Cortical and medullary thymic epithelial cells provide essential signals for a normal programme of T-cell development. Current models of thymus development suggest that thymocyte-derived signals play an important role in establishing thymic microenvironments, a process termed thymus crosstalk. Studies on CD3epsilontg26 mice lacking intrathymic T-cell progenitors provided evidence that normal development of the thymic cortex depends upon thymocyte-derived signals. Importantly, the reported failure to effectively reconstitute adult CD3epsilontg26 mice raised the possibility that such crosstalk must occur within a developmental window, and that closure of this window during the postnatal period renders thymic epithelium refractory to crosstalk signals and unable to effectively impose T-cell selection. We have re-investigated the timing of provision of crosstalk in relation to development of functional thymic microenvironments. We show that transfer of either fetal precursors or adult T-committed precursors into adult CD3epsilontg26 mice initiates key parameters of successful thymic reconstitution including thymocyte development and emigration, restoration of cortical and medullary epithelial architecture, and establishment of thymic tolerance mechanisms including maturation of Foxp3(+) Treg and autoimmune regulator-expressing medullary epithelium. Collectively, our data argue against a temporal window of thymocyte crosstalk, and instead demonstrates continued receptiveness of thymic epithelium for the formation of functionally competent thymic microenvironments.