Behavioral discrimination procedures clearly demonstrate that nicotine elicits interoceptive stimulus effects in humans that are malleable by various pharmacological manipulations as well as by some behavioral manipulations. The parameters of nicotine discrimination and both chronic and acute factors that may alter discrimination behavior are addressed in this chapter, which emphasizes research by the author involving nicotine delivered by nasal spray. Human discrimination of nicotine is centrally mediated, as the central and peripheral nicotine antagonist mecamylamine blocks discrimination but the peripheral antagonist trimethaphan does not. The threshold dose for discrimination of nicotine via spray appears to be very low in smokers as well as nonsmokers. Because smoked tobacco delivers nicotine more rapidly than spray, the threshold dose of nicotine via smoking is probably even lower. In terms of individual differences, smokers may become tolerant to the discriminative stimulus effects of higher nicotine doses but not of low doses. Men may be more sensitive than women to nicotine's discriminative stimulus effects, consistent with other research suggesting that nicotine is more reinforcing in men than in women. Other potential individual differences in nicotine discrimination have not been clearly tested, but may include genetics, obesity, and dependence on other drugs. Acute environmental factors that alter nicotine discrimination include the specific training and testing conditions, pointing to the need for careful control over such conditions during research. Other factors, such as concurrent acute use of alcohol or caffeine, do not appear to alter nicotine discrimination, suggesting that changes in nicotine discrimination are not likely explanations for the association of smoking behavior with use of those drugs. Concurrent physical activity also does not appear to alter nicotine discrimination, indicating that results from studies of discrimination in subjects at quiet rest, the standard approach in this research, generalize well to discrimination in subjects engaged in various activities, as often occurs in the natural environment. Future research should more clearly examine the potential role of nicotine's discriminative stimulus effects in nicotine reinforcement and determine the generalizability of these findings to nicotine delivered by other means, particularly tobacco smoking.