When reward reinforcement in a two-choice discrimination task is regularly changed from one stimulus to another immediately after one learning acquisition session, the learning efficiency of a rat increases as if the rat has come to recognize this regularity of reversal. To investigate how the rat medial prefrontal cortex (mPFC) is involved in such improvement, we examined the performance of mPFC-lesioned rats in a serial reversal task of olfactory discrimination. The performance of other mPFC-lesioned rats in a delayed alternation task was also analyzed using the same apparatus to evaluate the contribution of the mPFC to working memory. The mPFC-lesioned rats demonstrated selective difficulty in the second reversal session in the serial reversal task and also showed performance impairment in the delayed alternation task. These results suggest that the rat mPFC mediating working memory is involved in early progress in learning efficiency during experiences of multiple reversals, which may be relevant to cognitive operations in reversal learning beyond a one-time reversal of stimulus response associations.