheimer's disease, septal cholinergic neur ons derived from rat embryonic brains were exposed to chronic excitoto xic stress by glutamate, Counts of choline acetyltransferase (ChAT)-im munopositive neurons and measurement of ChAT activity revealed that co ncentrations of glutamate on the order of 70 mu M killed 50% of cholin ergic neurons after 24 hr of treatment, Neurotoxic effects were not ai med at cholinergic neurons specifically, since other populations of ce lls present in these cultures were also affected at similar concentrat ions, The noncompetitive N-methyl-D-aspartate (NMDA) receptor channel antagonist MK-801 (10 mu M) abolished acute neuronal swelling and resc ued from late degeneration both cholinergic and noncholinergic cells w hen concentrations of glutamate up to 500 mu M were added to the cultu res, Protective effects declined progressively with increasing concent rations of the amino acid, even when MK-801 was raised to its highest nontoxic levels, e.g., 50 mu M. The kainate/quisqualate receptor antag onist CNQX provided no protection alone or in combination with MK-801, Nerve growth factor (NGF), used in standard culture conditions to sti mulate the expression of the cholinergic phenotype, was shown not to i nfluence excitotoxic neurodegenerative changes, Several observations s uggested that nitric oxide (NO) may act as an intercellular messenger of NMDA-mediated cell death in septal cultures: 1) Most of the choline rgic neurons contained the NO synthase enzyme as characterized by NADP H-diaphorase (NADPH-d) staining; 2) sodium nitroprusside (SNP) [a chem ical with the ability of generating NO] was capable of mimicking some of the aspects of the glutamate-induced degenerative process; 3) the r ise in cyclic GMP which was observed in the presence of toxic levels o f glutamate and which is usually taken as an index of NO production, w as antagonized by MK-801 and by the inhibitor of the NO synthase enzym e, L-NOARG. Yet, the fact that L-NOARG and its congener, L-NAME, were ineffective in preventing glutamate-induced neurodegenerative changes in our culture system did not substantiate our working hypothesis, Alt ogether these results suggest that glutamate-induced cholinergic neuro nal death is the consequence of an overstimulation of NMDA receptors a nd that neither NGF nor NO plays a key role in the degenerative proces s. (C) 1995 Wiley-Liss, Inc.