Traumatic brain injury (TBI) initiates a complex genetic response that may include the expression of organelle specific stress genes. We investigated the effects of brain trauma on the expression of a number of stress genes by in situ hybridization and Western blot analysis including the endoplasmic reticulum (ER) stress gene grp78, ER protein processing enzymes calnexin and protein disulphide isomerase (PDI), the mitochondrial stress gene hsp60, and the cytoplasmic stress gene hsp70. Male Sprague-Dawley rats were subjected either to sham-surgery or moderate (1.8-2.2 atm) parasagittal fluid-percussion (F-P) brain injury followed by 30 min of either normoxic or hypoxic (30-40 mm Hg) gas levels. Expression of grp78 was increased in the ipsilateral cerebral cortex and dentate gyrus beginning 4 h after trauma plus hypoxia. Similarly, mRNA encoding the mitochondrial hsp60 was induced in the ipsilateral outer cortical layers at 4-24 h after TBI plus hypoxia. Calnexin and PDI mRNAs were not significantly altered following TBI with or without secondary hypoxia. In contrast, mRNA of the cytoplasmic hsp70 was strongly induced at 4 h after brain injury in multiple brain regions within the injured hemisphere, and this expression was greatly enhanced by secondary hypoxia. Because subcellular stress gene expression may reflect where unfolded or damaged proteins are abundant, these findings suggest that abnormal proteins are localized mainly in the cytoplasm, and to a lesser degree in the ER lumen and mitochondria after brain trauma. Thus, distinct parts of the cellular machinery respond to traumatic and metabolic stresses in specific ways.