The Korean Journal of Physiology and Pharmacology

The Korean Society of Pharmacology (대한약리학회)
권호 표지
DOI QR코드

DOI QR Code

Acute Hypoxia Activates an ENaC-like Channel in Rat Pheochromocytoma (PC12) Cells

  • Bae, Yeon Ju (Department of Physiology, Institute of Health Sciences and Medical Research Center for Neural Dysfunction, Gyeongsang National University School of Medicine) ;
  • Yoo, Jae-Cheal (Department of Physiology, Institute of Health Sciences and Medical Research Center for Neural Dysfunction, Gyeongsang National University School of Medicine) ;
  • Park, Nammi (Department of Physiology, Institute of Health Sciences and Medical Research Center for Neural Dysfunction, Gyeongsang National University School of Medicine) ;
  • Kang, Dawon (Department of Physiology, Institute of Health Sciences and Medical Research Center for Neural Dysfunction, Gyeongsang National University School of Medicine) ;
  • Han, Jaehee (Department of Physiology, Institute of Health Sciences and Medical Research Center for Neural Dysfunction, Gyeongsang National University School of Medicine) ;
  • Hwang, Eunmi (Department of Physiology, Institute of Health Sciences and Medical Research Center for Neural Dysfunction, Gyeongsang National University School of Medicine) ;
  • Park, Jae-Yong (Department of Physiology, Institute of Health Sciences and Medical Research Center for Neural Dysfunction, Gyeongsang National University School of Medicine) ;
  • Hong, Seong-Geun (Department of Physiology, Institute of Health Sciences and Medical Research Center for Neural Dysfunction, Gyeongsang National University School of Medicine)
  • Received : 2012.10.31
  • Accepted : 2012.12.04
  • Published : 2013.02.28
Download PDF
⟨ Previous Next ⟩

Abstract

Cells can resist and even recover from stress induced by acute hypoxia, whereas chronic hypoxia often leads to irreversible damage and eventually death. Although little is known about the response(s) to acute hypoxia in neuronal cells, alterations in ion channel activity could be preferential. This study aimed to elucidate which channel type is involved in the response to acute hypoxia in rat pheochromocytomal (PC12) cells as a neuronal cell model. Using perfusing solution saturated with 95% $N_2$ and 5% $CO_2$, induction of cell hypoxia was confirmed based on increased intracellular $Ca^{2+}$ with diminished oxygen content in the perfusate. During acute hypoxia, one channel type with a conductance of about 30 pS (2.5 pA at -80 mV) was activated within the first 2~3 min following onset of hypoxia and was long-lived for more than 300 ms with high open probability ($P_o$, up to 0.8). This channel was permeable to $Na^+$ ions, but not to $K^+$, $Ca^+$, and $Cl^-$ ions, and was sensitively blocked by amiloride (200 nM). These characteristics and behaviors were quite similar to those of epithelial sodium channel (ENaC). RT-PCR and Western blot analyses confirmed that ENaC channel was endogenously expressed in PC12 cells. Taken together, a 30-pS ENaC-like channel was activated in response to acute hypoxia in PC12 cells. This is the first evidence of an acute hypoxia-activated $Na^+$ channel that can contribute to depolarization of the cell.

