In the present study, variations of cholinergic and GABAergic markers in the medial septum/vertical limb of the diagonal band of Broca (MS/vDB) and the hippocampus of eight different inbred mouse strains were investigated. By means of immunocytochemistry against the acetylcholine-synthesizing enzyme choline acetyltransferase (ChAT), the cholinergic neurons were visualized and the number of ChAT-positive neuronal profiles in the MS/vDB was counted. Cholinergic and GABAergic septo-hippocampal projection neurons were detected with a combined retrograde tracing and immunocytochemical approach. In order to quantify the cholinergic innervation of various hippocampal sub-regions, we estimated the density of acetylcholinesterase (AChE)-containing fibers as visualized by AChE histochemistry. Additionally, the densities of muscarinic receptors (mainly the subtypes M1 and M2) in different hippocampal areas of seven inbred strains were measured by means of quantitative receptor autoradiography. We found significant strain differences for the number of ChAT-positive neurons in the MS/vDB; in the numbers of cholinergic septo-hippocampal projection neurons; in the density of cholinergic fibers in hippocampal subfields CA3c, CA1, and in the dentate gyrus; and in the density of muscarinic receptors in the hippocampus. In contrast the GABAergic component of the septo-hippocampal projection did not differ between the strains investigated. The number of ChAT-reactive neurons in the MS/vDB was not correlated with either hippocampal cholinergic markers. This might be attributed to different collateralization of cholinergic neurons or to different projections of these neurons to other brain regions. These results show a strong hereditary variability within the septo-hippocampal cholinergic system in mice. In view of the role of the cholinergic system in learning and memory processes, strain differences in cholinergic markers might be helpful in explaining behavioral variation.