The aim of this work was to characterize quantitatively the arrangement of mitochondria in heart and skeletal muscles. We studied confocal images of mitochondria in nonfixed cardiomyocytes and fibers from soleus and white gastrocnemius muscles of adult rats. The arrangement of intermyofibrillar mitochondria was analyzed by estimating the densities of distribution of mitochondrial centers relative to each other (probability density function). In cardiomyocytes (1,820 mitochondrial centers marked), neighboring mitochondria are aligned along a rectangle, with distance between the centers equal to 1.97 Ϯ 0.43 and 1.43 Ϯ 0.43 m in the longitudinal and transverse directions, respectively. In soleus (1,659 mitochondrial centers marked) and white gastrocnemius (621 pairs of mitochondria marked), mitochondria are mainly organized in pairs at the I-band level. Because of this organization, there are two distances characterizing mitochondrial distribution in the longitudinal direction in these muscles. The distance between mitochondrial centers in the longitudinal direction within the same I band is 0.91 Ϯ 0.11 and 0.61 Ϯ 0.07 m in soleus and white gastrocnemius, respectively. The distance between mitochondrial centers in different I bands is ϳ3.7 and ϳ3.3 m in soleus and gastrocnemius, respectively. In the transverse direction, the mitochondria are packed considerably closer to each other in soleus than in white gastrocnemius, with the distance equal to 0.75 Ϯ 0.22 m in soleus and 1.09 Ϯ 0.41 m in gastrocnemius. Our results show that intermyofibrillar mitochondria are arranged in a highly ordered crystal-like pattern in a muscle-specific manner with relatively small deviation in the distances between neighboring mitochondria. This is consistent with the concept of the unitary nature of the organization of the muscle energy metabolism. confocal microscopy; quantitative analysis; cardiac and skeletal muscles; probability density function; unitary structure of cells RECENT STUDIES HAVE SHOWN the existence of multiple specific functional interactions among mitochondria, sarcoplasmic reticulum (SR), and myofibrils in permeabilized muscle fibers (5,14,30,34). Namely, endogenous ATP has been shown to be more efficient than exogenous ATP in maintaining calcium uptake into SR (14). In addition, kinetic studies have shown a direct supply of endogenous ADP from ATPases to mitochondria (30, 34). Such interaction can be explained by the existence of localized intracellular diffusion restrictions (28, 39). A mild treatment of the fibers with trypsin leads to the removal of these diffusion restrictions, and at the same time, distribution of mitochondria in the fiber is changed from regular arrangement in the control to random distribution after the treatment (28). Similarly, in ischemic hearts, various alterations in mitochondrial function such as the significant decrease in maximal respiration rate and half-saturation constant for ADP were observed in parallel with the changes in structural organization of the cardiac muscle cells (7,1...

Heterogeneity of ADP diffusion and regulation of respiration were studied in permeabilized cardiomyocytes and cardiac fibers in situ and in silico. Regular arrangement of mitochondria in cells was altered by short-time treatment with trypsin and visualized by confocal microscopy. Manipulation of matrix volumes by changing K(+) and sucrose concentrations did not affect the affinity for ADP either in isolated heart mitochondria or in skinned fibers. Pyruvate kinase (PK)-phosphoenolpyruvate (PEP) were used to trap ADP generated in Ca,MgATPase reactions. Inhibition of respiration by PK-PEP increased 2-3 times after disorganization of regular mitochondrial arrangement in cells. ADP produced locally in the mitochondrial creatine kinase reaction was not accessible to PK-PEP in intact permeabilized fibers, but some part of it was released from mitochondria after short proteolysis due to increased permeability of outer mitochondrial membrane. In in silico studies we show by mathematical modeling that these results can be explained by heterogeneity of ADP diffusion due to its restrictions at the outer mitochondrial membrane and in close areas, which is changed after proteolysis. Localized restrictions and heterogeneity of ADP diffusion demonstrate the importance of mitochondrial functional complexes with sarcoplasmic reticulum and myofibrillar structures and creatine kinase in regulation of oxidative phosphorylation.

In this review we analyze the concepts and the experimental data on the mechanisms of the regulation of energy metabolism in muscle cells. Muscular energetics is based on the force-length relationship, which in the whole heart is expressed as a Frank-Starling law, by which the alterations of left ventricle diastolic volume change linearly both the cardiac work and oxygen consumption. The second basic characteristics of the heart is the metabolic stability--almost constant levels of high energy phosphates, ATP and phosphocreatine, which are practically independent of the workload and the rate of oxygen consumption, in contrast to the fast-twitch skeletal muscle with no metabolic stability and rapid fatigue. Analysis of the literature shows that an increase in the rate of oxygen consumption by order of magnitude, due to Frank-Starling law, is observed without any significant changes in the intracellular calcium transients. Therefore, parallel activation of contraction and mitochondrial respiration by calcium ions may play only a minor role in regulation of respiration in the cells. The effective regulation of the respiration under the effect of Frank-Starling law and metabolic stability of the heart are explained by the mechanisms of functional coupling within supramolecular complexes in mitochondria, and at the subcellular level within the intracellular energetic units. Such a complex structural and functional organisation of heart energy metabolism can be described quantitatively by mathematical models.