The Impact Of Cerebral Ischemic Stroke On Synaptic Structures In Mice

Cortical microcirculation is more than sufficient to support the integrity and activity of the brain network under normal physiological conditions.Previous studies have showed that stroke can cause drastic decrease in cerebral blood flow and disorders in the microcirculation,thus resulting in destruction of neuronal structures.Fortunately,the destroyed neuronal structures can be restored if reperfusion occurred in time.However,the mechanism and influential factors for reversible recovery of neuronal structures are still unknown.In the present study,we used a reversible global cerebral ischemia model in mice combined with transcranial two-photon imaging and electron microscopy to study the structural basis for the restoration of synaptic structures after ischemia and the effects of different durations of ischemia on damage and restoration of neuronal structures.Two-photon live imaging results showed that global cerebral ischemia caused rapid disruption of neuronal structures,and 13.2% of spines disappeared during 20 min of transient ischemia.The damaged neuronal structures can be rapidly restored following transient ischemia/reperfusion.Remarkably,the restoration of dendritic structures following transient ischemia was associated with a rapid and sustained reorganization of spines,which is mainly reflected in the reappearance of lost spines in situ and appearance of newly formed spines.Electron microscopy results showed that organelle,cytoplasm components and synaptic ultrastructures also experienced the process of disruption and restoration after transient ischemia/reperfusion.Long-term in vivo imaging results found that the restored synaptic structures survived the initial ischemia-reperfusion challenge and were stable over 30 days,and spines on the survived dendrites underwent a sustained remodeling following transient ischemia.Behavioral test results showed that the effects of transient ischemia on behavior were only limited to the first few days after reperfusion(0-3 days).The rapid recovery of animal behaviors may be related to the rapid restoration of synaptic structures,and the rapid remodeling of synaptic connections may be a compensation for damaged synaptic circuits.In addition,the recovery of dendritic and spine structures was depended on the duration of ischemia,and it was more and more difficult for them to restore with the extension of ischemic time.The percentages of restored dendrites at 1 h after reperfusion for 20 min,1 h,3 h and 6 h of ischemia were 93%,56%,17% and 0%,respectively.The recovery of dendritic structures was declined dramatically when the ischemic time was more than 3 h(there was 36% recovery of dendritic structures after 3 h of ischemia followed by 6 h of reperfusion),and it was hard to restore for damaged dendritic structures when the ischemic time was extended to more than 6 h(only 2% recovery of dendritic structures after 6 h of ischemia followed by 6 h of reperfusion).Live imaging showed that the time window for revisable recovery of damaged dendritic structures was 3-6 h after ischemia,which was consistent with the clinical time window for the treatment of ischemic stroke.Histolodical analysis using YFP brain slice and Golgi staining showed that dendritic beading mainly located within the dendritic tuft when ischemia was less than 3 h,but can spread to the entire dendritic shaft after 6 h of ischemia.Furthermore,3-6 h of ischemia caused an increasing number of degenerating neurons in cortex and caudate putamen(striatum).Importantly,TEM data showed that nuclear chromatin clumping occurred in neuron after transient global ischemia and then recovered after reperfusion;however significant chromatin margination and karyopyknosis of neuron were observed after 6 h of ischemia.Consistently,behavioral performance of post-ischemic animals experienced an ischemia time-dependent recovery.Taken together,our results suggested that disrupted synapses after transient ischemia could be rapidly restored following reperfusion,and the recovery of neuronal structures following reperfusion was dependent on the degree of neuronal damage.These results can provide valuable experimental data for clinical treatment of ischemic stroke.