Females are significantly more likely to become addicted to nicotine as demonstrated by studies reporting that women have greater difficulty quitting smoking compared to men. Intensive research focuses on nicotinic acetylcholine receptors as the agents responsible for nicotine addiction. However, the mechanism by which females are more susceptible to nicotine dependency is not fully understood. In order to address these sex differences in response to nicotine, estrogen and progesterone levels of 6 female Sprague Dawley rodents, Rattus norvegicus, were monitored at the following time intervals: pre-nicotine injection, 30 minutes post-6 days of daily nicotine injection, and one hour post-6 days of daily nicotine injection. Exfoliate cytology of vaginal mucosa was concurrently sampled at these three time periods, stained, and analyzed via microscopy. The number of nucleated epithelial cells, anucleated cornified cells, and leukocytes was estimated to determine estrous cycle phase: proestrus, estrus, metestrus, or diestrus. Animals injected with saline were used as a control. Injections were administered intraperitoneally and separated by at least 24 hours to eliminate residual drug effects. Before nicotine injection, animals were in various estrous cycle stages with no significant difference in the number of animals per stage. Thirty minutes post-6 days of nicotine administration, a significantly greater number of non-prescient mammals transitioned into estrus from their original stage rather than transitioning into the subsequent cycle stage (p < 0.001). One hour post-6 days of nicotine administration, these values remained significant as more animals transitioned into estrus at this time interval (p < 0.001). Correspondingly, an overwhelming majority of nucleated epithelial cells became anucleated cornified cells at both 30 minutes and one hour post-6 days of daily nicotine injection. Animals injected with saline remained in the same stage throughout administration. Our results demonstrate that nicotine incites a transformation to estrus in the murine model, regardless of cycle phase before nicotine administration. During the estrus stage, both estrogen and progesterone were low as confirmed by an increase in nucleated cells on vaginal smear. Due to the fact that nicotine administration was associated with a decrease in ovarian hormones, a lack of estrogen and progesterone could be paramount to elucidating the sexual dimorphism of addiction.