Protection by imidazol(ine) drugs and agmatine of glutamate-induced neurotoxicity in cultured cerebellar granule cells through blockade of NMDA receptor

1 This study was designed to assess the potential neuroprotective effect of several imidazol(ine) drugs and agmatine on glutamate-induced necrosis and on apoptosis induced by low extracellular K+ in cultured cerebellar granul e cells. 2 Exposure (30 min) of energy deprived cells to L-glutamate (1-100 mu M) ca used a concentration-dependent neurotoxicity, as determined 24 h later by a decrease in the ability of the cells to metabolize 3-(4,5-dimethylthiazol- 2-yl)-2,5-diphenyltetrazoliumbromide (MTT) into a reduced formazan product. L-glutamate-induced neurotoxicity (EC50=5 mu M) was blocked by the specifi c NMDA receptor antagonist Mg-801 (dizocilpine). 3 Imidazol(ine) drugs and agmatine fully prevented neurotoxicity induced by 20 mu M (EC100) L-glutamate with the rank order (EC50 in mu M): antazoline (13)> cirazoline (44)> LSL 61122 [2-styryl-2-imidazoline] (54)>LSL 60101 [ 2-(2-benzofuranyl) imidazole] (75)>idazoxan (90)>LSL 60129 [2(1,4-benzodiox an-6-yl)-4,5-dihydroimidazole (101) > RX821002 (2-methoxy idazoxan) (106)> agmatine (196). No neuroprotective effect of these drugs was observed in a model of apoptotic neuronal cell death (reduction of extracellular K+) whic h does not involve stimulation of NMDA receptors. 4 Imidazol(ine) drugs and agmatine fully inhibited [H-3]-(+)-MK-801 binding to the phencyclidine site of NMDA receptors in rat brain. The profile of d rug potency protecting against L-glutamate neurotoxicity correlated well (r =0.90) with the potency of the same compounds competing against [H-3]-(+)-M K-801 binding. 5 In HEK-293 cells transfected to express the NR1-1a and NR2C subunits of t he NMDA receptor, antazoline and agmatine produced a voltage- and concentra tion-dependent block of glutamate-induced currents. Analysis of the voltage dependence of the block was consistent with the presence of a binding site for antazoline located within the NMDA channel pore with an IC50 of 10-12 mu M at 0 mV. 6 It is concluded that imidazol(ine) drugs and agmatine are neuroprotective against glutamate-induced necrotic neuronal cell death in vitro and that t his effect is mediated through NMDA receptor blockade by interacting with a site located within the NMDA channel pore.