摘要The formation of axonal spheroid is a common feature following spinal cord injury.To further understand the source of Ca2+that mediates axonal spheroid formation,we used our previously characterized ex vivo mouse spinal cord model that allows precise perturbation of extracellular Ca2+.We performed two-photon excitation imaging of spinal cords isolated from Thy1YFP+transgenic mice and applied the lipophilic dye,Nile red,to record dynamic changes in dorsal column axons and their myelin sheaths respectively.We selectively released Ca2+from internal stores using the Ca2+ionophore ionomycin in the presence or absence of external Ca2+.We reported that ionomycin dose-dependently induces pathological changes in myelin and pronounced axonal spheroid formation in the presence of normal 2 mM Ca2+artificial cerebrospinal fluid.In contrast,removal of external Ca2+significantly decreased ionomycin-induced myelin and axonal spheroid formation at 2 hours but not at 1 hour after treatment.Using mice that express a neuron-specific Ca2+indicator in spinal cord axons,we confirmed that ionomycin induced significant increases in intra-axonal Ca2+,but not in the absence of external Ca2+.Periaxonal swelling and the resultant disruption in the axo-myelinic interface often precedes and is negatively correlated with axonal spheroid formation.Pretreatment with YM58483(500 nM),a well-established blocker of store-operated Ca2+entry,significantly decreased myelin injury and axonal spheroid formation.Collectively,these data reveal that ionomycin-induced depletion of internal Ca2+stores and subsequent external Ca2+entry through store-operated Ca2+entry contributes to pathological changes in myelin and axonal spheroid formation,providing new targets to protect central myelinated fibers.