Effects of Development on Bone Mineral Density and Mechanical Properties in the Aquatic Frog, Xenopus Laevis, and a Terrestrial Frog, Lithobates Catesbianus

Abstract

The limb bones of vertebrates have a critical role in supporting the weight of the body and transmitting forces that power locomotion. The loads that limb bones experience can vary in association with a range of factors, including locomotor environment or developmental stage. Limbed vertebrates that are habitually found in environments with low locomotor loads (e.g., water) might be predicted to also exhibit limb bones with less elevated mechanical properties, such as yield stiffness and yield stress. Frogs provide a distinctive case, in which these ideas can be tested as they experience changes in both locomotor style andhabitat as they develop. However, whilemany frog taxa shift fromaquatic to terrestrial habitats as theymetamorphose, some lineages, such as pipids, maintain an aquatic lifestyle even after metamorphosis, providing a comparative framework for the effects of habitat shifts on developing limbs in vertebrates. This study compares the material composition and mechanical properties of the femur between frog species that are aquatic specialists (Xenopus laevis) vs generalists that spend considerable time both on land and in water (Lithobates catesbeianus) as they transition from metamorphic tadpoles to fully grown adults. MicroCT scanning was used to determine changes in bone density related to developmental stage and hindlimb use during swimming. Microindentation was then used to collect hardness values from the cortical bone of each femur, which was used to evaluate bone material properties.We found that aquatic frogs had less overall bone mineral density (BMD) than terrestrial frogs and that BMD wasmore elevated in the cortical region of the diaphysis than trabeculae and distal and proximal epiphyses. Despite its less elevated BMD, bone mechanical properties were not significantly different in aquatic specialist X. laevis than in more terrestrial L. catesbeianus. Our results suggest that the limb bones of aquatic frogs may experience compensatory effects through development to offset their lower BMD. Furthermore, changes in bone density and material properties across development may help to explain some of the differences in locomotor performance found between aquatic and terrestrial metamorphic frogs, providing insight into how environmental factors might correlate with bone ossification.