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. 1993 Nov;65(5):2002–2012. doi: 10.1016/S0006-3495(93)81242-6

Gap junction gating sensitivity to physiological internal calcium regardless of pH in Novikoff hepatoma cells.

A Lazrak 1, C Peracchia 1
PMCID: PMC1225936  PMID: 8298030

Abstract

Gap junction conductance (Gj) and channel gating sensitivity to voltage, Ca2+, H+, and heptanol were studied by double whole-cell clamp in Novikoff hepatoma cell pairs. Channel gating was observed at transjunctional voltages (Vj) > +/- 50 mV. The cells readily uncoupled with 1 mM 1-heptanol. With heptanol, single (gap junctional) channel events with unitary conductances (gamma j) of 46 and 97 pS were detected. Both Ca(2+)-loading (EGTA.Ca) and acidifying (100% CO2) solutions caused uncoupling. However, CO2 was effective when Ca2+i was buffered with EGTA (a H(+)-sensitive Ca-buffer) but not with BAPTA (1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid) (a H(+)-insensitive Ca-buffer), suggesting a Ca(2+)-mediated H+ effect on gap junctions. This was tested by monitoring the Gj decay at different pCai values (9, 6.9, 6.3, 6, and 5.5; 1 mM BAPTA) and pHi values (7.2 or 6.1, 10 mM 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid and 2-(N-morpholino)ethansulphonic acid, respectively). With pCai > or = 6.9 (pH 7.2 or 6.1), Gj decreased to 10-70% of initial values in approximately 40 min, following single exponential decays (tau = approximately 28 min). With pCai 6-6.3 (pH 7.2 or 6.1), Gj decreased to 10-25% of initial values in 15 min (tau = approximately 5 min); the Student t gave a P = 0.0178. With pCa 5.5 the cells uncoupled in less than 1 min (tau = approximately 20 s). Low pHi affected neither time course nor shape of Gj decay at any pCai tested. The data indicate that these gap junctions are sensitive to [Ca2+]i in the physiological range (< or = 500 nM) and that low pHi, without an increase in [Ca2+]i, neither decreases Gj nor increases channel sensitivity to Ca2+.

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Selected References

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  1. Adler E. M., Augustine G. J., Duffy S. N., Charlton M. P. Alien intracellular calcium chelators attenuate neurotransmitter release at the squid giant synapse. J Neurosci. 1991 Jun;11(6):1496–1507. doi: 10.1523/JNEUROSCI.11-06-01496.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Anumonwo J. M., Wang H. Z., Trabka-Janik E., Dunham B., Veenstra R. D., Delmar M., Jalife J. Gap junctional channels in adult mammalian sinus nodal cells. Immunolocalization and electrophysiology. Circ Res. 1992 Aug;71(2):229–239. doi: 10.1161/01.res.71.2.229. [DOI] [PubMed] [Google Scholar]
  3. Arellano R. O., Ramón F., Rivera A., Zampighi G. A. Lowering of pH does not directly affect the junctional resistance of crayfish lateral axons. J Membr Biol. 1986;94(3):293–299. doi: 10.1007/BF01869725. [DOI] [PubMed] [Google Scholar]
  4. Beyer E. C., Paul D. L., Goodenough D. A. Connexin43: a protein from rat heart homologous to a gap junction protein from liver. J Cell Biol. 1987 Dec;105(6 Pt 1):2621–2629. doi: 10.1083/jcb.105.6.2621. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Burt J. M. Block of intercellular communication: interaction of intracellular H+ and Ca2+. Am J Physiol. 1987 Oct;253(4 Pt 1):C607–C612. doi: 10.1152/ajpcell.1987.253.4.C607. [DOI] [PubMed] [Google Scholar]
