AbstractAn intriguing feature of thymocyte differentiation is that the competence to express both interleukin‐(IL)2 and CD25 is acquired even prior to T cell recept or (TcR) expression. When T cell receptor‐independent stimuli are used, immature cells can express IL‐2 at levels comparable to mature cells, but unlike the mature cells, immature cells require IL‐1 as a costimulus. Here we present evidence that IL‐1 affects a variety of responses by members of the CD25+subset of immature thymocytes. Cells in this population are IL‐1 dependent not only for induction of IL‐2 expression, but also for high‐level maintenance of CD25 expression. CD25+expression is amplified by IL‐1 through a mechanism highly sensitive to changes in Ca2+ionophore concentration. The effects of IL‐1 on CD25 maintenance are not mediated by IL‐2, because of the divergent effects of cAMP on IL‐2 and CD25+expression. IL‐1 costimulation also increases RNA accumulation in the cell cycle, and this effect too seems to be separable from the effects on IL‐2 and CD25+expression. All these effects of IL‐1 are developmentally stage‐specific, manifest in the CD25+subset of immature thymocytes but not in later‐stage thymocytes or splenic T cells. Multiparameter cell sorting experiments that dissect the transitional stages between immature and TcR+thymocytes imply that all immature cells pass through an IL‐1 responsive state. Responsiveness to IL‐1 costimulation is then lost by these cells, apparently irreversibly, at a stage just prior to detectable cell‐surface TcR expression. These results indicate that IL‐1 responsiveness is a defining characteristic of the activation physiology of cells in a p