Distribution of cortactin in cerebellar Purkinje cell spines.

Dendritic spines are the primary sites of excitatory transmission in the mammalian brain. Spines of cerebellar Purkinje Cells (PCs) are plastic, but they differ from forebrain spines in a number of important respects, and the mechanisms of spine plasticity differ between forebrain and cerebellum. Our previous studies indicate that in hippocampal spines cortactin-a protein that stabilizes actin branch points-resides in the spine core, avoiding the spine shell. To see whether the distribution of cortactin differs in PC spines, we examined its subcellular organization using quantitative preembedding immunoelectron microscopy. We found that cortactin was enriched in the spine shell, associated with the non-synaptic membrane, and was also situated within the postsynaptic density (PSD). This previously unrecognized distribution of cortactin within PC spines may underlie structural and functional differences in excitatory spine synapses between forebrain, and cerebellum.

Quantitative Electron Microscopic Assay Using Random Sampling from Single Sections to Test Plastic Synaptic Changes in Hippocampus.

Studies over several decades on the organization of the CA1 hippocampus-a particularly favorable model for learning, memory and certain forms of cognition-have shown that the synaptic network in this brain region is plastic ( Fortin et al., 2012 ). Recent evidence suggests that a number of environmental and endogenous stimuli may have a substantial effect on hippocampus-dependent cognitive function, implying enhanced synaptic plasticity in this brain region. Stimuli (e.g., food restriction, enriched environment, social interaction, gene-loss [knock-out animals], etc.) can trigger structural and functional plasticity (e.g., spine formation, increased expression of neurotrophic factors, synaptic function and neurogenesis) in the hippocampus ( Stewart et al., 1989 ; Andrade et al., 2002 ; Babits et al., 2016 ). Using quantitative electron microscopy, we can study the synaptic neuropil of CA1 hippocampus in rodents during short- or long-term treatments and/or stimuli. Within the scope of this electron microscopic methodological construct, the density of various synaptic connections, the morphology and internal structure of excitatory spine synapses (e.g., the mean length and width of postsynaptic densities) can be quantified. Such quantitative ultrastructural measurement using high-resolution electron-microscopy may be applied to observe structural manifestations of synaptic plasticity in rodent brain tissue. The presented ultrastructural protocol may empower researchers to reveal details and synaptic changes which may not be obvious using only light microscopy. Ultrastructural data may provide substantial advances in our understanding of the changes in hippocampal synaptic architecture under different conditions.