Host: The Japanese Pharmacological Society
Name : The 97th Annual Meeting of the Japanese Pharmacological Society
Number : 97
Location : [in Japanese]
Date : December 14, 2023 - December 16, 2023
Osteocytes play a crucial role in regulating bone metabolism through their interaction with osteoclasts and osteoblasts. While osteocytes are believed to act as bone mechanosensors, this hypothesis has only been confirmed in vitro and remains unexplored at the in vivo level. In this study, we developed a novel method to visualize osteocyte responses to mechanical loading in live mice using a two-photon excitation microscope.
To facilitate our research, we developed specialized fixtures using an optical 3D printer and securely anchored bone tissue using biocompatible cement. We selected a strain of mice expressing GO-ATeam2, a fluorescence resonance energy transfer (FRET)-based biosensor for ATP, either in the cytosol or mitochondria, as ATP dynamics are recognized as indicative of osteocyte mechanosensing. We obtained fluorescent images from distinct organelles within the osteocytes. Subsequently, we applied static loads along the tibia's long axis using an actuator and monitored the applied force with a load cell. Fluorescent images were captured under various loading conditions. Our investigation into cytosolic ATP dynamics in osteocytes under bone loading, as analyzed through cytosol-localized GO-ATeam2, revealed no significant differences in ratio values at 1N and 3N compared to 0N. However, a noteworthy decrease was observed at 5N and 7N. Similarly, our examination of intramitochondrial ATP dynamics using mitochondria-localized GO-ATeam2 displayed no significant deviations in ratio values across loading conditions ranging from 1N to 7N when compared to 0N.
Our findings suggest that osteocytes do not respond to low loads below 3N. In contrast, excessive loading above 5N results in alterations in the cellular state of osteocytes. Furthermore, our study implies that osteocytes subjected to 5N and 7N mechanical loading exhibit distinct ATP dynamics within osteocyte organelles as a response to mechanical stimuli at the in vivo level.
