Employing Scanning Probe Microscopy to Directly Probe Mitochondrial Physical Properties and Function

Mitochondria play a central role in the metabolism and energy production of eukaryotic cells, through several chains of events entailing many protein complexes. Mitochondrial function is governed by all parts of its structure – the outer membrane which transports ions and metabolites as well as housing active enzymes; the inner membrane which contains proteins that mediate electron transport, ATP synthesis and metabolite passage; the region between the two membranes, the intermembrane space, which contains proteins that signal the transport activities. The majority of the mitochondrial proteins reside inside the inner membrane, in a volume called the matrix. Notwithstanding the desire to understand mitochondrial function at the microscopic level, the multitude of simultaneous and sequential activities occurring in the overall process can be studied functionally by measuring physical signals under different stimuli. The size of mitochondria range from diameters of several hundred nanometers up to several micrometers. Various forms of microscopy have been used to study mitochondria. Whereas electron microscopy provides the highest resolution of their inner structure, it cannot observe their function in real time. Optical microscopy including various fluorescent techniques is able to indirectly capture electrochemical activity and function of live mitochondria but with limited resolution. In this talk I will present a multi-faceted scanning probe microscopy-based approach to the study of mitochondrial function. SPM offers several advantages in such studies: Firstly, the measurements can be made in buffer solution with viable mitochondria, so that their response to different additives can be directly observed in real time. Secondly, the high resolving power allows us to obtain 3D images of individual mitochondria, which can simultaneously be characterized mechanically through fast force measurements. Finally, by measuring “noise spectra” of the mitochondria under the influence of different additives, we can follow the organelle’s activity, and how it changes when specific mitochondrial complexes are inhibited/activated, as well as under pathologic conditions. In this talk, I will summarize this work, emphasizing the application of these different SPM measurements and how combining them gives a nice picture of healthy mitochondrial function.