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Principal Investigator: Lori A. Setton
Organization: WASHINGTON UNIVERSITY
Fiscal Year: 2024
Award: $514,885
Funding agency: National Institute of Arthritis and Musculoskeletal and Skin Diseases
Intervertebral disc (IVD) degeneration is one of the greatest contributors to low back pain, yet how the IVD can
generate pain remains poorly understood. To date, our knowledge of “cross-talk” between degenerating IVD and
sensory nerves involved in transmitting pain is limited to findings of altered protein and RNA expression in tissues
of the IVD, the spinal cord or dorsal root ganglia (DRG). Recent advances in the imaging of sensory neuron
activation via recording of Ca2+ sensitive fluorescent indicators, together with pre-clinical models of IVD
degeneration, now enable the study of temporal and spatial changes in neuronal function and their “cross-talk”
to changes in the degenerating IVD.
We propose to evaluate action potential-driven Ca2+ transients and molecular changes in sensory
neurons in a mouse model of injury-induced IVD degeneration. In Specific Aim 1, we will document
temporal changes to pain-related behaviors and sensitivity in Thy1-GCaMP6s mice following puncture of a
lumbar IVD to induce IVD degeneration. These mice carry a transgene for the calcium-sensitive fluorophore,
GCaMP6s, that is expressed in sensory nerves of the DRG. We will also evaluate the presence of neuronal
markers and key ion channels in innervating DRGs, and anatomic changes and nerve fiber infiltration in IVDs,
to test for changes with IVD degeneration as compared to sham controls. This work will document molecular
changes to IVD and DRGs for this model from 6 to 52 weeks of IVD degeneration, and test for relationships
between injured IVD and the innervating lumbar DRGs as a first measure of “cross-talk.” In Specific Aim 2, we
will evaluate action potential-driven Ca2+ transients in lumbar DRG neurons in the IVD degeneration model of
Specific Aim 1. We will record Ca2+ transients in intact DRG of Thy1-GCaMP6s mice in vitro following electric
field stimulation, and measure threshold voltage, 50% maximum (IC50), and numbers of responding DRG
neurons and their nearest neighbor response. DRGs will also be tested before and after incubation with sodium
channel blockers to screen for remodeling of specific ion channel function with periods of IVD degeneration. Our
goal is to identify temporal and spatial changes in DRG function and “cross-talk” with changes of IVD
degeneration. Finally, in Specific Aim 3, we will evaluate sensory stimuli-induced responses in the DRG of
living mice using in vivo fluorescence microscopy. Working with 2-photon confocal microscopy and motion
correction algorithms developed for brain imaging, we will identify the threshold response of DRG neurons in
Thy1-GCaMP6s mice with and without IVD degeneration, following in vivo stimulation of brush, pinch, heat and
cold. Our goal is to test for relationships between in vivo activation of lumbar DRG neurons with behavioral and
sensitivity changes following onset of IVD degeneration. Completion of this study would identify functional
changes to sensory neurons at sites distant to the degenerated IVD and reveal new information about IVD-
nervous system “cross-talk” that may suggest novel interventions for treatment of discogenic pain.
Terms: <2-photon><Action Potentials><Acute><Affect><Afferent Neurons><Algorithms><Anatomic Sites><Anatomic structures><Anatomy><Animal Model><Animal Models and Related Studies><Animals><Behavior><Behavior monitoring><Behavioral><Body Tissues><Brain imaging><CNS Nervous System><Calcium><Cell Body><Cells><Central Nervous System><Chronic><Common Rat Strains><Confocal Microscopy><Dissociation><Distant><Dorsal Root Ganglia><Exposure to><Fluorescence Light Microscopy><Fluorescence Microscopy><Fugu Toxin><Gene Transcription><Generations><Genetic Transcription><Goals><Harvest><Image><Impairment><In Vitro><Incubated><Infiltration><Inflammation Mediators><Injury><Intervention><Intervention Strategies><Intervertebral Disc Degenerative Disease><Intervertebral Disc Degenerative Disorder><Intervertebral Disk><Intervertebral disc structure><Ion Channel><Ionic Channels><Knowledge><Low Back Ache><Low Back Pain><Low Backache><Lumbago><Measures><Medulla Spinalis><Membrane Channels><Methods><Mice><Mice Mammals><Modeling><Molecular><Molecular Target><Monitor><Morphology><Motion><Murine><Mus><Nerve><Nerve Cells><Nerve Fibers><Nerve Impulse Transmission><Nerve Transmission><Nerve Unit><Nervous System><Neural Cell><Neuraxis><Neurocyte><Neurologic Body System><Neurologic Organ System><Neuronal Transmission><Neurons><Operative Procedures><Operative Surgical Procedures><Organ><Organ Culture><Organ Culture Techniques><Pain><Painful><Pathology><Pattern><Phenotype><Population><Pre-Clinical Model><Preclinical Models><Puncture procedure><Punctures><RNA Expression><Rat><Rats Mammals><Rattus><Role><Sensory Neurons><Site><Sodium Channel Blockers><Spinal Cord><Spinal Ganglia><Surgical><Surgical Interventions><Surgical Procedure><TRPV1><TRPV1 gene><Tarichatoxin><Testing><Tetradotoxin><Tetrodotoxin><Tissues><Transcription><Transgenes><Transmission><Visualization><Work><X-ray microtomography><Xray microtomography><afferent nerve><arm><axon signaling><axon-glial signaling><axonal signaling><behavioral monitoring><brain visualization><cohort><contrast enhanced><degenerated intervertebral disc><degenerative disc disease><disability><disc degeneration><discogenic back pain><discogenic pain><dorsal root ganglion><electric field><fluorophore><glia signaling><glial signaling><imaging><in vitro Organ Culturing><in vitro vertebrate organ culturing><in vivo><in vivo fluorescence><inflammatory mediator><injured><injuries><interventional strategy><intervertebral disk degeneration><intervertebral disk injury><micro CT><micro computed tomography><microCT><microtomography><model of animal><mouse model><murine model><nerve signaling><neural signaling><neuronal><neuronal signaling><neurotransmission><neurotrophic factor><neurotrophin><neutrophin><new approaches><novel><novel approaches><novel strategies><novel strategy><perceptual stimulus><physicochemical phenomena related to the senses><promoter><promotor><protein expression><puffer fish toxin><response><second harmonic><sensory nerve><sensory stimulus><social role><spinal disk injury><surgery><transgene><transient receptor potential cation channel V1><transmission process><two-photon><voltage>