Indeed, the majority of mitochondria in mammalian skeletal muscle are located in pairs on the z-line side of t-tubules (e.g. Mitochondrial localization in mammalian fibres is similar in oxidative fatigue-resistant and glycolytic fatigue-sensitive muscle fibres. The densely arranged filaments of actin and myosin in muscle dedicated to contraction, which make up more than 80% of the cell volume in skeletal muscle, leaves little free space for mitochondria to move around. ![]() In adult skeletal and cardiac muscle, it is unclear as to whether functional networks of mitochondria exist and how extensive mitochondrial interconnections are –. These connections can form and break quite readily and allow diffusion of fluorescent labels between distant mitochondrial areas which suggest that the mitochondrial matrix has a uniform internal ionic and protein solution –. Dynamic formation of mitochondria to mitochondria connections has been demonstrated in cells as diverse as cortical neurones and fibroblasts. For example, in adult neurones and immature or foetal cardiomyocytes, mitochondria can reversibly elongate and fuse under hypoxic and other stressful conditions. Mitochondria in organisms as diverse as fungi and mice can also adapt quickly to metabolic disturbances within the cell. In some cells, mitochondria are dynamic organelles that change their shape and develop protrusions within minutes even in the absence of any external stimulus –. ![]() The spatiotemporal distribution of mitochondria is not fixed in most cell types but frequently, large numbers are found close to sites of high metabolic demand. In mammalian cells, mitochondria exist in a variety of forms from the almost universal picture of an ovoid structure not more than one µm long seen in cells ranging in size from hepatocytes to neurones to the long thread-like branching structures attaining a length of 50 µm or longer found in human fibroblasts. Furthermore, the limited and reversible effects of targeted FCCP application with the multifunctional pipette highlight its advantages over bulk application of compounds to isolated cells. In conclusion, our results indicate that extensive networks of interconnected mitochondria do not exist in skeletal muscle. After a pulse of FCCP, cytosolic was unchanged and fibres contracted in response to electrical stimulation. Similar results were observed when two sites along the fibre were pulsed sequentially with FCCP. At distances greater than 50 µm away from the site of FCCP application, the mitochondrial TMRE signal was unchanged. After washout of FCCP, the TMRE signal partially recovered. A pulse of carbonyl cyanide 4-(trifluoromethoxy) phenylhydrazone (FCCP, 100 µM) applied to a small area of the muscle fibre (30 µm in diameter) produced a rapid decrease in the mitochondrial TMRE signal (indicative of depolarization) to 38% of its initial value. Cytosolic free was monitored with fluo-3. Mitochondrial membrane potential was monitored with tetramethylrhodamine ethyl ester (TMRE). Compounds were delivered locally to the end or side of single adult mouse skeletal muscle fibres to test whether changes in mitochondrial membrane potential were transmitted to more distant located mitochondria. Two channels on either side of the central channel use suction to create a hydrodynamically confined flow zone and remove compounds completely from the bulk solution to internal waste compartments. ![]() The central channel in the pipette delivers compounds to a restricted region of the sarcolemma, typically 30 µm in diameter. Here, we use a novel three channelled microflow device, the multifunctional pipette, to test whether mitochondria in mouse skeletal muscle connect to each other. In contrast, mitochondria in adult mammalian skeletal muscle fibres show little motility over several hours. In cells, such as neurones and immune cells, mitochondria can form dynamic and extensive networks that change over the minute timescale.
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