1. Variation in Metamorphic Timing can be Induced by Variation in Density without Differences in Growth Rate in the Frog, Rana utricularia Janel Richter 1, Lincoln Martin 2, and Christopher K. Beachy 1 1 Department of Biology, Minot State University, Minot, ND, 2 Sawyer School, Sawyer, ND DENSITY EFFECTS EFFECTS OF FOOD REDUCTION Wilbur-Collins Predicts: Our Results: Pure Density Food reduction yesno yesno n=1n=4n=8n=10n=20n=2 n=1n=4n=8n=10n=20n=2 Constant food supply Reduction in food supply SCHEMATIC REPRESENTING EXPERIMENTAL DESIGN: We created all possible combinations of six density levels and two feeding regimes. Thus, twelve different treatments were imposed. Tadpoles were randomly assigned to a treatment, and treatments were placed in spatial blocks in order to compensate for spatially induced variation in growth and metamorphosis. Each treatment was replicated six times, for a total of 72 boxes arranged in 12 spatial blocks. Each spatial block contained the full complement of density treatments, thus 45 tadpoles. We used a total 12 X 45 = 540 tadpoles. Survival to metamorphosis was approximately 40% in density treatment of > 4 tadpoles per box. Mortality was higher in the low (I.e., 1 and 2 tadpoles per box) treatments. For other reasons the 1 and 2 density treatments were not used in analyses. Literature cited: Beachy, C.K. 2003. Copeia; Hensley, F.A., 1994. Ecology; Scott, D. 1990. Ecology; Wilbur, H.M., Collins, J.P. 1973. Science. HUSBANDRY, PROTOCOL AND DESIGN RESULTS A food reduction is expected to reduce growth, and cause metamorphosis PREDICTED EFFECTS OF FOOD REDUCTION As long as growth rates are maintained to be equal, variation in density is not expected to cause metamorphosis PREDICTED DENSITY EFFECTS INTRODUCTION AND HYPOTHESES The larval (or tadpole) stage of the amphibian life cycle is generally considered as an adaptation to take advantage of a transient, yet high growth, larval environment, e.g., a temporary pool. Wilbur and Collins (1973) posited an optimality model that uses the hypothesis that all environmental variables can be transduced via a larval amphibian’s growth rate to initiate an Abundant evidence exists variable growth among larvae results in variation in metamorphic timing (e.g., Hensley, 1994; Beachy, 2003); however, exactly how variable growth maps to metamorphic timing remains unclear. Furthermore, while there exists a large literature on how natural variation in larval density results in variation in metamorphic timing, the usual explanation for this variation appeals to presumptive variable rates of growth (e.g., Scott, 1990). These studies never contain individual growth schedules, and the assumption tends that density results in variable growth and that this cues mediates metamorphic timing. Additionally, there is a growing body of experimental endocrinological evidence that supports the notion that stress hormone systems (e.g., corticotropins) may act to mediate metamorphosis. This means that the hypothesis that larval amphibians may be “using” density as a stressor to initiate metamorphosis, regardless of growth rate. We used tadpoles of the southern Leopard Frog, Rana utricularia, to test the hypotheses that (a) growth influecnes metamorphic timing and (b) that density alone can affect metamorphic timing. R. Utricularia is found throughout the southeastern United States, and most commonly courts and lays eggs in somewhat ephemeral ponds in late Autumn, and tadpoles usually complete metamorphosis 7-8 months later the following Spring. endocrine system response, i.e., initiate metamorphosis and depart from a poor growth habitat. Food application was via a shaker that was calibrated to provide 25 mg aliquots of a finely- ground 1:1 mix of TetraMin fish flakes and Purina Rabbit Chow. Tadpoles were contained in standard size shoeboxes. Shoeboxes were placed in spatial block of the 12 treatments. Block #4, 4 tadpole treatment Metamorphosis was defined as Gosner (1960) Stage 42, emergence of forelimb. Tadples were weighed every 7 days and upon metamorphosis. Growth profiles of all six density treatments. Because the growth of 1 and 2-density treatments was slower, these treatments were excluded from subsequent analysis and discussion. Complete growth profiles of 4-, 8-, 10-, and 20- density treatments. Profile terminate at the mean mass and date at metamorphosis. A food reduction is expected to reduce growth, and cause metamorphosis -- REJECTED NS P<.01 yes no food reduction? tadpole density NS P<.05 a a,b b As long as growth rates are maintained to be equal, variation in density is not expected to cause metamorphosis -- REJECTED 204810 204810 SUMMARY ABOUT METAMORPHIC TIMING