  6. Burt J. M., Spray D. C. Single-channel events and gating behavior of the cardiac gap junction channel. Proc Natl Acad Sci U S A. 1988 May;85(10):3431–3434. doi: 10.1073/pnas.85.10.3431. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Cooper K., Rae J. L., Gates P. Membrane and junctional properties of dissociated frog lens epithelial cells. J Membr Biol. 1989 Nov;111(3):215–227. doi: 10.1007/BF01871007. [DOI] [PubMed] [Google Scholar]
  8. Dahl G., Isenberg G. Decoupling of heart muscle cells: correlation with increased cytoplasmic calcium activity and with changes of nexus ultrastructure. J Membr Biol. 1980 Mar 31;53(1):63–75. doi: 10.1007/BF01871173. [DOI] [PubMed] [Google Scholar]
  9. De Mello W. C. Effect of intracellular injection of calcium and strontium on cell communication in heart. J Physiol. 1975 Sep;250(2):231–245. doi: 10.1113/jphysiol.1975.sp011051. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. De Mello W. C. The influence of pH on the healing-over of mammalian cardiac muscle. J Physiol. 1983 Jun;339:299–307. doi: 10.1113/jphysiol.1983.sp014717. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Délèze J. Cell-to-cell communication in the heart: structure-function correlations. Experientia. 1987 Oct 15;43(10):1068–1075. doi: 10.1007/BF01956041. [DOI] [PubMed] [Google Scholar]
  12. Délèze J., Hervé J. C. Effect of several uncouplers of cell-to-cell communication on gap junction morphology in mammalian heart. J Membr Biol. 1983;74(3):203–215. doi: 10.1007/BF02332124. [DOI] [PubMed] [Google Scholar]
  13. Délèze J., Loewenstein W. R. Permeability of a cell junction during intracellular injection of divalent cations. J Membr Biol. 1976 Aug 27;28(1):71–86. doi: 10.1007/BF01869691. [DOI] [PubMed] [Google Scholar]
  14. Délèze J. The recovery of resting potential and input resistance in sheep heart injured by knife or laser. J Physiol. 1970 Jul;208(3):547–562. doi: 10.1113/jphysiol.1970.sp009136. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Fenwick E. M., Marty A., Neher E. A patch-clamp study of bovine chromaffin cells and of their sensitivity to acetylcholine. J Physiol. 1982 Oct;331:577–597. doi: 10.1113/jphysiol.1982.sp014393. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Harris A. L., Spray D. C., Bennett M. V. Kinetic properties of a voltage-dependent junctional conductance. J Gen Physiol. 1981 Jan;77(1):95–117. doi: 10.1085/jgp.77.1.95. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Johnson R., Hammer M., Sheridan J., Revel J. P. Gap junction formation between reaggregated Novikoff hepatoma cells. Proc Natl Acad Sci U S A. 1974 Nov;71(11):4536–4540. doi: 10.1073/pnas.71.11.4536. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Johnston M. F., Ramón F. Electrotonic coupling in internally perfused crayfish segmented axons. J Physiol. 1981 Aug;317:509–518. doi: 10.1113/jphysiol.1981.sp013840. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Lal R., Arnsdorf M. F. Voltage-dependent gating and single-channel conductance of adult mammalian atrial gap junctions. Circ Res. 1992 Sep;71(3):737–743. doi: 10.1161/01.res.71.3.737. [DOI] [PubMed] [Google Scholar]
  20. Loewenstein W. R. Junctional intercellular communication: the cell-to-cell membrane channel. Physiol Rev. 1981 Oct;61(4):829–913. doi: 10.1152/physrev.1981.61.4.829. [DOI] [PubMed] [Google Scholar]
