Naunyn-Schmiedeberg's Archives of Pharmacology

, Volume 392, Issue 8, pp 949–959 | Cite as

Pinacidil, a KATP channel opener, stimulates cardiac Na+/Ca2+ exchanger function through the NO/cGMP/PKG signaling pathway in guinea pig cardiac ventricular myocytes

  • Keisuke Iguchi
  • Masao Saotome
  • Kanna Yamashita
  • Prottoy Hasan
  • Miyuki Sasaki
  • Yuichiro Maekawa
  • Yasuhide WatanabeEmail author
Original Article


Pinacidil, a nonselective ATP-sensitive K+ (KATP) channel opener, has cardioprotective effects for hypertension, ischemia/reperfusion injury, and arrhythmia. This agent abolishes early afterdepolarizations, delayed afterdepolarizations (DADs), and abnormal automaticity in canine cardiac ventricular myocytes. DADs are well known to be caused by the Na+/Ca2+ exchange current (INCX). In this study, we used the whole-cell patch-clamp technique and Fura-2/AM (Ca2+-indicator) method to investigate the effect of pinacidil on INCX in isolated guinea pig cardiac ventricular myocytes. In the patch-clamp study, pinacidil enhanced INCX in a concentration-dependent manner. The half-maximal effective concentration values were 23.5 and 23.0 μM for the Ca2+ entry (outward) and Ca2+ exit (inward) components of INCX, respectively. The pinacidil-induced INCX increase was blocked by L-NAME, a nitric oxide (NO) synthase inhibitor, by ODQ, a soluble guanylate cyclase inhibitor, and by KT5823, a cyclic guanosine monophosphate (cGMP)-dependent protein kinase (PKG) inhibitor, but not by N-2-mercaptopropyonyl glycine (MPG), a reactive oxygen species (ROS) scavenger. Glibenclamide, a nonselective KATP channel inhibitor, blocked the pinacidil-induced INCX increase, while 5-HD, a selective mitochondria KATP channel inhibitor, did not. In the Fura-2/AM study pinacidil also enhanced intracellular Ca2+ concentration, which was inhibited by L-NAME, ODQ, KT5823, and glibenclamide, but not by MPG and 5-HD. Sildenafil, a phosphodiesterase 5 inhibitor, increased further the pinacidil-induced INCX increase. Sodium nitroprusside, a NO donor, also increased INCX. In conclusion, pinacidil may stimulate cardiac Na+/Ca2+ exchanger (NCX1) by opening plasma membrane KATP channels and activating the NO/cGMP/PKG signaling pathway.


Pinacidil Nonselective KATP channel opener Na+/Ca2+ exchange current (INCXCardiac myocytes Patch-clamp technique 



We thank Dr. Junko Kimura and Prof. Yuichi Hattori for helpful and critical comments on the manuscript. This study was supported by Grant-in-Aids for Scientific Research (17 K11047, 16 K09428) from the Japan Society for Promotion of Science.

Author contributions

M.S., Y.M., and Y.W. conceived and designed the experiments. K.I., K.Y., H.P., and M.S. performed the experiments. K.I., K.Y., and Y.W. analyzed the data. K.I. and Y.W. wrote the article. All authors read and approved the manuscript.

Compliance with ethical standards

Conflict of interest

The authors of this manuscript have no conflict of interest to declare.


