EFFECT OF BILIRUBIN INFUSION ON THE GLUTAMATE SITE OF THE N-METHYL-D-ASPARTATE (NMDA) RECEPTOR IN THE BRAIN OF THE NEWBORN PIGLET† 1280

Numerous mechanisms have been proposed to describe the neurotoxic effects of bilirubin. Previous studies have shown that the administration of bilirubinin vivo to newborn piglets increased the affinity (decreased Kd) of the NMDA receptor for ³[H]MK-801. The present study tests the hypothesis that modification of the NMDA receptor by bilirubin is specifically dependent on alteration of the glutamate site of the NMDA receptor/ion channel complex. Studies were performed in 2-4 day old anesthetized and ventilated piglets. Piglets in the bilirubin-exposed [B] group (n=5) received a 35 mg/kg bolus of bilirubin followed by a four hour continuous infusion (25 mg/kg/h) containing 0.1 N NaOH, 5% albumin and 0.055 Na₂HPO₄ with 3 mg/ml of bilirubin. The control [C] group (n=5) received a bilirubin-free buffer solution. Sulfisoxazole, 160 mg/kg, was injected into each animal in three divided doses. P2 membrane fractions were prepared from the cerebral cortex and ³H-glutamate binding was performed in a concentration range from 25 nM to 1000 nM. Specific NMDA-displaceable ³H-glutamate binding was obtained in the presence of NMDA (100μM). Non-specific binding was performed in the presence of unlabelled glutamate (1mM). The number of NMDA-specific glutamate sites (Bmax) in the B and C groups were 579 ± 60 and 522 ± 107 fmol/mg protein (mean±SD). Kd in the same groups were 252 ± 133 and 293 ± 120 nM. The data show that bilirubin administration to piglets did not modify the glutamate site of the NMDA receptor, in contrast to the decrease in Kd from 6.21 to 3.97 nM (higher affinity) of the receptor channel for MK801 previously reported. We speculate that bilirubin neurotoxicity may be due to the incorporation of bilirubin molecules in neuronal membranes resulting in conformational alteration of the MK-801 binding site within the ion channel. (Funded by NIH-HD-20337, MOD-FY94-0135, UCPR 506-93)