New Tools to Visualize Mitochondria

To accurately assess mitochondria morphology in healthy and diseased brain tissue, Dr. Trushina's team — in collaboration with Mayo Clinic's Microscopy and Cell Analysis Core — developed a novel 3-D electron microscopy (EM) reconstruction technique. The method allows scientists to see 3-D details of mitochondrial structure in the context of a 3-D human or mouse brain.

The group also developed conditions that allow tissue preparation and processing that's free from artifacts often associated with the dynamic nature of mitochondrial morphological changes.

Using the technique they developed, researchers in the Mitochondrial Neurobiology and Therapeutics Lab identified a novel mitochondrial phenotype termed mitochondria-on-a-string (MOAS). Further, the lab linked MOAS to the fundamental mitochondrial response to stress.

The development of this advanced tool led to multiple collaborations with researchers at and external to Mayo Clinic, including Roberta Diaz Brinton, Ph.D., at the University of Arizona, and Stella Papa, M.D., at the Emory University School of Medicine, among others. Collaborations are focused on the molecular mechanisms of Alzheimer's disease and Parkinson's disease using human, primate and mouse brain tissue affected in each disease.

Detailed visualization of mitochondrial morphology using the lab's 3-D EM reconstruction technique to better understand disease progression could help researchers identify relevant therapeutics.

The video above is an animated 3-D reconstruction of mitochondrial structure in the hippocampal tissue of an APP-PS1 mouse at 24 weeks of age.

For 3-D reconstruction of the mitochondrial structure, Dr. Trushina's team began by inverting the grayscale of the individual electron microscopy section images so that the organelles became bright objects.

Researchers then sequentially coregistered the inverted images using the Normalized Mutual Information 2-D registration program in Analyze, a comprehensive multidimensional medical image processing, visualization and analysis software package developed by the Biomedical Imaging Resource Core at Mayo Clinic. Sequential coregistering is an automated procedure that aligns similar images based on the statistical distribution of paired pixels compared to the distribution in either image alone.

Finally, the lab rendered the inverted, coregistered stack using maximum intensity projection. Each pixel in the rendered image represents the brightest voxel in a ray from the viewer's eye through the entire stack of sections, greatly improving scientists' ability to see and understand 3-D mitochondrial structure in context.

The video above shows 3-D electron microscopy reconstruction of mitochondria in multiple neuropils in the hippocampi of nontransgenic (left) and APP-PS1 (right) mice. Mitochondria in Alzheimer's disease mice form MOAS, much longer structures compared with nontransgenic mice. Formation of MOAS could protect mitochondria from degradation, preserving residual functions and promoting cell survival under stress.

Two representative sets of standard TEM images of 0.09 μm thickness; from 10 to 40 consecutive serial sections were taken into reconstruction.