  21. Loewenstein W. R. Permeability of membrane junctions. Ann N Y Acad Sci. 1966 Jul 14;137(2):441–472. doi: 10.1111/j.1749-6632.1966.tb50175.x. [DOI] [PubMed] [Google Scholar]
  22. Marty A., Neher E. Potassium channels in cultured bovine adrenal chromaffin cells. J Physiol. 1985 Oct;367:117–141. doi: 10.1113/jphysiol.1985.sp015817. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Maurer P., Weingart R. Cell pairs isolated from adult guinea pig and rat hearts: effects of [Ca2+]i on nexal membrane resistance. Pflugers Arch. 1987 Aug;409(4-5):394–402. doi: 10.1007/BF00583793. [DOI] [PubMed] [Google Scholar]
  24. Meyer R. A., Laird D. W., Revel J. P., Johnson R. G. Inhibition of gap junction and adherens junction assembly by connexin and A-CAM antibodies. J Cell Biol. 1992 Oct;119(1):179–189. doi: 10.1083/jcb.119.1.179. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Meyer R. A., Lampe P. D., Malewicz B., Baumann W. J., Johnson R. G. Enhanced gap junction formation with LDL and apolipoprotein B. Exp Cell Res. 1991 Sep;196(1):72–81. doi: 10.1016/0014-4827(91)90457-6. [DOI] [PubMed] [Google Scholar]
  26. Moreno A. P., Fishman G. I., Spray D. C. Phosphorylation shifts unitary conductance and modifies voltage dependent kinetics of human connexin43 gap junction channels. Biophys J. 1992 Apr;62(1):51–53. doi: 10.1016/S0006-3495(92)81775-7. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Mullins L. J., Brinley F. J., Jr Sensitivity of calcium efflux from squid axons to changes in membrane potential. J Gen Physiol. 1975 Feb;65(2):135–152. doi: 10.1085/jgp.65.2.135. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. NOVIKOFF A. B. A transplantable rat liver tumor induced by 4-dimethylaminoazobenzene. Cancer Res. 1957 Nov;17(10):1010–1027. [PubMed] [Google Scholar]
  29. Neyton J., Trautmann A. Acetylcholine modulation of the conductance of intercellular junctions between rat lacrimal cells. J Physiol. 1986 Aug;377:283–295. doi: 10.1113/jphysiol.1986.sp016187. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Neyton J., Trautmann A. Single-channel currents of an intercellular junction. 1985 Sep 26-Oct 2Nature. 317(6035):331–335. doi: 10.1038/317331a0. [DOI] [PubMed] [Google Scholar]
  31. Noma A., Tsuboi N. Dependence of junctional conductance on proton, calcium and magnesium ions in cardiac paired cells of guinea-pig. J Physiol. 1987 Jan;382:193–211. doi: 10.1113/jphysiol.1987.sp016363. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Obaid A. L., Socolar S. J., Rose B. Cell-to-cell channels with two independently regulated gates in series: analysis of junctional conductance modulation by membrane potential, calcium, and pH. J Membr Biol. 1983;73(1):69–89. doi: 10.1007/BF01870342. [DOI] [PubMed] [Google Scholar]
  33. Peracchia C. Effects of caffeine and ryanodine on low pHi-induced changes in gap junction conductance and calcium concentration in crayfish septate axons. J Membr Biol. 1990 Jul;117(1):79–89. doi: 10.1007/BF01871567. [DOI] [PubMed] [Google Scholar]
  34. Peracchia C. Increase in gap junction resistance with acidification in crayfish septate axons is closely related to changes in intracellular calcium but not hydrogen ion concentration. J Membr Biol. 1990 Jan;113(1):75–92. doi: 10.1007/BF01869608. [DOI] [PubMed] [Google Scholar]
