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Cell Biochemistry and Biophysics

, Volume 74, Issue 2, pp 263–276 | Cite as

Functional Expression Profile of Voltage-Gated K+ Channel Subunits in Rat Small Mesenteric Arteries

  • Robert H. Cox
  • Samantha Fromme
Original Paper

Abstract

Multiple K v channel complexes contribute to total K v current in numerous cell types and usually subserve different physiological functions. Identifying the complete compliment of functional K v channel subunits in cells is a prerequisite to understanding regulatory function. It was the goal of this work to determine the complete K v subunit compliment that contribute to functional K v currents in rat small mesenteric artery (SMA) myocytes as a prelude to studying channel regulation. Using RNA prepared from freshly dispersed myocytes, high levels of K v 1.2, 1.5, and 2.1 and lower levels of K v 7.4 α-subunit expressions were demonstrated by quantitative PCR and confirmed by Western blotting. Selective inhibitors correolide (K v 1; COR), stromatoxin (K v 2.1; ScTx), and linopirdine (K v 7.4; LINO) decreased K v current at +40 mV in SMA by 46 ± 4, 48 ± 4, and 6.5 ± 2 %, respectively, and K v current in SMA was insensitive to α-dendrotoxin. Contractions of SMA segments pretreated with 100 nmol/L phenylephrine were enhanced by 27 ± 3, 30 ± 8, and 7 ± 3 % of the response to 120 mmol/L KCl by COR, ScTX, and LINO, respectively. The presence of K v 6.1, 9.3, β1.1, and β1.2 was demonstrated by RT-PCR using myocyte RNA with expressions of K vβ1.2 and K v 9.3 about tenfold higher than K vβ1.1 and K v 6.1, respectively. Selective inhibitors of K v 1.3, 3.4, 4.1, and 4.3 channels also found at the RNA and/or protein level had no significant effect on K v current or contraction. These results suggest that K v current in rat SMA myocytes are dominated equally by two major components consisting of K v 1.2–1.5–β1.2 and K v 2.1–9.3 channels along with a smaller contribution from K v 7.4 channels but differences in voltage dependence of activation allows all three to provide significant contributions to SMA function at physiological voltages.

Keywords

Kv subunits Gene expressions Protein expression Smooth muscle cells Toxin inhibitors Kv currents Contractile effects 

