T cells are present in large numbers in the epithelial lining of the small and large intestine of humans and mice. Those cells, referred to as intraepithelial lymphocytes (IELs), are critical for maintaining an effective mucosal immune response against the onslaught of enteric infectious agents and intestinal neoplasia. However, because intestinal immunity must by necessity occur rapidly and efficiently, it is concomitantly important that the local intestinal immune response be curtailed so as not to result in conditions that lead to a destructive inflammatory environment as occurs in inflammatory bowel disease (IBD). Although many aspects of the IEL activation process remain to be understood, emerging evidence indicates that costimulatory molecules on IELs are critical for activation and that they hold the key to regulating intestinal immunity across many levels. In this article, the involvement of three IEL costimulatory molecules (CD43, OX40, and Ly-6C) - working independently or in collaboration-will be discussed in the context of immunity and disease in the human and mouse intestine, and the involvement of those in sustaining the IELs in a uniquely precarious but effective state of activation readiness will be explored.