Südamerakkude sünnijärgse arengu bioenergeetilised aspektid: struktuuri ja funktsiooni vaheliste seoste väljakujunemine

Taust ja eesmärk. Täiskasvanud südamerakkude bioenergeetikas on valdavaks ATP genereerimisemehhanismiks mitokondriaalne oksüdatiivne fosforüülimine, mis katab tavatingimustel üle 90% südame energeetilisest vajadusest. Mitokondrid paiknevad kardiomüotsüütides korrapäraselt müofibrillide vahel, asetudes kohakuti libisevate filamentide kontaktalaga (sarkomeeri anisotroopne (A) vööt). Aktomüosiinisüsteem, mitokondrid, sarkoplasmaatiline võrgustik ja nendega seotud tsütoskeleti valgud moodustavad rakus ühtse struktuurse ja funktsionaalse terviku, nn energeetilise üksuse (EÜ), mis reguleerib efektiivselt energia tootmist ja fosforüülrühma ülekannet. Vahetult pärast sündi on mitokondrite paigutus ebakorrapärane, täiskasvanud kardiomüotsüüdiga võrreldes on oluliselt erinev ka südamerakkude metabolism ning energiaülekande regulatsioon. Töö eesmärgiks oli uurida südame mitokondriaalse hingamise regulatsiooni mehhanismide väljakujunemist südame sünnijargses arengus ning selle seotust mitokondrite ja tubuliini isovormide rakusisese paigutusega. Töö tulemused võimaldavad selgitada südamerakkude teatud patoloogiliste seisundite etioloogiat. Metoodika. Kardiomüotsüüdid isoleeriti, perfuseerides katseloomade (Wistari liini rotid) südant kollagenaas A lahusega. Skineeritud kiudude eraldamiseks kasutati meetodit, mille käigus lihaskiud eraldatakse õrnalt pintsettidega ja töödeldakse seejärel saponiiniga. Permeabiliseeritud kardiomüotsüütide ja skineeritud kiudude hapnikutarbimine registreeriti suure lahutusvõimega oksügraafil. Preparaatide visualiseerimiseks kasutati konfokaalmikroskoope Zeiss LSM 510 ja Olympus FluoView FV10i-W. Tulemused. Katseloomade sünni järel toimuvad esimese pooleteise kuu jooksul südamerakuenergiaülekande regulatsioonis kiired muutused: mitokondrite paigutus muutub korrapäraseks, toimub tsütoskeleti funktsionaalselt oluliste komponentide paigutumine mitokondrite lähedusse ja sellega samal ajal kasvavad oluliselt difusioonitakistused adenosiindifosfaadile (Km(ADP) väärtus suureneb 75,0 ・} 4,5 μM 3 päeva vanuste rottide kardiomüotsüütides kuni 317 ・} 29,5 μM vorreldes 84päevaste katseloomadega) ning käivitub kreatiinkinaasi-fosfokreatiini ülekandevõrgustik mitokondrite ja tsütosoolsete ATPaaside vahel. Järeldused. Katseloomade sünnijargse arengu käigus toimuvad dünaamilised muutusedkardiomüotsüütide struktuuris, millega kaasnevad muutused nende funktsioonis. Funktsionaalsete vastasmõjude tekkimine mitokondrite ja tsütoskeleti komponentide vahel on eelduseks täiskasvanud südamerakule omase energiametabolismi väljakujunemiseks. Eesti Arst 2013; 92(7):372–38

Molecular System Bioenergics of the Heart: Experimental Studies of Metabolic Compartmentation and Energy Fluxes versus Computer Modeling †

In this review we analyze the recent important and remarkable advancements in studies of compartmentation of adenine nucleotides in muscle cells due to their binding to macromolecular complexes and cellular structures, which results in non-equilibrium steady state of the creatine kinase reaction. We discuss the problems of measuring the energy fluxes between different cellular compartments and their simulation by using different computer models. Energy flux determinations by 18O transfer method have shown that in heart about 80% of energy is carried out of mitochondrial intermembrane space into cytoplasm by phosphocreatine fluxes generated by mitochondrial creatine kinase from adenosine triphosphate (ATP), produced by ATP Synthasome. We have applied the mathematical model of compartmentalized energy transfer for analysis of experimental data on the dependence of oxygen consumption rate on heart workload in isolated working heart reported by Williamson et al. The analysis of these data show that even at the maximal workloads and respiration rates, equal to 174 μmol O2 per min per g dry weight, phosphocreatine flux, and not ATP, carries about 80–85% percent of energy needed out of mitochondria into the cytosol. We analyze also the reasons of failures of several computer models published in the literature to correctly describe the experimental data

Matters of the heart in bioenergetics: mitochondrial fusion into continuous reticulum is not needed for maximal respiratory activity.

International audienceMitochondria are dynamic structures for which fusion and fission are well characterized for rapidly dividing cells in culture. Based on these data, it has recently been proposed that high respiratory activity is the result of fusion and formation of mitochondrial reticulum, while fission results in fragmented mitochondria with low respiratory activity. In this work we test the validity of this new hypothesis by analyzing our own experimental data obtained in studies of isolated heart mitochondria, permeabilized cells of cardiac phenotype with different mitochondrial arrangement and dynamics. Additionally, we reviewed published data including electron tomographic investigation of mitochondrial membrane-associated structures in heart cells. Oxygraphic studies show that maximal ADP-dependent respiration rates are equally high both in isolated heart mitochondria and in permeabilized cardiomyocytes. On the contrary, these rates are three times lower in NB HL-1 cells with fused mitochondrial reticulum. Confocal and electron tomographic studies show that there is no mitochondrial reticulum in cardiac cells, known to contain 5,000-10,000 individual, single mitochondria, which are regularly arranged at the level of sarcomeres and are at Z-lines separated from each other by membrane structures, including the T-tubular system in close connection to the sarcoplasmic reticulum. The new structural data in the literature show a principal role for the elaborated T-tubular system in organization of cell metabolism by supplying calcium, oxygen and substrates from the extracellular medium into local domains of the cardiac cells for calcium cycling within Calcium Release Units, associated with respiration and its regulation in Intracellular Energetic Units

Formation of highly organized intracellular structure and energy metabolism in cardiac muscle cells during postnatal development of rat heart

AbstractAdult cardiomyocytes have highly organized intracellular structure and energy metabolism whose formation during postnatal development is still largely unclear. Our previous results together with the data from the literature suggest that cytoskeletal proteins, particularly βII-tubulin, are involved in the formation of complexes between mitochondria and energy consumption sites. The aim of this study was to examine the arrangement of intracellular architecture parallel to the alterations in regulation of mitochondrial respiration in rat cardiomyocytes during postnatal development, from 1day to 6months.Respirometric measurements were performed to study the developmental alterations of mitochondrial function. Changes in the mitochondrial arrangement and cytoarchitecture of βII- and αIV-tubulin were examined by confocal microscopy.Our results show that functional maturation of oxidative phosphorylation in mitochondria is completed much earlier than efficient feedback regulation is established between mitochondria and ATPases via creatine kinase system. These changes are accompanied by significant remodeling of regular intermyofibrillar mitochondrial arrays aligned along the bundles of βII-tubulin. Additionally, we demonstrate that formation of regular arrangement of mitochondria is not sufficient per se to provide adult-like efficiency in metabolic feed-back regulation, but organized tubulin networks and reduction in mitochondrial outer membrane permeability for ADP are necessary as well. In conclusion, cardiomyocytes in rat heart become mature on the level of intracellular architecture and energy metabolism at the age of 3months