Date of Award
Doctor of Philosophy (PhD)
Neocortical circuits can adapt to changes in sensory input by modifying the strength or number of synapses. These changes have been well-characterized electrophysiologically in primary somatosensory (barrel) cortex of rodents across different ages and with different patterns of whisker stimulation. Previous work from our lab has identified layer-specific critical periods for synaptic potentiation after selective whisker experience (SWE), where all but one row of facial whiskers has been removed. Although whole-cell patch-clamp recording methods enable a mechanistic understanding of how synaptic plasticity can occur in vivo, they are painstakingly slow, typically focus on a small number of observed events, and are focused on a single pathway or restricted anatomical area. For example, most studies of plasticity in barrel cortex have focused on analyses of experience-dependent synaptic changes in layer 4 and layer 2/3, at a single time point, but it is unclear whether such changes are limited to these layers, or whether they persist over long time periods. Here we employ an established electron-microscopic technique that selectively intensifies synaptic contacts, in combination with unbiased, automated synapse detection, to broadly explore experience-dependent changes in synaptic size and density across many neocortical layers, regions, and time periods in a high-throughput fashion. To validate the method, we focused on imaging synaptic contacts at time points surrounding the critical period for strengthening of excitatory synapses in mouse barrel cortex, and compared these to electrophysiological analyses that show a doubling of synaptic events targeting layer 2/3 pyramidal neurons following SWE. We found that the pattern of occurrence of synapses across the cortical layers is significantly different following SWE. Also, an increase in length was observed specifically in layer 3 synapses. Furthermore, we uncovered potential bidirectional plasticity in L6 synapses depending on the developmental state of circuit and a potential critical period onset for L5A synapse at PND 18. The high resolution imaging and unbiased synapse detection has enabled us to potentially tease apart synaptic changes that occur in a laminar specific fashion. This high-throughput method will facilitate analysis of experience-dependent changes in synaptic density by age, sensory experience, genotype, pharmacological treatments or behavioral training, and will enable classification of synaptic structure to identify key parameters that can be changed by these variables.
Chandrasekaran, Santosh, "Unbiased, High-Throughput Electron Microscopy Analysis of Experience-Dependent Synaptic Changes" (2015). Dissertations. 462.