  1. Ago Y, Kawasaki T, Nashida T, Ota Y, Cong Y, Kitamoto M, Takahashi T, Takuma K, Matsuda T (2011) SEA0400, a specific Na+/Ca2+ exchange inhibitor, prevents dopaminergic neurotoxicity in an MPTP mouse model of Parkinson’s disease. Neuropharmacology 61:1441–1451CrossRefGoogle Scholar
  2. Baczkó I, Giles WR, Light PE (2004) Pharmacological activation of plasma-membrane KATP channels reduces reoxygenation-induced Ca2+ overload in cardiac myocytes via modulation of the diastolic membrane potential. Br J Pharmacol 141:1059–1067CrossRefGoogle Scholar
  3. Bers DM (2000) Calcium fluxes involved in control of cardiac myocyte contraction. Cir Res 87:275–281CrossRefGoogle Scholar
  4. Blaustein MP, Lederer WJ (1999) Sodium/calcium exchange: its physiological implications. Physiol Rev 79:763–854CrossRefGoogle Scholar
  5. Carlsson L, Abrahamsson C, Drews L, Duker G (1992) Antiarrhythmic effects of potassium channel openers in rhythm abnormalities related to delayed repolarization. Circulation 85:1491–1500CrossRefGoogle Scholar
  6. Chen S-J, Wu C-C, Yang S-N, Lin C-I, Yen M-H (2000) Abnormal activation of K+ channels in aortic smooth muscle of rats with endotoxic shock: electrophysiological and function evidence. Br J Pharmacol 131:213–222CrossRefGoogle Scholar
  7. Cuong DV, Kim N, Youm JB, Joo H, Warda M, Lee JW, Park WS, Kim T, Kang S, Kim H, Han J (2006) Nitric oxide-cGMP-protein kinase G signaling pathway induces anoxic preconditioning through activation of ATP-sensitive K+ channels in rat hearts. Am J Physiol Heart Circ Physiol 290:H1808–H1817CrossRefGoogle Scholar
  8. Eigel BN, Gursahani H, Hardley RW (2004) ROS are required for rapid reactivation of Na+/Ca2+ exchanger in hypoxic reoxygenated Guinea pig ventricular myocytes. Am J Physiol Heart Circ Physiol 286:H955–H963CrossRefGoogle Scholar
  9. Fernandes G, Dasai N, Kozlova N, Mojadadi A, Gall M, Drew E, Barratt E, Madamidola OA, Brown SG, Milne AM, Martins da Siva SJ, Whalley KM, Barratt CLR, Jovanovic A (2016) A spontaneous increase in intracellular Ca2+ in metaphase II human oocytes in vitro can be prevented by drugs targeting ATP-sensitive K+ channels. Hum Reprod 31:287–297Google Scholar
  10. Foster MN, Coetzee WA (2016) KATP channel in the cardiovascular system. Physiol Rev 96:177–252CrossRefGoogle Scholar
  11. Furukawa K, Ohshima N, Tawada-Iwata Y, Shigekawa M (1991) Cyclic GMP stimulates Na+/Ca2+ exchange in vascular smooth muscle cells in primary culture. J Biol Chem 266:12337–12341Google Scholar
  12. Gendron ME, Thorin E, Perrault LP (2004) Loss of endothelial KATP channel-dependent, NO-mediated dilation of endocardial resistance coronary arteries in pigs with left ventricular hypertrophy. Br J Pharmacol 143:285–291CrossRefGoogle Scholar
  13. Goldhaber JI (1996) Free radical enhance Na+/Ca2+ exchange in ventricular myocytes. Am J Physiol 271:H823–H833Google Scholar
  14. Gonzalez DR, Treuer A, Sun Q-A, Stamler JS, Hare JM (2009) S-nitrosylation of cardiac ion channels. J Cardiovasc Pharmacol 54:188–195CrossRefGoogle Scholar
  15. Han J, Kim N, Joo H, Kim E, Earm Y (2002a) ATP-sensitive K+ channel activation by nitric oxide and protein kinase G in rabbit ventricular myocytes. Am J Physiol Heart Circ Physiol 283:H1545–H1554CrossRefGoogle Scholar