References

  1. 1.
    Nelson, M. T., & Quayle, L. M. (1995). Physiological roles and properties of potassium channels in arterial smooth muscle. American Journal of Physiology, 268, C799–C822.PubMedGoogle Scholar
  2. 2.
    Stekiel, W. J. (1989). Electrophysiological mechanisms of force development by vascular smooth muscle membrane in hypertension. In R. M. K. W. Lee (Ed.), Blood Vessel Changes in Hypertension: Structure and Fucntion (Vol. II, pp. 127–170). Boca Raton: CRC Press.Google Scholar
  3. 3.
    Nelson, M. T., Patlak, J. B., Worley, J. F., & Standen, N. B. (1990). Calcium channels, potassium channels, and voltage dependence of arterial smooth muscle tone. American Journal of Physiology, 259, C3–C18.PubMedGoogle Scholar
  4. 4.
    Cox, R. H. (2005). Molecular determinants of voltage gated potassium currents in vascular smooth muscle. Cell Biochemistry and Biophysics, 42, 167–195.CrossRefPubMedGoogle Scholar
  5. 5.
    Li, Y., Um, S. Y., & McDonald, T. V. (2006). Voltage-gated potassium channels: regulation by accessory subunits. Neuroscientist, 12, 199–210.CrossRefPubMedGoogle Scholar
  6. 6.
    Torres, Y. P., Morera, F. J., Carvacho, I., & Latorre, R. (2007). A marriage of convenience: β-subunits and voltage-dependent K + channels. Journal of Biological Chemistry, 282, 24485–24489.CrossRefPubMedGoogle Scholar
  7. 7.
    Coetzee, W. A., Amarillo, Y., Chiu, J., Chow, A., Lau, D., McCormick, T., et al. (1999). Molecular diversity of K+ channels. Annals of the New York Academy of Sciences, 868, 233–285.CrossRefPubMedGoogle Scholar
  8. 8.
    Chandy, K. G., & Gutman, G. A. (1994). Voltage-gated potassium channels. In R. A. North (Ed.), Handbook of receptors and channels. ligand and voltage-gated ion channels (pp. 1–71). Boca Raton: CRC Press.Google Scholar
  9. 9.
    Fergus, D. J., Martens, J. R., & England, S. K. (1998). K v channel subunits that contribute to voltage-gated K+ current in renal vascular smooth muscle. Pflugers Archiv, 445, 697–704.CrossRefGoogle Scholar
  10. 10.
    Yuan, X. J., Wang, J., Juhaszova, M., Golovina, V. A., & Rubin, L. J. (2003). Molecular basis and function of voltage-gated K+ channels in pulmonary arterial smooth muscle cells. American Journal of Physiology, 274, L621–L635.Google Scholar
  11. 11.
    Thorneloe, K. S., Chen, T. T., Grier, E. F., Horowitz, B., Cole, W. C., & Walsh, M. P. (2001). Molecular composition of 4-aminopyridine-sensitive voltage-gated K+ channels of vascular smooth muscle. Circulation Research, 9, 1030–1037.CrossRefGoogle Scholar
  12. 12.
    Cheong, A., Dedman, A. M., Xu, S. Z., & Beech, D. J. (2001). K vα1 channels in murine arterioles: differential cellular expression and regulation of diameter. American Journal of Physiology, 281, H1057–H1065.PubMedGoogle Scholar
  13. 13.
    Xu, C., Lu, Y., Tang, G., & Wang, R. (1999). Expression of voltage-dependent K+ channel genes in mesenteric artery smooth muscle cells. American Journal of Physiology, 277, G1055–G1063.PubMedGoogle Scholar
  14. 14.
    McGahon, M. K., Dawicki, J. M., Arora, A., Simpson, D. A., Gardiner, T. A., Stitt, A. W., et al. (2007). K v 1.5 is a major component underlying the A-type potassium current in retinal arteriolar smooth muscle. American Journal of Physiology, 292, H1001–H1008.PubMedGoogle Scholar
  15. 15.
    Mackie, A. R., Brueggemann, L. I., Henderson, K. K., Shiels, A. J., Cribbs, L. L., Scrogin, K. E., & Byron, K. L. (2008). Vascular KCNQ potassium channels as novel targets for the control of mesenteric artery constriction by vasopressin, based on studies in single cells, pressurized arteries, and in vivo measurements of mesenteric vascular resistance. Journal of Pharmacology and Experimental Therapeutics, 325, 475–483.CrossRefPubMedPubMedCentralGoogle Scholar
