Mitochondrion Reconstructed by Electron Tomography
Author: T.G. Frey
Research sponsored by the Western States Affiliate of the American Heart Association
[bibliplug last_name=Perkins year=1997]
Tilt Series of a Mitochondrion
The first step is to collect a tilt series from a semi-thick section.
The tilt series shows the three-dimensional structure viewed from many different angles in projection onto two-dimensional images; this is the information that it needed to calculate its structure in three-dimensions. The three-dimensional structure or “Tomogram” can be calculated from these images.
60 Electron Micrographs Recorded at 2 Degree Intervals on the JEOL 4000 400 kV Electron Microscope
A series of electron micrographs recorded from a 0.5 micron section of a mitochondrion in a chick cerebellum dendrite. There are 61 images in the tilt series recorded at two degree intervals from -60° to +60°. This movie shows the images sequentially 12 per second as if viewed live while the section is being rotated.
Electron Tomogram of a Mitochondrion
A tomogram is a thee-dimensional picture. The movie shows sections of the tomogram one right after the other.
This is a three-dimensional reconstruction of a dendritic mitochondrion from chick cerebellum reconstructed tomographically from a tilt series of a 0.5 micron section. This Quicktime Movie steps through every fourth section of the three dimensional tomogram.
Segmenting the Tomogram
The tracings from all sections are modeled as three-dimensional surfaces and displayed as a three-dimensional model by the program Synu developed at the National Center for Microscopy and Imaging Research at UCSD.
Three-Dimensional Model of a Fully Segmented Mitochondrion
The tomogram was segmented by manually contouring into regions bounded by the outer, inner boundary, and cristal membranes shown in dark blue, light blue, and yellow respectively. Only the initial frame shows the outer membrane; in subsequent frames the inner boundary membrane is displayed as translucent so that the cristal membrane can be viewed through it. Note that the inner boundary membrane and the cristal membrane are components of the inner mitochondrial membrane and are one continuous surface joined at 30 nm diameter tubular connections or crista junctions.
Four Cristae and Endoplasmic Reticulum
In this case only four crista are displayed in separate colors to demonstrate the different morphologies observed in this mitochondrion. These range from the red crista at upper right which is a single tubule through somewhat more complex cristae with several tubular crista junctions to the inner boundary membrane to the large yellow crista consisting of a large lamellar compartment with many crista junctions. Two segments of the endoplasmic reticulum adjacent to the mitochondrion are shown in light brown and green.
Three-dimensional reconstruction of a dendritic mitochondrion from chick cerebellum reconstructed tomographically from a tilt series of a 0.5 micron section. The tomogram was segmented by manually contouring into regions bounded by the outer and inner boundary membranes shown in dark blue and light blue respectively. Four different cristae are displayed in red, yellow, green and grey along with two segments of endoplasmic reticulum in green and light brown. The outer membrane and the inner boundary membrane are rendered translucent so that the cristal membranes can be viewed through it. Note that the inner boundary membrane and the cristal membrane are components of the inner mitochondrial membrane and are one continuous surface joined at 30 nm diameter tubular connections or crista junctions.