Regional regulation of glutaminase by phosphate and calcium was examined in the temporal cortex (TCX), striatum (STR) and hippocampus (HIPP) from adult and aged male F344 rats. Phosphate-dependent glutaminase activity in adult rats was significantly lower (35-43%) in the HIPP (100 and 150 mM) and STR (150 mM) compared to PAG activity in the TCX. Phosphate activation in aged rats was 50-60% lower in the HIPP at concentrations greater than 25 mM compared to the aged TCX or STR. PAG activity in the TCX and STR was unaffected by age, but was significantly reduced (30-50%) in the HIPP from aged rats at phosphate concentrations of 25 mM and greater when compared to adult rats. In adult rats at concentrations of CaCl2 above 1 mM, PAG activity was significantly lower (60-75%) in the STR and HIPP when compared to the TCX. In aged rats, PAG activity (1 mM CaCl2) in the HIPP was significantly less (50%) than STR PAG activity in aged rats. Diminished PAG activity was seen only in the TCX (2.5 mM; 32%), and the HIPP (0.5 mM; 25% and 1 mM; 38%) at higher calcium concentrations compared to adult. Phosphate-independent calcium activation of PAG occurred in the HIPP but not in either the TCX or the STR. Addition of phosphate resulted in a synergistic activation of PAG in the STR and TCX, but not in the HIPP. These findings suggest that PAG is regionally regulated by phosphate and calcium, and this regulation is impaired in aged rats. These data also support the hypothesis that isozymes of PAG exist with different regulatory properties.