  16. 16.
    Ng, F. L., Davis, A. J., Jepps, T. A., Harhun, M. I., Yeung, S. Y., Wan, A., et al. (2011). Expression and function of the K+ channel KCNQ genes in human arteries. British Journal of Pharmacology, 162, 42–53.CrossRefPubMedPubMedCentralGoogle Scholar
  17. 17.
    Lu, Y., Zhang, J., Tang, G., & Wang, R. (2001). Modulation of voltage-dependent K+ channel current in vascular smooth muscle cells from rat mesenteric arteries. The Journal of Membrane Biology, 180, 163–175.CrossRefPubMedGoogle Scholar
  18. 18.
    Albarwani, S., Nemetz, L. T., Madden, J. A., Tobin, A. A., England, S. K., Pratt, P. F., & Rusch, N. J. (2003). Voltage-gated K+ channels in rat small cerebral arteries: molecular identity of the functional channels. Journal of Physiology, 551, 751–763.CrossRefPubMedPubMedCentralGoogle Scholar
  19. 19.
    Amberg, G. C., & Santana, L. F. (2006). K v 2 channels oppose myogenic constriction of rat cerebral arteries. American Journal of Physiology, 291, C348–C356.CrossRefPubMedGoogle Scholar
  20. 20.
    Zhong, X. Z., Abd-Elrahman, K. S., Liao, C. H., El-Yazbi, A. F., Wash, E. J., Walsh, M. P., & Cole, W. C. (2010). Stromatoxin-sensitive, heteromultimeric K v 2.1/K v 9.3 channels contribute to myogenic control of cerebral arterial diameter. Journal of Physiology, 588, 4519–4537.CrossRefPubMedPubMedCentralGoogle Scholar
  21. 21.
    Grissmer, S., Nguyen, A. N., Aiyar, J., Hanson, D. C., Mather, R. J., Gutman, G. A., et al. (1994). Pharmacological characterization of five cloned voltage-gated channels, types K v 1.1, 1.2, 1.3, 1.5 and 3.1, stably expressed in mammalian cell lines. Molecular Pharmacology, 45, 1227–1234.PubMedGoogle Scholar
  22. 22.
    Colinas, O., Gallego, M., Setien, R., Lopez-Lopex, J. M., Perez-Garcia, M. T., & Casis, O. (2006). Differential modulation of K v 4.2 and K v 4.3 channels by calmodulin-dependent protein kinase II in rat cardiac myocytes. American Journal of Physiology, 291, H1978–H1987.PubMedGoogle Scholar
  23. 23.
    Lu, Z., Abe, J., Taunton, J., Lu, Y., Shishido, T., McClain, C., et al. (2008). Reactive oxygen species-induced activation of p90 ribosomal S6 kinase prolongs cardiac repolarization through inhibiting outward K+ channel activity. Circulation Research, 103, 269–278.CrossRefPubMedPubMedCentralGoogle Scholar
  24. 24.
    Bett, G. C., & Rasmusson, R. L. (2008). Modification of K+ channel-drug interactions by ancillary subunits. Journal of Physiology, 586, 929–950.CrossRefPubMedGoogle Scholar
  25. 25.
    Gelband, C. H., & Hume, J. R. (1995). (Ca2 +)i inhibition of K + channels in canine renal artery. Novel mechanism for agonist-induced membrane depolarization. Circulation Research, 77, 121–130.CrossRefPubMedGoogle Scholar
  26. 26.
    Cox, R. H., & Petrou, S. (1999). Ca(2+) influx inhibits voltage-dependent and augments Ca( dependent K(+) currents in arterial myocytes. American Journal of Physiology, 277, C51–C63.PubMedGoogle Scholar
  27. 27.
    Brignell, J. L., Perry, M. D., Nelson, C. P., Willets, J. M., Challiss, R. A., & Davies, N. W. (2015). Steady-state modulation of voltage-gated K + channels in rat arterial smooth muscle by cyclic AMP-dependent protein kinase and protein phosphatase 2B. PLoS One., 10(3), e0121285. doi: 10.1371/journal.pone.0121285.CrossRefPubMedPubMedCentralGoogle Scholar
  28. 28.
    Fishman, J. A., Ryan, G. B., & Karnovsky, M. J. (1975). Endothelial regeneration in the rat carotid artery and the significance of endothelial denudation in the pathogenesis of myointimal thickening. Laboratory Investigation, 32, 339–351.PubMedGoogle Scholar
  29. 29.
    Cox, R. H., Folander, K., & Swanson, R. (2001). Differential expression of voltage-gated K + channel genes in arteries from spontaneously hypertensive and Wistar-Kyoto rats. Hypertension, 37, 1315–1322.CrossRefPubMedGoogle Scholar
