Detection of Mitochondrial ATP-Sensitive Potassium Channels in Rat Cardiomyocytes

  • Cuong, Dang Van (Department of Physiology and Biophysics, College of Medicine, and 2020 Cardiovascular Institute, Inje University) ;
  • Kim, Na-Ri (Department of Physiology and Biophysics, College of Medicine, and 2020 Cardiovascular Institute, Inje University) ;
  • Kim, Eui-Yong (Department of Physiology and Biophysics, College of Medicine, and 2020 Cardiovascular Institute, Inje University) ;
  • Lee, Young-Suk (Department of Physiology and Biophysics, College of Medicine, and 2020 Cardiovascular Institute, Inje University) ;
  • Kim, Hyun-Ju (Department of Physiology and Biophysics, College of Medicine, and 2020 Cardiovascular Institute, Inje University) ;
  • Kang, Sung-Hyun (Department of Physiology and Biophysics, College of Medicine, and 2020 Cardiovascular Institute, Inje University) ;
  • Hur, Dae-Young (Department of Physiology and Biophysics, College of Medicine, and 2020 Cardiovascular Institute, Inje University) ;
  • Joo, Hyun (Department of Physiology and Biophysics, College of Medicine, and 2020 Cardiovascular Institute, Inje University) ;
  • Park, Young-Shik (Department of Physiology and Biophysics, College of Medicine, and 2020 Cardiovascular Institute, Inje University) ;
  • Hong, Yong-Geun (Department of Physiology and Biophysics, College of Medicine, and 2020 Cardiovascular Institute, Inje University) ;
  • Lee, Sang-Kyung (Department of Physiology and Biophysics, College of Medicine, and 2020 Cardiovascular Institute, Inje University) ;
  • Chung, Joon-Yong (Department of Physiology and Biophysics, College of Medicine, and 2020 Cardiovascular Institute, Inje University) ;
  • Seog, Dae-Hyun (Department of Physiology and Biophysics, College of Medicine, and 2020 Cardiovascular Institute, Inje University) ;
  • Han, Jin (Department of Physiology and Biophysics, College of Medicine, and 2020 Cardiovascular Institute, Inje University)
  • 발행 : 2004.08.21

초록

Mitochondrial ATP-sensitive potassium $(mitoK_{ATP})$ channels play a role in early and late ischemic preconditioning. Nevertheless, the subunit composition of $mitoK_{ATP}$ channels remains unclear. In this study, we investigated the subunit composition of $mitoK_{ATP}$ channels in mitochondria isolated from rat cardiac myocytes. Mitochondria were visualized using the red fluorescence probe, Mitrotracker Red, while $mitoK_{ATP}$ channels were visualized using the green fluorescence probe, glibenclamide-BODIPY. The immunofluorescence confocal microscopy revealed the presence of Kir6.1, Kir6.2 and SUR2 present in the cardiac mitochondria. Western blot analysis was carried to further investigate the nature of $mitoK_{ATP}$ channels. For SUR proteins, a 140-kDa immunoreactive band that corresponded to SUR2, but no SUR1 was detected. For Kir6.2, three bands $({\sim}44,\;{\sim}46,\;and\;{\sim}30\;kDa)$ were detected, and a specific ${\sim}46-kDa$ immunoreactive band corresponding to Kir6.1 was also observed. These observations suggest that the subunits of $mitoK_{ATP}$ channels in rat myocytes include Kir6.1, Kir6.2, and a SUR2-related sulfonylurea-binding protein.

