Abstract
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.