Abstract 116: Single-synapse Analysis Of Synaptic Remodeling In The Post-stroke Rodent Brain

Introduction: The mechanisms of functional recovery after stroke are thought to be based on structural and functional changes in brain circuits adjacent to or connected with the stroke site. Deciphering these changes at the synaptic level is key to understanding the re-organization of the synaptic circuitry (i.e. the connectome). Quantitative information about such synapse rearrangements after stroke has been inadequate however, due to the technical limitations of available methodologies. Here we describe the use of array tomography, a new high-resolution proteomic imaging method, to determine the composition of glutamate and GABA synapses in the post-stroke mouse brain. Methods: A cortical lesion was induced in 12-week-old C57BL/6J male mice using the distal middle cerebral artery occlusion model of ischemia. Small tissue sections were removed from the peri-infarct cortex and ribbons of serial ultrathin (70 nm) sections were obtained using an ultramicrotome. Ribbons were stained with antibodies for the synaptic markers SynapsinI, VGlut1, VGlut2, PSD-95, GAD, VGAT. Analysis of the resultant staining pattern was used to identify subtypes of glutamatergic and GABAergic synapses. Results: At 1 week post-stroke, an increase in GABAergic synapses was observed in layer 5 of the peri-infarct cortex. A sub-analysis of the type of inhibitory interneurons (e.g. parvalbumin, somatostatin) expressing these synapses is pending. In addition, a trend for an increase of VGlut1+2 synapses was also observed. However, there were no detectable differences in total synapse number between stroke-injured and naive animals, thus suggesting that VGluT2 expression may be upregulated in existing glutamatergic VGluT1 synapses after stroke. Further analysis will be extended to cortical layers 2/3 and 4. Conclusion: These results provide new information about the organization of synaptic circuitry and its plasticity after stroke. Furthermore, it demonstrates how array tomography enables a previously unobtainable level of volumetric visualization and quantification of synapses.