Although nicotine is known to protect against β-amyloid (Aβ)-induced neurotoxicity, the effect of nicotine on colchicine-induced neurotoxicity remains unknown. Colchicine is a microtubule-interfering agent and is able to induce neural apoptosis. Here we investigated whether nicotine exhibits similar neuroprotective effects and the mechanism against colchicine-induced neurotoxicity of the primarily cultured cortical neurons. In this study, we investigated the effect of nicotine on the protection of neurons against colchicine damage and evaluated the associated intracellular signaling pathways. Nicotine-induced protection was blocked by an α7 nicotinic acetylcholine receptors (nAChRs) antagonist and a phosphatidylinositol 3-kinase (PI3K) inhibitor. These results suggest that the neuroprotective effects of nicotine are mediated by the α7 nAChRs and PI3K-Akt signaling pathway. In addition, we reveal that blockade of p38 and JNK (c-Jun N-terminal kinase) signaling increased Akt signaling, thus enhancing the survival of cell treatment with colchicine. On the other hand, inhibition of constitutively active Akt enhanced p38 or JNK signaling phosphorylation. These data suggested that crosstalk between PI3K Akt and p38 or JNK signaling pathways contributed to nicotine against colchicine-induced cytotoxicity.