The present study tests the hypothesis that during graded
hypoxia,
N-methyl-D-aspartate (
NMDA) receptor expression and phosphorylation are altered in the cerebral cortex of newborn piglets. Studies were performed in anesthetized, ventilated piglets, 6 normoxic and 9 exposed to different lengths of decreased fractions of inspired
oxygen to achieve varying biochemical levels of
phosphocreatine (PCr).
P(2)
membrane proteins were immunoprecipitated with antiphosphoserine, antiphosphotyrosine, or antiphosphothreonine
antibodies and separated by
sodium dodecyl sulfate polyacrylamide gel electrophoresis.
Proteins were transblotted and probed with
NMDA receptor subunit 1 (NR1), NR2A or NR2B
antibodies. As tissue PCr levels decreased from 3.5 to 0.5 micromol/g brain during
hypoxia, NR1, NR2A and NR2B
protein increased by 84, 56 and 38%, respectively. Phosphorylated
serine,
tyrosine and
threonine residues also increased during
hypoxia on the three subunits. However, the increase in
subunit protein exceeded the increase in phosphorylated residues for all three subunits. Therefore, the ratio of phosphorylated/dephosphorylated
serine,
tyrosine and
threonine residues decreased with worsening
hypoxia. We speculate that an alteration in the ratio of phosphorylated/dephosphorylated residues of the
NMDA receptor may regulate receptor activation during
hypoxia.