  16. Han J, Kim N, Park J, Seog D-H, Joo H, Kim E (2002b) Opening of mitochondrial ATP-sensitive potassium channels evokes oxygen radical generation in rabbit heart slices. J Biochem 131:721–727CrossRefGoogle Scholar
  17. Hinata M, Matsuoka I, Iwamoto T, Watanabe Y, Kimura J (2007) Mechanism of Na+/Ca2+ exchanger activation by hydrogen peroxide in Guinea-pig ventricular myocytes. J Pharmacol Sci 103:283–292CrossRefGoogle Scholar
  18. Iwamoto T, Nakamura TY, Pan Y, Uehara A, Imanaga I, Shigekawa M (1999) Unique topology of the internal repeats in the cardiac Na+-Ca2+ exchanger. FEBS Lett 446:264–268CrossRefGoogle Scholar
  19. Kitao T, Takuma K, Kawasaki T, Inoue Y, Ikehara A, Nashida T, Ago Y, Matsuda T (2010) The Na+/Ca2+ exchanger-mediated Ca2+ influx triggers nitric oxide-induced cytotoxicity in cultured astrocytes. Neurochem Int 57:58–66CrossRefGoogle Scholar
  20. Krenz M, Oldenburg O, Wimpee H, Cohen MV, Garlid KD, Critz SD, Downey JM, Benoit JN (2002) Opening of ATP-sensitive potassium channels causes generation of free radicals in vascular smooth muscle cells. Basic Res Cardiol 97:365–373CrossRefGoogle Scholar
  21. Lebuffe G, Schumacker PT, Shao Z-U, Anderson T, Iwase H, Vanden Hoek TL (2003) ROS and NO trigger early preconditioning: relationship to mitochondrial KATP channel. Am J Physiol Heart Circ Physiol 284:H299–H308CrossRefGoogle Scholar
  22. Liu D, Homan LL, Dillon JS (2004) Genistein acutely stimulates nitric oxide synthesis in vascular endothelial cells by a cyclic adenosine 5′-monophosphate-dependent mechanism. Endocrinology 145:5532–5539CrossRefGoogle Scholar
  23. Mączewski M, Beręsewicz A (1997) Inhibitors of nitric oxide synthesis and ischemia/reperfusion attenuate coronary vasodilator response to pinacidil in isolated rat heart. J Physiol Pharmacol 48:737–749Google Scholar
  24. Murphy ME, Brayden JE (1995) Nitric oxide hyperpolarizes rabbit mesenteric arteries via ATP-sensitive potassium channels. J Physiol 486:47–58CrossRefGoogle Scholar
  25. Newgreen DT, Bray KM, McHarg AD, Weston AH, Duty S, Brown BS, Kay PB, Edwards G, Longmore J, Southerton JS (1990) The action of diazoxide and minoxidil sulphate on rat blood vessels: a comparison with cromakalim. Br J Pharmacol 100:605–613CrossRefGoogle Scholar
  26. Nicoll DA, Ottolia M, Lu L, Lu Y, Philipson KD (1999) A new topological model of the cardiac sarcolemmal Na+-Ca2+ exchanger. J Biol Chem 274:910–917CrossRefGoogle Scholar
  27. Noda Y, Mori A, Packer L (1997) Gliclazide scavenges hydroxyl, superoxide and nitric oxide radicals: an ESR study. Res Commun Mol Pathol Pharmacol 96:115–124Google Scholar
  28. Oldenburg O, Yang XM, Krieg T, Garlid KD, Cohen MV, Grover GJ, Downey JM (2003) P1075 opens mitochondria KATP channels and generates reactive oxygen species resulting in cardioprotection of rabbit hearts. J Mol Cell Cardiol 35:1035–1042CrossRefGoogle Scholar
  29. Reppel M, Fleischmann BK, Reuter H, Sasse P, Schunkert H, Hescheler J (2007) Regulation of the Na+/Ca2+ exchanger (NCX) in the murine embryonic heart. Cardiovasc Res 75:99–108CrossRefGoogle Scholar
