Although the importance of the septohippocampal projections in learning and memory is generally accepted, the degree to which damage to particular set of septohippocampal projections contributes to deficits within specific cognitive domains is less clear. In the present study, using novelty-preference paradigm, we tested the behavioral effects of electrolytic, neurotoxic (ibotenic acid) or immunotoxic (192 IgG-saporin) lesions of medial septum (MS) to gain insights into the differential roles of septohippocampal cholinergic and noncholinergic projections in exploratory behavior and recognition memory. A total of 39 male Wistar rats (Department of Animal Care, I. Beritashvili Center of Experimental Biomedicine) were used in the present study. All experiments were approved by the Animal Care and Use Committee of the Center and were in accordance with the principles of laboratory animal care. The size and location of the electrolytic and ibotenic acid lesions were determined by microscopic examination of serial coronal sections (25 µm) stained with cresyl violet. The immunotoxic lesions of MS were verified by observing decreased Acetylcholinesterase (AChE) staining of the MS and hippocampal sections. Statistical analysis was performed using ANOVA. The results indicate that the MS lesioned rats did not differ from control rats in the activity level measured by grid crossing. The MS electrolytic lesioned rats were impaired in habituating to the environment in the repeated spatial environment, but rats with immuno- or neurotoxic lesions of the MS did not differ from control ones. The MS electrolytic and ibotenic acid lesioned rats showed an increase in their exploratory activity to the objects and were impaired in habituating to the objects in the repeated spatial environment; rats with immunolesions of the MS did not differ from control rats. The electrolytic lesions of the MS disrupt spatial recognition memory; rats with immuno- or neurotoxic lesions of the MS were normal in detecting spatial novelty; all of the MS-lesioned and control rats clearly reacted to the object novelty by exploring the new object more than familiar ones. In conclusion, our results observed across lesion techniques demonstrates dissociation between the two major components (cholinergic and noncholinergic) of the septohippocampal pathway in exploratory behavior assessed in the open field and indicate that: deficits after nonselective damage of MS are limited to a subset of cognitive processes dependent on the hippocampus; the selective loss of septohippocampal cholinergic or noncholinergic projections does not disrupt the function of the hippocampus to a sufficient extent to impair spatial recognition memory.