  35. Peracchia C. Structural correlates of gap junction permeation. Int Rev Cytol. 1980;66:81–146. doi: 10.1016/s0074-7696(08)61972-5. [DOI] [PubMed] [Google Scholar]
  36. Plagemann P. G., Swim H. E. Replication of mengovirus. I. Effect on synthesis of macromolecules by host cell. J Bacteriol. 1966 Jun;91(6):2317–2326. doi: 10.1128/jb.91.6.2317-2326.1966. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Pressler M. L. Intracellular pH and cell-to-cell transmission in sheep Purkinje fibers. Biophys J. 1989 Jan;55(1):53–65. doi: 10.1016/S0006-3495(89)82780-8. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Pusch M., Neher E. Rates of diffusional exchange between small cells and a measuring patch pipette. Pflugers Arch. 1988 Feb;411(2):204–211. doi: 10.1007/BF00582316. [DOI] [PubMed] [Google Scholar]
  39. Ramón F., Rivera A. Gap junction channel modulation--a physiological viewpoint. Prog Biophys Mol Biol. 1986;48(3):127–153. doi: 10.1016/0079-6107(86)90010-6. [DOI] [PubMed] [Google Scholar]
  40. Reber W. R., Weingart R. Ungulate cardiac purkinje fibres: the influence of intracellular pH on the electrical cell-to-cell coupling. J Physiol. 1982 Jul;328:87–104. doi: 10.1113/jphysiol.1982.sp014254. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Rook M. B., Jongsma H. J., van Ginneken A. C. Properties of single gap junctional channels between isolated neonatal rat heart cells. Am J Physiol. 1988 Oct;255(4 Pt 2):H770–H782. doi: 10.1152/ajpheart.1988.255.4.H770. [DOI] [PubMed] [Google Scholar]
  42. Rose B., Loewenstein W. R. Permeability of a cell junction and the local cytoplasmic free ionized calcium concentration: a study with aequorin. J Membr Biol. 1976 Aug 27;28(1):87–119. doi: 10.1007/BF01869692. [DOI] [PubMed] [Google Scholar]
  43. Rose B., Rick R. Intracellular pH, intracellular free Ca, and junctional cell-cell coupling. J Membr Biol. 1978 Dec 29;44(3-4):377–415. doi: 10.1007/BF01944230. [DOI] [PubMed] [Google Scholar]
  44. Rüdisüli A., Weingart R. Electrical properties of gap junction channels in guinea-pig ventricular cell pairs revealed by exposure to heptanol. Pflugers Arch. 1989 Oct;415(1):12–21. doi: 10.1007/BF00373136. [DOI] [PubMed] [Google Scholar]
  45. Spray D. C., Bennett M. V. Physiology and pharmacology of gap junctions. Annu Rev Physiol. 1985;47:281–303. doi: 10.1146/annurev.ph.47.030185.001433. [DOI] [PubMed] [Google Scholar]
  46. Spray D. C., Harris A. L., Bennett M. V. Equilibrium properties of a voltage-dependent junctional conductance. J Gen Physiol. 1981 Jan;77(1):77–93. doi: 10.1085/jgp.77.1.77. [DOI] [PMC free article] [PubMed] [Google Scholar]
  47. Spray D. C., Harris A. L., Bennett M. V. Gap junctional conductance is a simple and sensitive function of intracellular pH. Science. 1981 Feb 13;211(4483):712–715. doi: 10.1126/science.6779379. [DOI] [PubMed] [Google Scholar]
  48. Spray D. C., Harris A. L., Bennett M. V. Voltage dependence of junctional conductance in early amphibian embryos. Science. 1979 Apr 27;204(4391):432–434. doi: 10.1126/science.312530. [DOI] [PubMed] [Google Scholar]
  49. Spray D. C., Stern J. H., Harris A. L., Bennett M. V. Gap junctional conductance: comparison of sensitivities to H and Ca ions. Proc Natl Acad Sci U S A. 1982 Jan;79(2):441–445. doi: 10.1073/pnas.79.2.441. [DOI] [PMC free article] [PubMed] [Google Scholar]