키워드

참고문헌

  1. Aguilar-Bryan L, Clement JP 4th, Gonzalez G, Kunjilwar K, Babenko A, Bryan J. Toward understanding the assembly and structure of KATP channels. Physiol Rev 78: 227-245, 1998
  2. Bajgar R, Seetharaman S, Kowaltowski AJ, Garlid KD, Paucek P. Identification and properties of a novel intracellular (mitochondrial) ATP-sensitive potassium channel in brain. J Biol Chem 276: 33369-33374, 2001 https://doi.org/10.1074/jbc.M103320200
  3. Carroll R, Gant VA, Yellon DM. Mitochondrial KATP channel opening protects a human atrial-derived cell line by a mechanism involving free radical generation. Cardiovasc Res 51: 691- 700, 2001 https://doi.org/10.1016/S0008-6363(01)00330-3
  4. Dos Santos P, Kowaltowski AJ, Laclau MN, Seetharaman S, Paucek P, Boudina S, Thambo JB, Tariosse L, Garlid KD. Mechanisms by which opening the mitochondrial ATP-sensitive K$^+$ channel protects the ischemic heart. Am J Physiol Heart Circ Physiol 283: H284-H295, 2002
  5. Garlid KD, Paucek P, Yarov-Yarovoy V, Murray HN, Darbenzio RB, D'Alonzo AJ, Lodge NJ, Smith MA, Grover GJ. Cardioprotective effect of diazoxide and its interaction with mitochondrial ATP-sensitive K$^+$channels. Possible mechanism of cardioprotection. Circ Res 81: 1072-1082, 1997 https://doi.org/10.1161/01.RES.81.6.1072
  6. Gross GJ. The role of mitochondrial KATP channels in the antiarrhythmic effects of ischaemic preconditioning in dogs. Br J Pharmacol 137: 939-940, 2002 https://doi.org/10.1038/sj.bjp.0704965
  7. Han J, Kim N, Joo H, Kim E, Earm YE. ATP-sensitive K$^+$ channel activation by nitric oxide and protein kinase G in rabbit ventricular myocytes. Am J Physiol Heart Circ Physiol 283: H1545- H1554, 2002
  8. Han J, Kim N, Cuong DV, Kim C, Kim E. Thiol-dependent redox mechanism in the modification of ATP-sensitive K$^+$channels in rabbit ventricular myocytes. Korean J Physiol Pharmacol 7: 15- 23, 2003
  9. Hanley PJ, Mickel M, Loffler M, Brandt U, Daut J. KATP channelindependent targets of diazoxide and 5-hydroxydecanoate in the heart. J Physiol 542: 735-741, 2002 https://doi.org/10.1113/jphysiol.2002.023960
  10. Inagaki N, Gonoi T, Seino S. Subunit stoichiometry of the pancreatic beta-cell ATP-sensitive K$^+$ channel. FEBS Lett 409: 232- 236, 1997 https://doi.org/10.1016/S0014-5793(97)00488-2
  11. Ichinose M, Yonemochi H, Sato T, Saikawa T. Diazoxide triggers ardioprotection against apoptosis induced by oxidative stress. Am J Physiol Heart Circ Physio 284: H2235-H2241, 2003
  12. Inoue I, Nagase H, Kishi K, Higuti T. ATP-sensitive K$^+$ channel in the mitochondrial inner membrane. Nature 352: 244-247, 1991 https://doi.org/10.1038/352244a0
  13. Kuniyasu A, Kaneko K, Kawahara K, Nakayama H. Molecular assembly and subcellular distribution of ATP-sensitive potassium channel proteins in rat hearts. FEBS Lett 552: 259-263, 2003 https://doi.org/10.1016/S0014-5793(03)00936-0
  14. Lacza Z, Snipes JA, Miller AW, Szabo C, Grover G, Busija DW. Heart mitochondria contain functional ATP-dependent K$^+$ channels. J Mol Cell Cardiol, 35: 1339-1347, 2003 https://doi.org/10.1016/S0022-2828(03)00249-9
  15. Murry CE, Jennings RB, Reimer KA. Preconditioning with ischemia: a delay of lethal cell injury in ischemic myocardium. Circulation 74: 1124-1136, 1986 https://doi.org/10.1161/01.CIR.74.5.1124
  16. Peart JN, Gross GJ. Sarcolemmal and mitochondrial KATP channels and myocardial ischemic preconditioning. J Cell Mol Med 6: 453- 464, 2002 https://doi.org/10.1111/j.1582-4934.2002.tb00449.x
  17. Rouslin W. Regulation of the mitochondrial ATPase in situ in cardiac muscle: role of the inhibitor subunit. J Bioenerg Biomembr 23: 873-888, 1991 https://doi.org/10.1007/BF00786006
  18. Seharaseyon J, Ohler A, Sasaki N, Fraser H, Sato T, Johns DC, O'Rourke B, Marban E. Molecular composition of mitochondrial ATP-sensitive potassium channels probed by viral Kir gene transfer. J Mol Cell Cardiol 32: 1923-1930, 2000 https://doi.org/10.1006/jmcc.2000.1226
  19. Suzuki M, Kotake K, Fujikura K, Inagaki N, Suzuki T, Gonoi T, Seino S, Takata K. Kir6.1: a possible subunit of ATP-sensitive K$^+$ channels in mitochondria. Biochem Biophys Res Commun 241: 693-697, 1997 https://doi.org/10.1006/bbrc.1997.7891
  20. Tokuyama Y, Fan Z, Furuta H, Makielski JC, Polonsky KS, Bell GI, Yano H. Rat inwardly rectifying potassium channel Kir6.2: cloning electrophysiological characterization, and decreased expression in pancreatic islets of male Zucker diabetic fatty rats. Biochem Biophys Res Commun 220: 532-538, 1996 https://doi.org/10.1006/bbrc.1996.0439