Abstract Neck and back pain have a tremendous annual incidence and associated costs. The facet capsular ligament (FCL) encloses the bilateral articulating joints of the spinal vertebrae and is richly innervated to provide proprioception during spinal motions. The FCL is also innervated by nociceptors and may act as a pain sensor during abnormal conditions like injury and repeated loading. Although aberrant spinal motions and pathologic conditions are associated with pain, the relationship between tissue loading and nociceptor activation is unclear due to the complicated involvement of mechanics and physiology in the FCL across length scales. Accurately relating spinal motions to neuronal function within the FCL requires multi-scale modeling and experiments to identify the relevant mechanotransduction mechanisms by which tissue loading mediates neuronal function. Under this U01 renewal, we will expand our prior work defining how the neuronal response is governed by the forces from its local environment, which are determined by the complex interaction of macroscopic loads and the microscopic structure of the FCL. We extend that work by improving our multiscale models of FCL mechanics at the tissue, collagen fiber network, and neuronal scales. We will use those models to study the mechanical environment of neurons in the tissue and its collagen matrix and to predict responses under injury and pathologic conditions. Complementary experiments at the tissue and cell scales will characterize neuron and matrix structure and architectural descriptions of neurons, as well as define rate effects and the mechanical interactions between the FCL, collagen fibers, and neurons. We will integrate modeling and experimental work under coordinated specific aims to define how the organization of fibrillar and non-fibrillar material in the FCL govern its mechanical response to different loading scenarios, how the micro-scale fiber motion translates into forces on neurons, and how those forces affect neuronal architecture, signaling and function. In Aim 1, we will use advanced bioimaging, image-processing, and analytical tools to refine our existing multiscale model and capture the complex geometry and architecture of the matrix and neurons in the FCL. In Aim 2, we will define forces on the neurons during loading when its surrounding matrix deforms; Aim 3 includes adding viscoelasticity and interstitial flow to our model and studying rate effects on both the tissue, matrix and neurons. Finally, in Aim 4, we will insert our cellular/matrix model (µm-to-nm scale) into a whole-spine model (mm-µm scale) to link realistic macroscopic loading to neuronal deformation in clinically relevant contexts. By connecting the tissue and cellular scales, the project will facilitate efforts to include relevant physiological data on joint mechanics during injury and clinically relevant-spinal conditions with afferent neuronal function, which will promote understanding of the in vivo res...