  30. Secondo A, Molinaro P, Pannaccione A, Esposito A, Cantile M, Lippiello P, Sirabella R, Iwamoto T, Di Renzo G, Annunziato L (2011) Nitric oxide stimulates NCX1 and NCX2 but inhibits NCX3 isoform by three distinct molecular determinants. Mol Pharmacol 79:558–568CrossRefGoogle Scholar
  31. Southerton JS, Weston AH, Bray KM, Newgreen DT, Taylor SG (1988) The potassium channel opening action of pinacidil: studies using biochemical, ion flux and microelectrode techniques. Naunyn Schmiedeberg Arch Pharmacol 338:310–318CrossRefGoogle Scholar
  32. Spinelli W, Sorota S, Siegal M, Hoffman BF (1991) Antiarrhythmic actions of the ATP-regulated K+ current activated by pinacidil. Circ Res 68:1127–1137CrossRefGoogle Scholar
  33. Wei J, Watanabe Y, Takeuchi K, Yamashita K, Tashiro M, Kita S, Iwamoto T, Watanabe H, Kimura J (2016) Nicorandil stimulates a Na+/Ca2+ exchanger by activating guanylate cyclase in Guinea pig cardiac myocytes. Pflugers Arch 468:693–703Google Scholar
  34. Wu Y, He J-K, Y S-H, Ma S-Y, Huang W, Wei Y-Y, Kong H, Wang H, Zeng X-N, Xie W-P (2017) Activation of ATP-sensitive potassium channels facilitates the function of human endothelial colony-forming cells via Ca2+/Akt/eNOS pathway. J Cell Mol Med 21:609–620CrossRefGoogle Scholar
  35. Xia Y, Dawson VL, Dawson TM, Snyder SH, Zweier JL (1996) Nitric oxide synthase generates nitric oxide and superoxide in arginine-deleted cells leading to peroxynitrite-mediated cellular injury. Proc Natl Acad Sci USA 93:6770–6774CrossRefGoogle Scholar
  36. Xuan Y-T, Tang X-L, Qiu Y, Banerjee S, Takano H, Han H, Bolli R (2000) Biphasic response of cardiac NO synthase isoforms to ischemic preconditioning in conscious rabbits. Am J Physiol Heart Circ Physiol 279:H2360–H2371CrossRefGoogle Scholar
  37. Yamashita K, Watanabe Y, Kita S, Iwamoto T, Kimura J (2016) Inhibitory effect of YM-244769, a novel Na+/Ca2+ exchanger inhibitor on Na+/Ca2+ exchange current in Guinea-pig cardiac ventricular myocytes. Naunyn Schmiedebergs Arch Pharmacol 389:1205–1214CrossRefGoogle Scholar
  38. Zeitz O, Maass AE, Van Nguyen P, Hensmann G, Kogler H, Moller K, Hasenfuss G, Janssen PM (2002) Hydroxyl radical-induced acute diastolic dysfunction is due to calcium overload via reverse-mode Na+/Ca2+ exchange. Circ Res 90:988–995CrossRefGoogle Scholar
  39. Zhang B, Zhang Z, Ji H, Shi H, Chen S, Yan D, Jiang X, Shi B (2015) Grape seed proanthocyanidin extract alleviates urethral dysfunction in diabetic rats through modulating the NO-cGMP pathway. Exp Ther Med 15:1053–1061Google Scholar
  40. Zhang DM, Chai Y, Erickson JR, Brown JH, Bers DM, Lin YF (2014) Intracellular signaling mechanism responsible for modulation of sarcolemmal ATP-sensitive potassium channels by nitric oxide in ventricular cardiomyocytes. J Physiol 592(5):971–990CrossRefGoogle Scholar

Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2019

Authors and Affiliations

  • Keisuke Iguchi
    • 1
    • 2
  • Masao Saotome
    • 1
  • Kanna Yamashita
    • 2
  • Prottoy Hasan
    • 1
  • Miyuki Sasaki
    • 2
  • Yuichiro Maekawa
    • 1
  • Yasuhide Watanabe
    • 2
    Email author
  1. 1.Department of Internal Medicine III (Cardiology)Hamamatsu University School of MedicineHamamatsuJapan
  2. 2.Division of Pharmacological Science, Department of Health ScienceHamamatsu University School of MedicineHamamatsuJapan

Personalised recommendations