Abstract
High-frequency electrical stimulation is becoming a promising therapy for neurological disorders, however the response of the central nervous system to stimulation remains poorly understood. The current work investigates the response of myelin to electrical stimulation by laser-scanning coherent anti-Stokes Raman scattering (CARS) imaging of myelin in live spinal tissues in real time. Paranodal myelin retraction at the nodes of Ranvier was observed during 200 Hz electrical stimulation. Retraction was seen to begin minutes after the onset of stimulation and continue for up to 10 min after stimulation was ceased, but was found to reverse after a 2 h recovery period. The myelin retraction resulted in exposure of Kv 1.2 potassium channels visualized by immunofluorescence. Accordingly, treating the stimulated tissue with a potassium channel blocker, 4-aminopyridine, led to the appearance of a shoulder peak in the compound action potential curve. Label-free CARS imaging of myelin coupled with multiphoton fluorescence imaging of immuno-labeled proteins at the nodes of Ranvier revealed that high-frequency stimulation induced paranodal myelin retraction via pathologic calcium influx into axons, calpain activation, and cytoskeleton degradation through spectrin break-down.
Highlights
High-frequency electrical stimulation of the central nervous system (CNS) is becoming a promising therapy for treatment of a variety of disorders
Real-time E-coherent anti-Stokes Raman scattering (CARS) imaging showed that retraction began shortly after the onset of stimulation and continued throughout the duration of stimulation (Fig. 2F and Supplemental Movie S1)
Utilizing compound action potentials (CAPs) measurement to observe the collective response of many axons to stimulation as well as CARS microscopy to efficiently examine many individual nodes in live spinal tissues, we have explored mechanisms linking repetitive stimulation to paranodal myelin change
Summary
High-frequency electrical stimulation of the central nervous system (CNS) is becoming a promising therapy for treatment of a variety of disorders. The response of the CNS to extended high-frequency stimulation is not well understood [5,6]. It was further suggested that high-frequency stimulation could loosen the paranodal axoglial junctions thereby impacting the action potential [8]. Structural deformation of myelin following repetitive CAP propagation has been visualized by histology [9], X-ray diffraction [10], and electron microscopy [11,12]. Real-time visualization of structural changes of paranodal myelin upon repetitive stimulation was impossible with these traditional techniques. The mechanism linking electrical stimulation and paranodal myelin deformation has not been clarified
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