Keywords

References

  1. Cross JL, Meloni BP, Bakker AJ, Lee S, Knuckey NW. Modes of neuronal calcium entry and homeostasis following cerebral ischemia. Stroke Res Treat. 2010;2010:316862.
  2. Fung ML. Role of voltage-gated $Na^{+}$ channels in hypoxiainduced neuronal injuries. Clin Exp Pharmacol Physiol. 2000; 27:569-574. https://doi.org/10.1046/j.1440-1681.2000.03309.x
  3. Ratan RR, Siddiq A, Smirnova N, Karpisheva K, Haskew- Layton R, McConoughey S, Langley B, Estevez A, Huerta PT, Volpe B, Roy S, Sen CK, Gazaryan I, Cho S, Fink M, LaManna J. Harnessing hypoxic adaptation to prevent, treat, and repair stroke. J Mol Med (Berl). 2007;85:1331-1338. https://doi.org/10.1007/s00109-007-0283-1
  4. Conrad PW, Conforti L, Kobayashi S, Beitner-Johnson D, Rust RT, Yuan Y, Kim HW, Kim RH, Seta K, Millhorn DE. The molecular basis of $O_{2}$-sensing and hypoxia tolerance in pheochromocytoma cells. Comp Biochem Physiol B Biochem Mol Biol. 2001;128:187-204. https://doi.org/10.1016/S1096-4959(00)00326-2
  5. Conforti L, Millhorn DE. Selective inhibition of a slowinactivating voltage-dependent $K^{+}$ channel in rat PC12 cells by hypoxia. J Physiol. 1997;502:293-305. https://doi.org/10.1111/j.1469-7793.1997.293bk.x
  6. Zhu WH, Conforti L, Czyzyk-Krzeska MF, Millhorn DE. Membrane depolarization in PC-12 cells during hypoxia is regulated by an $O_{2}$-sensitive $K^{+}$ current. Am J Physiol. 1996; 271:C658-665. https://doi.org/10.1152/ajpcell.1996.271.2.C658
  7. Conforti L, Bodi I, Nisbet JW, Millhorn DE. $O_{2}$-sensitive $K^{+}$ channels: role of the Kv1.2 -subunit in mediating the hypoxic response. J Physiol. 2000;524:783-793. https://doi.org/10.1111/j.1469-7793.2000.00783.x
  8. Kim D. $K^{+}$ channels in $O_{2}$ sensing and postnatal development of carotid body glomus cell response to hypoxia. Respir Physiol Neurobiol. 2013;185:44-56. https://doi.org/10.1016/j.resp.2012.07.005
  9. Lopez-Lopez JR, Gonzalez C, Urena J, Lopez-Barneo J. Low p$O_{2}$ selectively inhibits K channel activity in chemoreceptor cells of the mammalian carotid body. J Gen Physiol. 1989;93: 1001-1015. https://doi.org/10.1085/jgp.93.5.1001
  10. Lopez-Lopez JR, Gonzalez C, Perez-Garcia MT. Properties of ionic currents from isolated adult rat carotid body chemoreceptor cells: effect of hypoxia. J Physiol. 1997;499:429-441. https://doi.org/10.1113/jphysiol.1997.sp021939
  11. Ji HL, Benos DJ. Degenerin sites mediate proton activation of deltabetagamma-epithelial sodium channel. J Biol Chem. 2004; 279:26939-26947. https://doi.org/10.1074/jbc.M401143200
  12. Wesch D, Miranda P, Afonso-Oramas D, Althaus M, Castro- Hernandez J, Dominguez J, Morty RE, Clauss W, Gonzalez- Hernandez T, Alvarez de la Rosa D, Giraldez T. The neuronalneuronal- specific SGK1.1 kinase regulates $\delta$ -epithelial $Na^{+}$ channel independently of PY motifs and couples it to phospholipase C signaling. Am J Physiol Cell Physiol. 2010;299: C779-790. https://doi.org/10.1152/ajpcell.00184.2010
  13. Seta KA, Spicer Z, Yuan Y, Lu G, Millhorn DE. Responding to hypoxia: lessons from a model cell line. Sci STKE. 2002; 2002:re11.
  14. Abe E, Fujiki M, Nagai Y, Shiqi K, Kubo T, Ishii K, Abe T, Kobayashi H. The phosphatidylinositol-3 kinase/Akt pathway mediates geranylgeranylacetone-induced neuroprotection against cerebral infarction in rats. Brain Res. 2010;1330:151-157. https://doi.org/10.1016/j.brainres.2010.02.074
  15. Tong Q, Gamper N, Medina JL, Shapiro MS, Stockand JD. Direct activation of the epithelial $Na^{+}$ channel by phosphatidylinositol 3,4,5-trisphosphate and phosphatidylinositol 3,4- bisphosphate produced by phosphoinositide 3-OH kinase. J Biol Chem. 2004;279:22654-22663. https://doi.org/10.1074/jbc.M401004200
  16. Chen Y, Shi G, Xia W, Kong C, Zhao S, Gaw AF, Chen EY, Yang GP, Giaccia AJ, Le QT, Koong AC. Identification of hypoxia-regulated proteins in head and neck cancer by proteomic and tissue array profiling. Cancer Res. 2004;64: 7302-7310. https://doi.org/10.1158/0008-5472.CAN-04-0899
  17. Lebowitz J, Edinger RS, An B, Perry CJ, Onate S, Kleyman TR, Johnson JP. Ikappab kinase-beta (ikkbeta) modulation of epithelial sodium channel activity. J Biol Chem. 2004;279: 41985-41990. https://doi.org/10.1074/jbc.M403923200