  50. Stern M. D. Buffering of calcium in the vicinity of a channel pore. Cell Calcium. 1992 Mar;13(3):183–192. doi: 10.1016/0143-4160(92)90046-u. [DOI] [PubMed] [Google Scholar]
  51. Swenson K. I., Jordan J. R., Beyer E. C., Paul D. L. Formation of gap junctions by expression of connexins in Xenopus oocyte pairs. Cell. 1989 Apr 7;57(1):145–155. doi: 10.1016/0092-8674(89)90180-3. [DOI] [PubMed] [Google Scholar]
  52. Turin L., Warner A. E. Intracellular pH in early Xenopus embryos: its effect on current flow between blastomeres. J Physiol. 1980 Mar;300:489–504. doi: 10.1113/jphysiol.1980.sp013174. [DOI] [PMC free article] [PubMed] [Google Scholar]
  53. Turin L., Warner A. Carbon dioxide reversibly abolishes ionic communication between cells of early amphibian embryo. Nature. 1977 Nov 3;270(5632):56–57. doi: 10.1038/270056a0. [DOI] [PubMed] [Google Scholar]
  54. Veenstra R. D., DeHaan R. L. Cardiac gap junction channel activity in embryonic chick ventricle cells. Am J Physiol. 1988 Jan;254(1 Pt 2):H170–H180. doi: 10.1152/ajpheart.1988.254.1.H170. [DOI] [PubMed] [Google Scholar]
  55. Veenstra R. D., DeHaan R. L. Measurement of single channel currents from cardiac gap junctions. Science. 1986 Aug 29;233(4767):972–974. doi: 10.1126/science.2426781. [DOI] [PubMed] [Google Scholar]
  56. Veenstra R. D. Developmental changes in regulation of embryonic chick heart gap junctions. J Membr Biol. 1991 Feb;119(3):253–265. doi: 10.1007/BF01868730. [DOI] [PubMed] [Google Scholar]
  57. Veenstra R. D. Voltage-dependent gating of gap junction channels in embryonic chick ventricular cell pairs. Am J Physiol. 1990 Apr;258(4 Pt 1):C662–C672. doi: 10.1152/ajpcell.1990.258.4.C662. [DOI] [PubMed] [Google Scholar]
  58. Wang H. Z., Li J., Lemanski L. F., Veenstra R. D. Gating of mammalian cardiac gap junction channels by transjunctional voltage. Biophys J. 1992 Jul;63(1):139–151. doi: 10.1016/S0006-3495(92)81573-4. [DOI] [PMC free article] [PubMed] [Google Scholar]
  59. Weingart R. Electrical properties of the nexal membrane studied in rat ventricular cell pairs. J Physiol. 1986 Jan;370:267–284. doi: 10.1113/jphysiol.1986.sp015934. [DOI] [PMC free article] [PubMed] [Google Scholar]
  60. Weingart R. The actions of ouabain on intercellular coupling and conduction velocity in mammalian ventricular muscle. J Physiol. 1977 Jan;264(2):341–365. doi: 10.1113/jphysiol.1977.sp011672. [DOI] [PMC free article] [PubMed] [Google Scholar]
  61. White R. L., Doeller J. E., Verselis V. K., Wittenberg B. A. Gap junctional conductance between pairs of ventricular myocytes is modulated synergistically by H+ and Ca++. J Gen Physiol. 1990 Jun;95(6):1061–1075. doi: 10.1085/jgp.95.6.1061. [DOI] [PMC free article] [PubMed] [Google Scholar]
  62. White R. L., Spray D. C., Campos de Carvalho A. C., Wittenberg B. A., Bennett M. V. Some electrical and pharmacological properties of gap junctions between adult ventricular myocytes. Am J Physiol. 1985 Nov;249(5 Pt 1):C447–C455. doi: 10.1152/ajpcell.1985.249.5.C447. [DOI] [PubMed] [Google Scholar]
  63. Wilders R., Jongsma H. J. Limitations of the dual voltage clamp method in assaying conductance and kinetics of gap junction channels. Biophys J. 1992 Oct;63(4):942–953. doi: 10.1016/S0006-3495(92)81664-8. [DOI] [PMC free article] [PubMed] [Google Scholar]

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