Menthol Decreases Oral Nicotine Aversion in C57BL/6 Mice Through a TRPM8-Dependent Mechanism Lu Fan, M.D., Ph.D 1 , Shrilatha Balakrishna, Ph.D. 1 , Sairam V. Jabba, D.V.M., Ph.D. 2 , Pamela E. Bonner, M.S. 2 , Seth R. Taylor, Ph.D. 1 , Marina R. Picciotto, Ph.D. 1 , and Sven-Eric Jordt, Ph.D. 1,2 1 Yale Tobacco Center of Regulatory Science (TCORS), Department of Psychiatry, Yale School of Medicine, New Haven, CT, USA 2 Department of Anesthesiology, Duke University School of Medicine, Durham, NC Abstract Background—Nicotine is a major oral irritant in smokeless tobacco products and has an aversive taste. Mentholated smokeless tobacco products are highly popular, suggesting that menthol increases their palatability and may facilitate initiation of product use. While menthol is known to reduce respiratory irritation by tobacco smoke irritants it is not known whether this activity extends to oral nicotine and its aversive effects. Study design—The two-bottle choice drinking assay was used to characterize aversion and preference in C57BL/6 mice to a range of menthol concentrations (10–200 μg/ml). Then, effects of menthol on oral nicotine aversion were determined. Responses were compared with those in mice deficient in the cold/menthol receptor, TRPM8, expressed in trigeminal sensory neurons innervating the oral cavity. Results—Mice showed aversion to menthol concentrations of 100 μg/ml and above. When presented with a highly aversive concentration of nicotine (200 μg/ml), mice preferred solutions with 50% or 100 μg/ml menthol added over nicotine alone. In contrast to wild-type mice, Trpm8−/ − showed a strong aversion to mentholated (100 μg/ml) nicotine (200 μg/ml) and preferred nicotine alone. Trpm8−/− mice show aversion to lower concentrations of menthol than wild-type mice. Conclusions—Oral menthol can reduce the aversive effects of oral nicotine and, at higher concentrations, acts as an irritant by itself. Menthol’s effects in relation to nicotine require TRPM8, the cool temperature sensing ion channel that activates analgesic and counterirritant mechanisms. These mechanisms may underlie preference for menthol-containing smokeless tobacco products and may facilitate initiation of product use. Correspondence to Dr. Sven-Eric Jordt, Department of Anesthesiology, Duke University School of Medicine, Box 3094 MS27, Durham, NC 27710-3094, USA; sven.jordt@duke.edu. Contributors SEJ and MRP: conceived of the study. LF, SB, SVJ, PEB and SRT performed experiments. LF, SB, SVJ, MRP and SEJ contributed to data analysis, interpretation of data, writing of the manuscript, and review, revision, and approval of the final article. Competing Interest The authors declare no competing interests. HHS Public Access Author manuscript Tob Control. Author manuscript; available in PMC 2017 November 01. Published in final edited form as: Tob Control. 2016 November ; 25(Suppl 2): ii50–ii54. doi:10.1136/tobaccocontrol-2016-053209. Author Manuscript Author Manuscript Author Manuscript Author Manuscript