Urocortin (UCN) is a new neuropeptide of the corticotrophin-releasing hormone (CRH) family which plays an important role in immune responses. Mycobacterium avium subspecies paratuberculosis (Map) is the etiological agent of paratuberculosis (Johne's disease). The role of UCN or CRH in the pathogenesis of Map-infection is unknown. In the present study, we first cloned the bovine UCN gene and demonstrated the profile of UCN or CRH expression in peripheral blood cells from Map-infected cattle and uninfected controls by real-time reverse transcription-polymerase chain reaction (RT-PCR) and ELISA analysis. These data are the first observations of the characteristic kinetics of these neuropeptides in Map-infection. UCN or CRH expression in non-stimulated blood samples from infected cattle was higher than that in similarly treated samples from uninfected controls; however, exposure to Map lysate and live Map resulted in down-regulated expression of UCN in infected cattle compared to their counterparts from uninfected controls. These results have provided a direction in understanding the pathogenesis of paratuberculosis and improving diagnostic methods for Map-infection.