EFFECTS OF NEUROPROTECTIVE DOSE OF FRUCTOSE-1,6-BISPHOSPHATE ON HYPOXIA-INDUCED EXPRESSION OF C-FOS AND HSP70 MESSENGER-RNA IN NEONATAL RATCEREBROCORTICAL SLICES

In situ hybridization (ISH) measurements of c-fos and hsp70 expression were made in brain slice studies of hypoxia, with or without fructose -1,6-bisphosphate (FBP) pretreatment. Each experiment used eighty 350 mu m thick cerebrocortical slices, obtained from twenty 7-day old rats . Thirty minute periods of hypoxia were followed by 8 h of hyperoxic p erfusion. Slices were removed at eight predetermined times, and proces sed for ISH and immunohistochemistry. In three of six hypoxia experime nts, slices were pretreated for 60 min with 2 mM FBP, a condition know n to maintain ATP level in brain slices during hypoxia. In three other hypoxia experiments slices received no pretreatment. In two control e xperiments slices were perfused for 11.5 h without hypoxia. In control experiments, hsp70 mRNA was barely detectable in slices at all times, although moderate c-fos mRNA expression occurred at 1 h after decapit ation. Hypoxia produced a modest but statistically significant increas e in c-fos mRNA and hsp70 mRNA induction 4 h following reoxygenation. At all times after hypoxia, FBP pretreatment reduced expression of c-f os and hsp70 mRNA. The absence of hsp70 mRNA in control slices suggest s that intracellular protein denaturation was minimal in this preparat ion. In slices made hypoxic, the decrease in c-fos and hsp70 mRNA caus ed by FBP pretreatment suggests ameliorated progression towards injury . Immunohistochemistry showed no HSP70 protein at any time following h ypoxia, with or without FBP pretreatment, presumably due to delayed HS P70 protein synthesis, or to a block in translation, as observed in vi vo in other studies.