To study the changes in the hippocampal cholinergic system of chronologically old and behaviorally impaired animals, old (21 months of age) and young (3 months of age) male, Fischer-344 rats were used. The aged animals were tested on a reference memory task (Morris water maze) and found to be functionally impaired as compared to the young controls. Carbachol-stimulated phosphoinositide metabolism was measured in hippocampal slices from young and old rats. Slices were prelabeled with (3)H-inositol for 120 min and subjected to muscarinic stimulation in the presence of lithium. Following extraction of the slices with acidified solvent mixture, the inositolphosphates present in the aqueous fraction were isolated by ion exchange chromatography. Receptor-stimulated release of inositolphosphates (IPs) was found to be increased in the hippocampus of older animals. The age-related enhancement of IP release was in contrast to the decrease in choline acetyltransferase (CHAT) activity in the hippocampus. The authors postulate that alterations in the G-protein coupling with the muscarinic receptor leads to an increase in the phosphoinositide turnover in part as a compensatory mechanism for neuronal cell death and reduced transmitter levels.