Abstract Cholinergic deficits occur in the aged hippocampus and they are significant in Alzheimer's disease. Using stereological and biochemical approaches, we characterized the cholinergic septohippocampal pathway in old (24 months) and young adult (3 months) rats. The total length of choline acetyltransferase (ChAT)-positive fibers in the dorsal hippocampus was significantly decreased by 32% with aging ( F (1,9) = 20.94, p = 0.0014), along with the levels of synaptophysin, a presynaptic marker. No significant changes were detected in ChAT activity or in the amounts of ChAT protein, nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF), tropomyosin related kinase receptor (Trk) A, TrkB, or p75 neurotrophin receptor (p75 NTR) in the aged dorsal hippocampus. The number and size of ChAT-positive neurons and the levels of ChAT activity, NGF and BDNF were not statistically different in the septum of aged and young adult rats. This study suggests that substantial synaptic loss and cholinergic axonal degeneration occurs during aging and reinforces the importance of therapies that can protect axons and promote their growth in order to restore cholinergic neurotransmission.