INTRAUTERINE HYPOXIA-ISCHEMIA INCREASES N-METHYL-D-ASPARTATE-INDUCED CGMP FORMATION AND GLUTAMATE ACCUMULATION IN CULTURED RAT CEREBELLAR GRANULE CELLS

Effects of intrauterine hypoxia-ischemia (HI) on brain functional development in the term rat were examined in cerebellar granule cell cultures obtained from an in utero HI model. On gestation d 17, HI conditions were achieved by complete clamping of the uterine vasculature for designated durations followed by removal of the clamps to permit reperfusion. Sham operation (surgery without vasculature clamping) was performed as the control. After surgery, the uterine horns were returned to dam's abdomen to let the pups deliver naturally. Severe HI insult from the surgical manipulation was evident in that the lactate levels of fetal brain increased, and fetal blood pH decreased with the duration of vasculature clamping up to 1 h. The experimental KI insult up to 1 h did not affect fetal survival rate, but retarded growth of fetuses or newborns was observed in the 1 h HI group. N-Methyl-D-aspartate (NMDA)- and kainate (KA)-stimulated cGMP formation and glutamate accumulation were measured in cerebellar granule cell cultures from 8-d-old pups suffering from various durations of antenatal HI insult. NMDA (100 mu M)-induced elevation of cGMP was further augmented by a 10-35-min HI insult as compared with the control cells (62.4-78.2 versus 49 pmol/mg protein). In the presence of L-N-G-monomethyl-arginine (L-NMMA, 150 mu M), a nitric oxide synthase inhibitor, NMDA-induced cGMP formation was blocked, and the blockade of cGMP formation by L-NMMA (10 mu M) could be reversed by simultaneous application of 1 mM arginine in both control and HI cells. Antenatal HI insult (20-35 min) also augmented NMDA-, but not KA-, stimulated accumulation of extracellular glutamate. The hypersensitive response in NMDA-induced glutamate accumulation could be suppressed by 150 mu M L-NMMA. The overall results suggest that antenatal HI occurring in the last half of gestation may result chronically in adverse effects on NMDA receptor-mediated neurotransmission and that nitric oxide is possibly involved in these effects.