Increased synthesis of a rat hippocampal protein with an apparent molecular weight (Mr) of 35,000 Da occurs in response to elevation of serum corticosterone levels. Subcellular fractionation has localized this protein in the cytosol. Two-dimensional gel electrophoresis indicated that this protein has an isoelectric point (IEP) of 6.6. A similar protein in liver has a slightly higher Mr and an IEP of 6.8. Increased synthesis of one additional hippocampal protein with an Mr of 46,000 Da and an IEP of 6.2 and of two other liver proteins, one with an Mr of 53,000 Da and an IEP of 6.2 and the other with an Mr of 45,000 Da and a range of IEPs from 8.7 to 7.8, was also seen after injection of corticosterone into rats. One possible identity of the 35,000 Da protein is glycerol 3-phosphate dehydrogenase (GPDH), based upon the reported Mr and IEP of this enzyme. The 35,000 Da hippocampal protein co-eluted from a gel filtration column with GPDH activity. No alteration of hippocampal GPDH activity was seen in intact rats 4 or 24 h after injection of either corticosterone or the type II receptor-specific agonist RU 28362. However, daily administration of corticosterone to rats beginning 10 days after adrenalectomy returned hippocampal GPDH activity to normal values after 2-3 days. In contrast, synthesis of the 35,000 Da protein was maximally increased 4 h after a single injection of steroid and not elevated at later times.