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Mitochondria networks unveiled with 3D volume electron microscopy using FIB-SEM
3D rendering of individual mitochondria (various colors) in adjacent oxidative (rear) and glycolytic (front) mouse skeletal muscle cells overlaid on raw FIB-SEM data (grayscale). Data collected with ZEISS Crossbeam. Credit: National Heart, Lung, and Blood Institute at the National Institutes of Health, USA
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3D Volume Electron Microscopy Explores Mitochondrial and Muscle Substructure

作者：ZEISS Microscopy 2023 6月 12 3 分鐘 閱讀
Introduction

3D Volume Electron Microscopy Explores Mitochondrial and Muscle Substructure

FIB-SEM used to understand spatial relationships between mitochondria and sites of energy storage, utilization, and signaling.

Skeletal muscle is the most abundant tissue in humans. In an instant, it must coordinate the movement of signals and materials through relatively large muscle cells to generate force through muscle contractions. These contractions are maintained anywhere from seconds to hours. The Muscle Energetics Laboratory, led by Dr. Brian Glancy, at the National Heart, Lung, and Blood Institute at the National Institutes of Health, USA, seeks to understand how mitochondria are optimized within muscle cells to help maintain energy homeostasis during the large change in energy demand caused by muscle contractions. 3D volume electron microscopy (vEM) utilizing ZEISS Crossbeam FIB-SEM is a key technology that Dr. Glancy’s team has utilized to explore mitochondrial and muscle substructure as evidenced in many of their recent publications highlighted below.

  

Dr. Alejandro Rojas-Fernandez

There are many, many different physical interactions that are happening within these cells and when we’re doing 3D electron microscopy, we can see all of these things. This allows us to take an integrated viewpoint on muscle cell structure.

Dr. Brian Glancy

Earl Stadtman Investigator, Muscle Energetics Laboratory, National Heart, Lung, and Blood Institute at the National Institutes of Health, USA

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3D rendering of individual mitochondria from a glycolytic (left), oxidative (middle), and cardiac (right) muscle FIB-SEM volumes. Various colors represent individual mitochondria. Data collected with ZEISS Crossbeam FIB-SEM. Credit: National Heart, Lung, and Blood Institute at the National Institutes of Health, USA

Subcellular Connectomics

Analyzing Mitochondrial Networks

In C.K.E. Bleck et al. 2018, Dr. Glancy and team applied a connectomics approach using volume electron microscopy with FIB-SEM and SBF-SEM to quantitatively assess the mitochondrial network in cardiac, oxidative, and glycolytic muscle.

They showed that each muscle type, with its differing contraction demands, had different mitochondrial network configurations.  They also assessed mitochondria-lipid droplet interactions and found evidence that individual mitochondria may be tuned to specialize in energy distribution or calcium cycling.

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3D rendering of a region of the fast-twitch myofibrillar matrix showing branching sarcomeres. Data acquired with ZEISS Crossbeam FIB-SEM. Credit: National Heart, Lung, and Blood Institute at the National Institutes of Health, USA

3D Electron Microscopy of Sarcomeres

A Branching Myofibrillar Matrix

In T.B.  Willingham et al. 2020, Dr. Glancy's group uses volume electron microscopy with FIB-SEM to unveil that striated muscle cells form a continuous myofibrillar matrix with frequent, branching sarcomeres. Their work examines changes in branching during postnatal development and in different muscle types. They suggest a new theory for how force is generated based on a mesh-like myofibrillar network rather than many individual, parallel myofibrils.

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3D rendering and rotation of the myosin filaments within a single mouse cardiac sarcomere. Adjacent mitochondria (red), sarcotubular network (green), and a lipid droplet (cyan) are also shown. Data collected with ZEISS Crossbeam FIB-SEM. Credit: National Heart, Lung, and Blood Institute at the National Institutes of Health, USA

3D Modeling of Sarcomere Interactions

Mitochondrial Networks Influences Sarcomere and Myosin Filament Structure

Mitochondria must supply a constant energy stream to actin and myosin filaments within muscle sarcomeres to sustain muscle contractions over time. In P. Katti et al. 2022, 3D electron microscopy is used to examine variations in sarcomere cross-sectional area and provide evidence that both sarcomere structure and myofilament interactions are influenced by the location and orientation of mitochondria within muscle cells.

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3D rendering and fly through of the Drosophila leg muscle myofibrillar network. Data collected with ZEISS Crossbeam FIB-SEM. Credit: National Heart, Lung, and Blood Institute at the National Institutes of Health, USA

3D EM of Drosophila Muscle Types

Analyzing Myofibrillar Connectivity

Continuing their work from T.B. Willingham et al. 2020 mentioned above, Dr. Glancy and team looked into the extent to which myofibrillar connectivity is evolutionarily conserved as well as mechanisms which regulate the specific architecture of sarcomere branching. In P.T. Ajayi et al. 2022,  they present 3D electron microscopy evidence which indicates fruit flies have a myofibrillar connectivity on/off switch that is regulated by both cell-type dependent and independent mechanisms.

3D electron microscopy allows us to see how all the pieces fit together and how this puzzle changes across different cell types and environments. As a result, we have a better understanding of how a muscle cell is built, how it works, and how we may be able to fix it when it doesn’t.

Dr. Brian Glancy

Earl Stadtman Investigator, Muscle Energetics Laboratory, National Heart, Lung, and Blood Institute at the National Institutes of Health, USA
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