During frog ontogeny, PHA and Con A responsiveness of splenocytes precedes that of thymocytes.


The in-vitro proliferation of splenocytes and thymocytes from Xenopus laevis-gilli (hybrid clone LG-15) to the T cell mitogens, concanavalin A (Con A) and phytohaemagglutinin-P (PHA), were examined at specific stages of larval development (stages 51-66 of Nieuwkoop & Faber, 1967) and at 2 months post-metamorphosis. The responses of splenic lymphocytes to each mitogen were significant at all stages with stimulation indices ranging from 1.9 to 50.5 and 2.6 to 45.5 for PHA and Con A, respectively. Stage-related differences in responses of splenocytes to both mitogens suggest two waves of emergence of proliferative activity during development, divided by periods of diminished responsiveness during the metamorphic crisis. In contrast to the responses observed with splenocytes, proliferation of thymocytes cultured with either mitogen was barely detectable, with stimulation indices ranging from 1.2 to 6.9 and 1.4 to 2.9 for PHA and Con A, respectively. These minimal responses were observed only when thymocytes were cultured at relatively high cell density (5 X 10(5) cells/ml); they were not improved by increased or decreased concentrations of mitogen or by increased concentrations of fetal calf serum (5 or 10%) in the medium. Co-culture of larval thymocytes with autologous splenocytes and each mitogen did not consistently increase thymocyte responses suggesting that the defect in thymocyte responsiveness is not due to lack of accessory cells. These findings suggest that if PHA- and Con A-reactive cells are present in the thymus, they are present in relatively low numbers at all stages of larval development. The pattern of early mitogen responsiveness in the spleen at a time when the thymus is unresponsive contrasts with that observed in mammalian development in which thymocytes become responsive to mitogens in fetal stages and mitogen responsiveness appears in the spleen only around the time of birth. The apparent inactivity of larval thymocytes may reflect a population of cells that can become tolerant to those neo-self-antigens that arise during and after metamorphosis. If so, the larval amphibian thymus may provide a model to study the early events of thymocyte 'education' and differentiation in a broader time framework than is possible with fetal mammals.