  30. 30.
    Cox, R. H., Fromme, S., Folander, K., & Swanson, R. (2008). Voltage gated K+ channel expression in arteries of Wistar Kyoto and spontaneously hypertensive rats. American Journal of Hypertension, 2008(21), 213–218.CrossRefGoogle Scholar
  31. 31.
    Livak, K. J., & Schmittgen, T. D. (2001). Analysis of relative gene expression data using real-time quantitative PCR and the 2−∆∆Ct method. Methods, 25, 402–408.CrossRefPubMedGoogle Scholar
  32. 32.
    Cox, R. H., Haas, K. S., Moisey, D. M., & Tulenko, T. N. (1989). Effects of endothelium regeneration on canine coronary artery function. American Journal of Physiology, 257, H1681–H1692.PubMedGoogle Scholar
  33. 33.
    Felix, J. P., Bugianesi, R. M., Schmalhofer, W. A., Borris, R., Goetz, M. A., Hensens, O. D., et al. (1999). Identification and biochemical characterization of a novel nortriterpene inhibitor of the human lymphocyte voltage-gated potassium channel, K v 1.3. Biochemistry, 38, 4922–4930.CrossRefPubMedGoogle Scholar
  34. 34.
    Russell, S. N., Overturf, K. E., & Horowitz, B. (1994). Heterotetramer formation and charybdotoxin sensitivity of two K + channels cloned from smooth muscle. American Journal of Physiology, 267, C1729–C1733.PubMedGoogle Scholar
  35. 35.
    Escoubas, P., Diochot, S., Célérier, M. L., Nakajima, T., & Lazdunski, M. (2002). Novel tarantula toxins for subtypes of voltage-dependent potassium channels in the K v 2 and K v 4 subfamilies. Molecular Pharmacology, 62, 48–57.CrossRefPubMedGoogle Scholar
  36. 36.
    Patel, A. J., Lazdubski, M., & Honore, E. (1997). K v 2.1/K v 9.3, a novel ATP-dependent delayed-rectifier K+ channel in oxygen-sensitive pulmonary artery myocytes. EMBO Journal, 6, 6615–6625.CrossRefGoogle Scholar
  37. 37.
    Kramer, J. W., Post, M. A., Brown, A. M., & Kirsch, G. E. (1998). Modulation of potassium channel gating by coexpression of K v 2.1 with regulatory K v 5.1 or K v 6.1 alpha-subunits. American Journal of Physiology, 274, C1501–C1510.PubMedGoogle Scholar
  38. 38.
    Amberg, G. C., Koh, S. D., Imaizumi, Y., Ohya, S., & Sanders, K. M. (2003). A-type potassium currents in smooth muscle. American Journal of Physiology, 284, C583–C595.CrossRefPubMedGoogle Scholar
  39. 39.
    Cartwright, T. A., Corey, M. J., & Schwalbe, R. A. (2007). Complex oligosaccharides are N-linked in K v 3 voltage-gated K+ channels in brain. Biochimica et Biophysica Acta, 1770, 666–671.CrossRefPubMedGoogle Scholar
  40. 40.
    Diochot, S., Schweitz, H., Béress, L., & Lazdunski, M. (1998). Sea anemone peptides with a specific blocking activity against the fast inactivating potassium channel K v 3.4. Journal of Biological Chemistry, 273, 6744–6749.CrossRefPubMedGoogle Scholar
  41. 41.
    Birnbaum, S. G., Varga, A. W., Yuan, L. L., Anderson, A. E., Sweatt, J. D., & Schrader, L. A. (2004). Structure and function of K v 4-family transient potassium channels. Physiological Reviews, 84, 803–833.CrossRefPubMedGoogle Scholar
  42. 42.
    Zhong, X. Z., Harhun, M. I., Olesen, S. P., Phya, S., Moffatt, J. D., Cole, W. C., & Greenwood, I. A. (2010). Participation of KCNQ (K v 7) potassium channels in myogenic control of cerebral artery diameter. Journal of Physiology, 588, 3277–3293.CrossRefPubMedPubMedCentralGoogle Scholar
  43. 43.
    Cox RH, Fromme S. (2015) Comparison of voltage gated K+ currents in arterial myocytes with K v subunits expressed in HEK293 cells. 2015. Submitted.Google Scholar
  44. 44.
    Misonou, H., & Trimmer, J. S. (2004). Determinants of voltage-gated potassium channel surface expression and localization in mammalian neurons. Critical Reviews in Biochemistry and Molecular Biology, 39, 125–145.CrossRefPubMedGoogle Scholar
