Three-dimensional distribution of cortical synapses: a replicated point pattern-based analysis.

Laura Anton-Sanchez,Josã©-Rodrigo Rodrã­Guez,Pedro Larraã±Aga,Javier Defelipe,Angel Merchã¡N-Pã©Rez,Concha Bielza

doi:10.3389/fnana.2014.00085

Abstract

The biggest problem when analyzing the brain is that its synaptic connections are extremely complex. Generally, the billions of neurons making up the brain exchange information through two types of highly specialized structures: chemical synapses (the vast majority) and so-called gap junctions (a substrate of one class of electrical synapse). Here we are interested in exploring the three-dimensional spatial distribution of chemical synapses in the cerebral cortex. Recent research has showed that the three-dimensional spatial distribution of synapses in layer III of the neocortex can be modeled by a random sequential adsorption (RSA) point process, i.e., synapses are distributed in space almost randomly, with the only constraint that they cannot overlap. In this study we hypothesize that RSA processes can also explain the distribution of synapses in all cortical layers. We also investigate whether there are differences in both the synaptic density and spatial distribution of synapses between layers. Using combined focused ion beam milling and scanning electron microscopy (FIB/SEM), we obtained three-dimensional samples from the six layers of the rat somatosensory cortex and identified and reconstructed the synaptic junctions. A total volume of tissue of approximately 4500μm3 and around 4000 synapses from three different animals were analyzed. Different samples, layers and/or animals were aggregated and compared using RSA replicated spatial point processes. The results showed no significant differences in the synaptic distribution across the different rats used in the study. We found that RSA processes described the spatial distribution of synapses in all samples of each layer. We also found that the synaptic distribution in layers II to VI conforms to a common underlying RSA process with different densities per layer. Interestingly, the results showed that synapses in layer I had a slightly different spatial distribution from the other layers.

Highlights

A very dense network of neuronal and glial processes occupies the space between the cell bodies of the neurons, glia, and blood vessels
Three-dimensional brain tissue samples were obtained from flat-embedded vibratome sections using a combined focused ion beam/scanning electron microscope (FIBSEM)
Virtually all cortical synapses can be accurately identified as asymmetric and symmetric using FIB/scanning electron microscopy (SEM) (Merchán-Pérez et al, 2009), we considered synaptic junctions as a whole

Summary

Introduction

A very dense network of neuronal and glial processes occupies the space between the cell bodies of the neurons, glia, and blood vessels. Given that most synapses are found here and the neuropil accounts for the largest volume of the cerebral cortex, it follows that most synaptic interactions take place in the neuropil (Alonso-Nanclares et al, 2008). Numerous researchers have been trying to find simple and accurate methods for estimating the distribution, size and number of synapses. To this end, two sampling procedures are currently available: one is based on serial reconstructions and the other on single sections. Serial sectioning transmission electron microscopy is a well-established and mature technology for collecting three-dimensional data

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Conclusion