Warning: include(phj2-admin/kernel/config.php): failed to open stream: No such file or directory in /zstorage3/vhosts/fz.kiev.ua/httpdocs/index.php on line 45

Warning: include(): Failed opening 'phj2-admin/kernel/config.php' for inclusion (include_path='.:/usr/local/share/pear') in /zstorage3/vhosts/fz.kiev.ua/httpdocs/index.php on line 45

Warning: Cannot modify header information - headers already sent by (output started at /zstorage3/vhosts/fz.kiev.ua/httpdocs/index.php:45) in /zstorage3/vhosts/fz.kiev.ua/httpdocs/kernel/content.php on line 3
Fiziologichnyi Zhurnal - <a id="lj1_ret_bk_to_jl0_href_id" href="?list0=17">Fiziol. Zh. 2015;</a> <a id="lj1_ret_bk_to_jl0_href_id" href="?list1=91">61(6):</a> 17-25
Українська Русский English

ISSN 2522-9028 (Print)
ISSN 2522-9036 (Online)
DOI: https://doi.org/10.15407/fz

Fiziologichnyi Zhurnal

is a scientific journal issued by the

Bogomoletz Institute of Physiology
National Academy of Sciences of Ukraine

Editor-in-chief: V.F. Sagach

The journal was founded in 1955 as
1955 – 1977 "Fiziolohichnyi zhurnal" (ISSN 0015 – 3311)
1978 – 1993 "Fiziologicheskii zhurnal" (ISSN 0201 – 8489)
1994 – 2016 "Fiziolohichnyi zhurnal" (ISSN 0201 – 8489)
2017 – "Fiziolohichnyi zhurnal" (ISSN 2522-9028)

Fiziol. Zh. 2015; 61(6): 17-25


CA2+ ACCUMULATION IN ISOLATED RAT HEART MITOCHONDRIA UNDER MAINTENANCE OF MITOCHONDRIAL POTENTIAL

A.Yu. Budko, N.A. Strutynska, I.Yu. Okhay, O.M. Semenykhina, V.F. Sagach

    O.O. Bogomoletz Institute of Physiology, National Academy of Sciences of Ukraine, Kyiv
DOI: https://doi.org/10.15407/fz61.06.017

Abstract

It is known that mitochondria can accumulate calcium, which regulates energy metabolism and cell death. About 90% of energy of cardiomyocytes is synthesized in mitochondria. Heart cells are also affected by the rapid changes in the Ca2+ concentration in the cytoplasm. Therefore, mitochondrial Ca2+-accumulation ability is crucial. The aim of our work was to study the accumulation of Ca2+ in isolated rat heart mitochondria in the presence of mitochondrial potential and different extramitochondrial Ca2+ concentrations. Isolated organelles were loaded with fluorescent dye Fluo-4 AM (2,5 µmol/l) at a temperature of 26°C for 30 min. It has been revealed that under these conditions high mitochondrial potential was maintained sufficiently, which is necessary for the functioning of the calcium transporting system in organelles. We established that mitochondria have a limited ability to store ionized calcium, as addition of Ca2+ ion in concentrations of 10, 20, 50 µmol/l ensures a certain level of accumulation in organelles with further fluorescent signal growth cessation. Addition of 100 µmol/l Ca2+ to isolated mitochondria resulted in a significant increase in fluorescence intensity (46% in the fifth minute, compared to the fluorescence when 20 µmol/l Ca2+ was added) and likely to activation of cation release. It was shown that ruthenium red (10-5 mol/l), an inhibitor of Ca2+ -uniporter, prevented accumulation of calcium ions in organelles by 89%, in the presence of 100 µmol/l Ca2+ . It was clearly seen that heart mitochondria require Mg2+-ATP complex (3 mmol/l) to accumulate Ca2 +, likely to maintain the inner membrane potential, activity of Ca2+ uniporter and energetic processes in organelles. Thus, the process of Ca2+ accumulation in rat heart mitochondria requires the maintenance of mitochondrial potential, activity of Ca2+-uniporter, depends on extramitochondrial Ca2 +concentration and presence of Mg2+-ATP complex.

Keywords: isolated mitochondria; heart; Ca2 +; flow cytometry; fluorescence probe Fluo-4 AM.

