After direct administration into the dorsal hippocampus nicotine decreased the time spent in social interaction, without changing locomotor activity, indicating an anxiogenic effect. The possibility that post-synaptic M1 muscarinic receptors mediated this effect was examined by determining whether dorsal hippocampal administration of a specific M1 receptor agonist (McN-A-343) had anxiogenic effects, and whether the anxiogenic effect of nicotine could be reversed by co-administration of the M1 receptor antagonist, pirenzepine. McN-A-343 (0.3, 1.6, 3.2, 15.8 nmol) was without effect on social interaction, and pirenzepine (0.7 and 2.4 nmol) injection into the dorsal hippocampus failed to reverse the decrease in social interaction caused by nicotine (6.3 nmol) injection into this area. However, the decrease in social interaction after nicotine (50 nmol) was completely reversed by the specific 5-HT1A receptor antagonist, WAY 100635 (0.4 nmol) after co-administration of both drugs into the dorsal hippocampus. Thus, the anxiogenic effect of nicotine in this brain region seems to be mediated by 5-HT1A, but not M1, receptors. In contrast to the effect of nicotine in naive animals, those retested after a second injection of 50 nmol did not show a significant anxiogenic effect. The theoretical implications of this are discussed and from a practical point of view this suggests caution in the retesting of animals after central injections.