Electron tomography of rabbit cardiomyocyte three-dimensional ultrastructure

Abstract

The field of cardiovascular research has benefitted from rapid developments in imaging technology over the last few decades. Accordingly, an ever growing number of large, multidimensional data sets have begun to appear, often challenging existing pre-conceptions about structure and function of biological systems. For tissue and cell structure imaging, the move from 2D section-based microscopy to true 3D data collection has been a major driver of new insight. In the sub-cellular domain, electron tomography is a powerful technique for exploration of cellular structures in 3D with unparalleled fidelity at nanometer resolution.Electron tomography is particularly advantageous for studying highly compartmentalised cells such as cardiomyocytes, where elaborate sub-cellular structures play crucial roles in electrophysiology and mechanics. Although the anatomy of specific ultra-structures, such as dyadic couplons, has been extensively explored using 2D electron microscopy of thin sections, we still lack accurate, quantitative knowledge of true individual shape, volume and surface area of sub-cellular domains, as well as their 3D spatial interrelations; let alone of how these are reshaped during the cycle of contraction and relaxation. Here we discuss and illustrate the utility of ET for identification, visualisation, and analysis of 3D cardiomyocyte ultrastructures such as the T-tubular system, sarcoplasmic reticulum, mitochondria and microtubules.

Axial tubule junctions control rapid calcium signaling in atria

“…AT and TT structures and isolated AM were preserved by high-pressure freezing with an HPM100 system (Leica) as described previously (51). ET at the Electron Microscopy core facility at the European Molecular Biology Laboratory (EMBL) Heidelberg (Heidelberg, Germany) and at the EM services of the University of Colorado Boulder (Boulder, Colorado, USA) was performed as described previously (23).…”

High resolution structural evidence suggests the Sarcoplasmic Reticulum forms microdomains with Acidic Stores (lysosomes) in the heart

“…Images from each tilt-series were aligned by fiducial marker tracking and back-projected to generate two single full-thickness reconstructed volumes (tomograms), which were then combined to generate a high-resolution 3D reconstruction of the original volume of interest. Tomograms were processed and analysed using the IMOD software, also used to segment and generate 3D models of the structures of interest6970. The models were smoothed and meshed to obtain a final 3D representation, in which spatial relationships of T-tubules, sarcoplasmic reticulum, mitochondria with lysosomes can be visualised.…”

Mitochondrial Deformation During the Cardiac Mechanical Cycle

“…All tomograms were processed and analyzed using IMOD software (Mastronarde, 2005), which was also used to generate 3D models of relevant structures (Kremer et al, 1996). Models were smoothed and meshed to obtain the final 3D representation, in which spatial relations of various cardiomyocyte substructures were quantified, as described before (Rog-Zielinska et al, 2016).…”