Ultrastructure of astrocytes using volume electron microscopy: A scoping review

Abstract

The morphological features of astrocytes are crucial for brain homeostasis, synaptic activity and structural support, yet remain poorly quantified. As a result of the nanometre-sized cross-section of neuropil astrocytic processes, electron microscopy (EM) is the only technique availabe to date capable of revealing their finest morphologies. Volume EM (vEM) techniques, such as serial block-face or focused ion beam scanning EM, enable high-resolution imaging of large fields and allow more extensive 3-D model analyses, revealing new astrocytic morphological features. This scoping review aims to summarize the state of the art of astrocyte ultrastructural analysis. This review included 45 of 439 non-duplicated articles from a Pubmed search, categorizing studies by research focus, animal models, brain region, vEM techniques and segmentation methods. By answering classical questions such as volume, surface area, branching complexity and synaptic ensheathment reported in the literature, this work is a valuable resource for scientists working on structural biology or computational neuroscience.

Keywords: FIB‐SEM; SBF‐SEM; astrocyte; leaflets; perisynaptic processes; ssTEM; ultrastucture; volume electron microscopy.

Figure 1. PRISMA‐ScR

Preferred Reporting Items for Systematic Reviews and Meta‐Analyses extension for Scoping Reviews chart for the identification of papers included in the scoping review.

Figure 2. Maximum volume of imaging per years

Maximum volume of imaging per years in the analysed papers, highlighting the vEM technique employed. ATUM‐SEM, automated tape‐collecting ultra Microtome‐scanning electron microscopy; FIB‐SEM, focused ion beam‐scanning electron microscopy; SBF‐SEM, serial block face‐scanning electron microscopy; ssTEM, serial section transmission electron microscopy; vEM, volume electron microscopy.

Figure 3. Frequency of use for each software

Frequency of use for each software extracted from the papers that reported this information and categorized by segmentation, visualization, analysis, integrated software and custom solutions.

Figure 4. Models and CNS regions of interest

Animal models and CNS regions of interest involved in studying astrocyte morphology.

Figure 5. Part A: Human Astrocytes from Cortical Layers I and V. Part B: Rat astrocytes from cortical layer VI

A, top: six astrocytes from layer I of the human temporal lobe forming a cell aggregate with intermingling arbours, visualized through a 3‐D rendering. On the bottom, two astrocytes from layer V of the human temporal lobe with closely connected cell bodies and overlapping territories. B, morphology of astrocytes from layer VI of the rat somatosensory cortex. Includes a full mesh rendering derived from volume segmentation and a volume rendering from a thinning procedure to emphasize primary processes. (A) and (B) are adapted from (Calì et al., ; Shapson‐Coe et al., 2021) and reproduced under the Creative Commons Attribution license.

Figure 6. Papers related to different questions regarding astrocyte ultrastructure

Three examples of papers related to different biological questions regarding astrocyte ultrastructure. A, detailed astrocyte morphology is shown. Left: an electron microscopy image and a 3‐D reconstruction of an astrocyte branchlet that divides into two processes, creating reflexive contacts near the endfoot process. Reflexive contacts are highlighted in bright yellow. Right: another example of an astrocytic leaflet process is depicted, looping back to the branchlet. A reflexive contact near the loop's apex is visible but does not form a closed loop structure. B, four electron micrographs and the 3‐D reconstruction of an entire synapse with the dendritic spine (S) and the bouton (B). The perisynaptic astrocytic process that ensheath the spine is marked with an asterisk (*). Dendritic spines where the astrocytic element completely surrounds the bouton–spine interface, exhibit a significant increase in number in mice undergoing whisker stimulation. C, the neurovascular unit morphology is reconstructed. Segmented data and 3‐D reconstructions highlight the intricate organization of macroglia surrounding the vasculature. Macroglia (red and purple) wrap around capillaries at varying depths, with astrocytes specifically identified in purple. (A), (B) and (C) are adapted from (Albargothy et al., ; Aten et al., ; Genoud et al., 2006) and reproduced under the Creative Commons attribution license.