Bone?s ability to respond and adapt to mechanical loading declines significantly with age, which is a major contributor to bone fragility. We do not understand how alterations in osteocyte mechanical sensing occur in aging bone. This is a critical knowledge gap given the central role for osteocytes in orchestrating bone formation and resorption responses. Recent breakthrough discoveries from my mentor?s laboratory revealed how osteocytes in living bone perceive and encode mechanical loading information. They created the first ever reporter mice with an osteocyte-targeted genetically encoded calcium indicator to allow measurement of osteocyte Ca2+ responses to loading in vivo. They discovered that osteocyte populations in living bone numerically encode load magnitude, with increasing strain levels recruiting more Ca2+ responding osteocytes in healthy young adult bone following a robust, linear response curve. Preliminary studies further revealed that this osteocyte loading response curve changes markedly with systemic challenge. The proposed F31 training grant will determine how osteocyte Ca2+ responses to mechanical loading in vivo change with age.
In Aim 1, we will create tamoxifen inducible GCaMP6f mice in which the Ca2+ reporter can be turned on at selective times throughout adulthood. This will address challenges inherent in long-term constitutive GCaMP expression as was used in the first-generation osteocyte Ca2+ reporter mice previously created by our laboratory. Metatarsal bones will be loaded in vivo through the range of physiological strain levels using a combined loading and multiphoton imaging approach and osteocyte Ca2+ responses in cortical bone will be measured.
In Aim 2, we will use these inducible GCaMP6f mice to assess osteocyte Ca2+ responses to loading in young adult, middle aged, and aged adult mice. Finally, a range of channels (eg, T-type and L-type channels, Panx1-P2X7, Piezo 1, TRPV channels) are implicated in osteocyte Ca2+ response to loading in vitro. However, the in vivo importance of these channels is poorly understood. In the exploratory studies in Aim 3, we will use validated selective channel antagonists to test how modulating channel function in vivo influences load- response curves. Relevance to F31 grant Scientific and Training goals: Research: These studies will be the first to examine osteocyte mechano-sensing in vivo throughout adult life. Training: The proposed fellowship incorporates a broad range of experimental and analytical approaches (mouse models, bone biomechanics, in vivo studies, multiphoton imaging, cell Ca2+ measurements, bone and osteocyte physiology, channel biology) that will provide a robust platform for research career growth.
Consensus over the last decade is that osteocytes, the cells that reside within the solid bone material, are the critical cells for sensing bone loading. This sensing and the resulting biochemical signals produced by osteocytes are key to growing the ?right? skeleton and maintaining it throughout life, yet we do not understand how alterations in osteocyte mechanical sensing occur in aging bone, and what that means for bone health and osteoporosis prevention. In the current studies, we will understand how osteocyte sensing of mechanical load differ in the young adult, middle aged and old skeleton; understanding these mechanisms may open a new window for novel therapeutic targets that modulate bone remodeling and prevent osteoporosis and bone fragility.