  45. 45.
    Bocksteins, E., & Snyders, D. J. (2012). Electrically silent K v subunits: Their molecular and functional characteristics. Physiology, 27, 73–84.CrossRefPubMedGoogle Scholar
  46. 46.
    Yeung, S. Y., Pucovský, V., Moffatt, J. D., Saldanha, L., Schwake, M., Ohya, S., & Greenwood, I. A. (2007). Molecular expression and pharmacological identification of a role for K(v)7 channels in murine vascular reactivity. British Journal of Pharmacology, 151, 758–770.CrossRefPubMedPubMedCentralGoogle Scholar
  47. 47.
    Miceli, F., Cilio, M. R., Taglialatela, M., & Bezanilla, F. (2009). Gating currents from neuronal K(V)7.4 channels: general features and correlation with the ionic conductance. Channels (Austin), 3, 274–2783.CrossRefGoogle Scholar
  48. 48.
    Plane, F., Johnson, R., Kerr, P., Wiehler, W., Thorneloe, K., Ishii, K., et al. (2005). Heteromultimeric K v 1 channels contribute to myogenic control of arterial diameter. Circulation Research, 96, 216–224.CrossRefPubMedGoogle Scholar
  49. 49.
    Nerbonne, J. M. (1998). Regulation of voltage-gated K+ channel expression in the developing mammalian myocardium. Journal of Neurobiology, 37, 37–59.CrossRefPubMedGoogle Scholar
  50. 50.
    Schmitt, N., Grunnet, M., & Olesen, S.-P. (2014). Cardiac potassium channel subtypes: new roles in repolarization and arrhythmia. Physiological Reviews, 94, 609–653.CrossRefPubMedGoogle Scholar
  51. 51.
    Schulz, D. J., Temporal, S., Barry, D. M., & Garcia, M. L. (2008). Mechanisms of voltage-gated ion channel regulation: from gene expression to localization. Cellular and Molecular Life Sciences, 65, 2215–2231.CrossRefPubMedGoogle Scholar
  52. 52.
    Núñez, L., Vaquero, M., Gómez, R., Caballero, R., Mateos-Cáceres, P., Macaya, C., et al. (2006). Nitric oxide blocks hKv1.5 channels by S-nitrosylation and by a cyclic GMP-dependent mechanism. Cardiovascular Research, 72, 80–89.CrossRefPubMedGoogle Scholar
  53. 53.
    Watanabe, I., Zhu, J., Sutachan, J. J., Gottschalk, A., Recio-Pinto, E., & Thornhill, W. B. (2007). The glycosylation state of K v 1.2 potassium channels affects trafficking, gating, and simulated action potentials. Brain Research, 1144, 1–18.CrossRefPubMedGoogle Scholar
  54. 54.
    Wang, Z., Kiehn, J., Yang, Q., Brown, A. M., & Wible, B. A. (1996). Comparison of binding and block produced by alternatively spliced K vβ1 subunits. Journal of Biological Chemistry, 271, 28311–28317.CrossRefPubMedGoogle Scholar
  55. 55.
    Schleifenbaum, J., Köhn, C., Voblova, N., Dubrovska, G., Zavarirskaya, O., Gloe, T., et al. (2010). Systemic peripheral artery relaxation by KCNQ channel openers and hydrogen sulfide. Journal of Hypertension, 28, 1875–1882.CrossRefPubMedGoogle Scholar
  56. 56.
    Yuan, X. J., Goldman, W. F., Tod, M. L., Rubuin, L. J., & Blaustein, M. P. (1993). Ionic currents in rat pulmonary and mesenteric arterial myocytes in primary culture and subculture. American Journal of Physiology, 264, L107–L115.PubMedGoogle Scholar
  57. 57.
    Cox, R. H. (1979). Contribution of smooth muscle to arterial wall mechanics. Basic Research in Cardiology, 74, 1–9.CrossRefPubMedGoogle Scholar
  58. 58.
    Strutz-Seebohm, N., Seebohm, G., Fedorenko, O., Baltaev, R., Engel, J., Knirsch, M., & Lang, F. (2006). Functional coassembly of KCNQ4 with KCNE-beta- subunits in Xenopus oocytes. Cellular Physiology and Biochemistry, 18, 57–66.CrossRefPubMedGoogle Scholar

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© Springer Science+Business Media New York 2015

Authors and Affiliations

  1. 1.Program in Cardiovascular DiseaseLankenau Institute for Medical Research, Main Line Health SystemWynnewoodUSA

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