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Abstract. In adult regenerating cardiomyocytes in culture, in contrast to fetal cells, mitochondrial creatine kinase (Mi-CK) was expressed. In the same cell, two populations of mitochondria, differing in shape, in distribution within the cell and in content of Mi-CK, could be distinguished. Immunofluorescence studies using antibodies against Mi-CK revealed a characteristic staining pattern for the two types of mitochondria: giant, mostly cylindrically shaped, and, as shown by confocal laser light microscopy, randomly distributed mitochondria exhibited a strong signal for Mi-CK, whereas small, "normal" mitochondria, localized in rows between myofibrils, gave a much weaker signal. Transmission EM of the giant mitochondria demonstrated paracrystalline inclusions located between cristae membranes. Immunogold labeling with anti-Mi-CK antibodies revealed a specific decoration of these inclusions for Mi-CK. Addition of 20 mM creatine, the substrate of Mi-CK, to the essentially creatine-free culture medium caused the disappearance of the giant cylindrically shaped mitochondria as well as of the paracrystalline inclusions, accompanied by an increase of the intracellular level of total creatine. Replacement of creatine in the medium by the creatine analogue and competitor B-guanidinopropionic acid caused the reappearance of the enlarged mitochondria. It is believed that the accumulation of Mi-CK within the paracrystalline inclusions, similar to those observed in certain myopathies, represents a compensatory effect of the cardiomyocytes to cope with a metabolic stress situation caused by low intracellular total creatine levels. IT was shown in preceding publications (3,5,9,11,17,29) that adult rat cardiomyocytes (ARCs) t in culture represent a suitable in vitro system for the investigation of cardiac differentiation. There it was demonstrated that ARCs in long term cultures in many ways repeat embryonic and fetal differentiation steps. Fully differentiated heart cells express mitochondrial creatine kinase (Mi-CK) in conjunction with the cytosolic creatine kinase isoforms MM-, MB-, or BB-CK, with a significant fraction of MM-CK being specifically associated subcellularly in a compartmented fashion at intracellular sites of high energy turnover, e.g., at the myofibriUar M-band, the sarcoplasmic reticulum, etc. (39, 59), which has led us to the postulation of the phosphocreatine-circuit model (60, 61). Expression and appearance of Mi-CK in rat heart in vivo, however, occurs only during postnatal development paralleling more or less the appearance of MM-CK within the M-line structure of myofibrils which does not occur before 2 wk after birth (35). Concomitantly, fetal cardiomyocytes in culture do not express and accumulate Mi-CK. ARCs in culture though accumulate Mi-CK 1. Abbreviations used in this paper: ARC, adult rat cardiomyoeyte; Mi-CK, mitochondrial creatine kinase.in their mitochondria, a feature characteristic for adult heart tissue and freshly isolated rod-shaped ARC. This is converse to contractile structures in cult...

The phosphocreatine content of smooth muscle is of similar magnitude to ATP. Thus the function of the creatine kinase system in this tissue cannot simply be regarded as an energy buffer. Thus an understanding of its role in smooth muscle behavior can point to CK function in other systems. From our perspective CK function in smooth muscle is one example of a more general phenomenon, that of the co-localization of ATP synthesis and utilization. In an interesting and analogous fashion distinct glycolytic cascades are also localized in regions of the cell with specialized energy requirements. Similar to CK, glycolytic enzymes are known to be localized on thin filaments, sarcoplasmic reticulum and plasma membrane. In this chapter we will describe the relations between glycolysis and smooth muscle function and compare and contrast to that of the CK system. Our goal is to more fully understand the significance of the compartmentation of distinct pathways for ATP synthesis with specific functions in smooth muscle. This organization of metabolism and function seen most clearly in smooth muscle is likely representative of many other cell types.

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