  18. Choi SW, Ahn JS, Kim HK, Kim N, Choi TH, Park SW, Ko EA, Park WS, Song DK, Han J. Increased expression of atp-sensitive $K^{+}$ channels improves the right ventricular tolerance to hypoxia in rabbit hearts. Korean J Physiol Pharmacol. 2011;15:189-194. https://doi.org/10.4196/kjpp.2011.15.4.189
  19. Wood JN, Boorman JP, Okuse K, Baker MD. Voltage-gated sodium channels and pain pathways. J Neurobiol. 2004;61:55-71. https://doi.org/10.1002/neu.20094
  20. Catterall WA, Goldin AL, Waxman SG. International Union of Pharmacology. XLVII. Nomenclature and structure-function relationships of voltage-gated sodium channels. Pharmacol Rev. 2005;57:397-409. https://doi.org/10.1124/pr.57.4.4
  21. Kellenberger S, Schild L. Epithelial sodium channel/degenerin family of ion channels: a variety of functions for a shared structure. Physiol Rev. 2002;82:735-767. https://doi.org/10.1152/physrev.00007.2002
  22. Hammarström AK, Gage PW. Hypoxia and persistent sodium current. Eur Biophys J. 2002;31:323-330. https://doi.org/10.1007/s00249-002-0218-2
  23. Waldmann R, Champigny G, Bassilana F, Heurteaux C, Lazdunski M. A proton-gated cation channel involved in acidsensing. Nature. 1997;386:173-177. https://doi.org/10.1038/386173a0
  24. Alexander SPH, Mathie A, Peters JA. Guide to receptors and Channels (GRAC), 5th edition. Br J Pharmacol. 2011;164 Suppl 1:S1-324. https://doi.org/10.1111/j.1476-5381.2011.01649_1.x
  25. Ohbuchi T, Sato K, Suzuki H, Okada Y, Dayanithi G, Murphy D, Ueta Y. Acid-sensing ion channels in rat hypothalamic vasopressin neurons of the supraoptic nucleus. J Physiol. 2010; 588:2147-2162. https://doi.org/10.1113/jphysiol.2010.187625
  26. Chu XP, Miesch J, Johnson M, Root L, Zhu XM, Chen D, Simon RP, Xiong ZG. Proton-gated channels in PC12 cells. J Neurophysiol. 2002;87:2555-2561. https://doi.org/10.1152/jn.00741.2001
  27. Garty H, Palmer LG. Epithelial sodium channels: function, structure, and regulation. Physiol Rev. 1997;77:359-396. https://doi.org/10.1152/physrev.1997.77.2.359
  28. Kellenberger S, Hoffmann-Pochon N, Gautschi I, Schneeberger E, Schild L. On the molecular basis of ion permeation in the epithelial $Na^{+}$ channel. J Gen Physiol. 1999;114:13-30. https://doi.org/10.1085/jgp.114.1.13
  29. Anantharam A, Palmer LG. Determination of epithelial $Na^{+}$ channel subunit stoichiometry from single-channel conductances. J Gen Physiol. 2007;130:55-70. https://doi.org/10.1085/jgp.200609716
  30. Ismailov II, Berdiev BK, Bradford AL, Awayda MS, Fuller CM, Benos DJ. Associated proteins and renal epithelial $Na^{+}$ channel function. J Membr Biol. 1996;149:123-132. https://doi.org/10.1007/s002329900013
  31. Kelly O, Lin C, Ramkumar M, Saxena NC, Kleyman TR, Eaton DC. Characterization of an amiloride binding region in the alpha-subunit of ENaC. Am J Physiol Renal Physiol. 2003;285: F1279-1290. https://doi.org/10.1152/ajprenal.00094.2003
  32. Ma HP, Al-Khalili O, Ramosevac S, Saxena S, Liang YY, Warnock DG, Eaton DC. Steroids and exogenous γ-ENaC subunit modulate cation channels formed by $\alpha$ -ENaC in human B lymphocytes. J Biol Chem. 2004;279:33206-33212. https://doi.org/10.1074/jbc.M405455200
  33. Ma HP, Chou CF, Wei SP, Eaton DC. Regulation of the epithelial sodium channel by phosphatidylinositides: experiments, implications, and speculations. Pflugers Arch. 2007;455: 169-180. https://doi.org/10.1007/s00424-007-0294-3
  34. Yamashima T, Saido TC, Takita M, Miyazawa A, Yamano J, Miyakawa A, Nishijyo H, Yamashita J, Kawashima S, Ono T, Yoshioka T. Transient brain ischaemia provokes $Ca^{2+}$, PIP2 and calpain responses prior to delayed neuronal death in monkeys. Eur J Neurosci. 1996;8:1932-1944. https://doi.org/10.1111/j.1460-9568.1996.tb01337.x
  35. Donaldson SH, Hirsh A, Li DC, Holloway G, Chao J, Boucher RC, Gabriel SE. Regulation of the epithelial sodium channel by serine proteases in human airways. J Biol Chem. 2002;277: 8338-8345. https://doi.org/10.1074/jbc.M105044200

Cited by

  1. Hypoxia Induces Internalization of κ-Opioid Receptor vol.126, pp.5, 2017, https://doi.org/10.1097/aln.0000000000001571