References

  1. Garlid KD, Costa AD, Quinlan CL, Pierre SV, Santos PD. Cardioprotective Signaling to Mitochondria. J Mol Cell Cardiol. 2009; 46:858-66. CrossRef PubMed PubMedCentral
  2.  
  3. Javadov S, Karmazyn M, Escobales N. Mitochondrial Permeability Transition Pore Opening as a Promising Therapeutic Target in Cardiac Diseases. J Pharmacol and Exp Therap. 2009; 330(3):670-8. CrossRef PubMed
  4.  
  5. Dedkova EN, Blatter LA. Measuring mitochondrial function in intact cardiac myocytes. J Mol and Cell Cardiol. 2012; 52:48-61. CrossRef PubMed PubMedCentral
  6.  
  7. Frasier ChR. The role of cardiac mitochondria in myocardial ischemia/reperfusion injury [dissertation]. East Carolina University; 2012.
  8.  
  9. Boyman L, Chikando AC, Williams GSB, Khairallah RJ, Kettlewell S, Ward ChW, Smith GL, Kao JPY, Lederer WJ. Calcium Movement in Cardiac Mitochondria. Biophysical J. 2014; 107:1289-1301. CrossRef PubMed PubMedCentral
  10.  
  11. Glancy B, Balaban RS. Role of Mitochondrial Ca2+ in the Regulation of Cellular Energetics. Biochemistry. 2012; 51:2959-73. CrossRef PubMed PubMedCentral
  12.  
  13. Brookes PS, Yoon Y, Robotham JL, Anders MW, Sheu SS. Calcium, ATP, and ROS: a mitochondrial love-hate triangle. AJP-Cell Physiol. 2004; 287:817-33. CrossRef PubMed
  14.  
  15. Dedkova EN, Blatter LA. Mitochondrial Ca2+ and the heart. Cell Calcium. 2008; 44:77-91. CrossRef PubMed
  16.  
  17. Akopova OV. The role of permeability transition pore in transmembrane Ca2+-exchange in mitochondria. Ukr Biochem J. 2008; 80(3):40-7 [Ukrainian].
  18.  
  19. Strutynska NA, Timoshchuk SV, Vavilova GL, Kotsuruba AV, Sagach VF. Expression of mitochondrial uncoupling protein 3 and the sensitivity of mitochondrial permeability transition pore opening to Ca2+ in old rat heart under activation of biosynthesis of coenzyme Q. Fiziol Zh. 2009; 55(3):44-54 [Ukrainian].
  20.  
  21. Kolomiets OV, Danylovych YuV, Danylovych GV, Kosterin SO. Ca2+ accumulation study in isolated smooth muscle mitochondria using Fluo-4 AM. Ukr Biochem J. 2013; 85(4):30-9 [Ukrainian].
  22.  
  23. Babich LG, Shlykov SG, Naumova NV, Kosterin SO. Investigation of Ca2+-induced changes of membrane potential of smooth muscle mitochondria using flow cytometric analysis. Ukr Biochem J. 2007; 79(6):34-41 [Ukrainian] .
  24.  
  25. Babich LG, Shlykov SG, Naumova NV, Kosterin SO. Use of flow cytometry method to determine Ca2+ content in mitochondria and influence of calmodulin antagonists on it. Ukr Biochem J. 2008; 80(4):51-8 [Ukrainian].
  26.  
  27. Zalessky VN. Molecular diagnostics: laser-scanning and flow cytometry in the analysis of apoptosis. Ukr Med Chasopys. 2010; 78(4):27-31 [Ukrainian].
  28.  
  29. Sagach VF, Vavilova GL, Strutyns'ka NA, Rudyk OV. The aging increase in the sensitivity of the mitochondrial permeability transition pore opening to inductors in rat heart. Fiziol Zh. 2004; 50(2): 49-63 [Ukrainian].
  30.  
  31. Lowry OH, Rosebrough NJ, Farr AL, Randall RJ. Protein measurement with the Folin phenol reagent. J Biol Chem. 1951; (1):265-75.
  32.  
  33. Brand MD, Brown GC, Cooper CE, Editors, Bioenergetics: a practical approach. Oxford.: IRL Press. 1995; 39–62.
  34.  
  35. Borutaite V, Mildaziene V, Brown GC, Brand MD. Control and kinetic analysis of ischemia-damaged heart mitochondria: which parts of the oxidative phosphorylation system are affected by ischemia? Biochim and Biophys Acta. 1995; (1272):154-8.
  36.  
  37. Jean-Quartier C, Bondarenko AI, Alam MR, Trenker M, WaldeckWeiermair M, Malli R, Graier WF. Studying mitochondrial Ca2+ uptake – a revisit. Mol and Cell Endocrinol. 2012; 353:114-127. CrossRef PubMed PubMedCentral
  38.  
  39. Szabadkai G, Duchen MR. Mitochondria: The Hub of Cellular Ca2+ signaling. Physiology. 2008; 23:84-94. CrossRef PubMed
  40.  
  41. Dedkova EN, Blatter LA. Calcium signaling in cardiac mitochondria. J Mol and Cell Cardiol. 2013; 58:125-33. CrossRef PubMed PubMedCentral
  42.  

© National Academy of Sciences of Ukraine, Bogomoletz Institute of Physiology, 2014-2019.