Skip to main content
PMC home page
Biophysical Journal logoLink to Biophysical Journal
. 1986 Mar;49(3):767–771. doi: 10.1016/S0006-3495(86)83703-1

Sidedness of reconstituted calcium channels from muscle transverse tubules as determined by D600 and D890 blockade.

H Affolter, R Coronado
PMCID: PMC1329523  PMID: 2421795

Abstract

The verapamil-type calcium antagonist, D600, and its charged quaternary derivative, D890, were used to assess the sidedness of blockade in single calcium channels reconstituted from purified transverse tubules of skeletal muscle. Spontaneous single channel openings were induced with the agonist Bay-K8644 and recordings were made in a two-chamber planar bilayer setup so that drugs could be delivered to either side of the channel. Micromolar drug addition resulted in a greater than 10-fold decrease in probability of open channel events (po) without a significant change in single channel currents. Changes in po occurred in parallel with changes in mean open time and both parameters could be titrated with a similar IC50. At pH 7.2, cis or trans D600 blocked with an IC50 of 5 microM but for D890 the IC50 was cis 3 microM and trans greater than 75 microM (cis is the intracellular-equivalent side as defined by the voltage-dependent activation). The asymmetry of D890 blockade indicates that the drug can readily gain access to the blocking site from the aqueous phase adjacent to the inner but not extracellular end of the channel.

Full text

PDF
767

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Affolter H., Coronado R. Agonists Bay-K8644 and CGP-28392 open calcium channels reconstituted from skeletal muscle transverse tubules. Biophys J. 1985 Aug;48(2):341–347. doi: 10.1016/S0006-3495(85)83789-9. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Borsotto M., Norman R. I., Fosset M., Lazdunski M. Solubilization of the nitrendipine receptor from skeletal muscle transverse tubule membranes. Interactions with specific inhibitors of the voltage-dependent Ca2+ channel. Eur J Biochem. 1984 Aug 1;142(3):449–455. doi: 10.1111/j.1432-1033.1984.tb08307.x. [DOI] [PubMed] [Google Scholar]
  3. Cachelin A. B., de Peyer J. E., Kokubun S., Reuter H. Ca2+ channel modulation by 8-bromocyclic AMP in cultured heart cells. Nature. 1983 Aug 4;304(5925):462–464. doi: 10.1038/304462a0. [DOI] [PubMed] [Google Scholar]
  4. Curtis B. M., Catterall W. A. Purification of the calcium antagonist receptor of the voltage-sensitive calcium channel from skeletal muscle transverse tubules. Biochemistry. 1984 May 8;23(10):2113–2118. doi: 10.1021/bi00305a001. [DOI] [PubMed] [Google Scholar]
  5. Hamilton S. L., Yatani A., Hawkes M. J., Redding K., Brown A. M. Atrotoxin: a specific agonist for calcium currents in heart. Science. 1985 Jul 12;229(4709):182–184. doi: 10.1126/science.3160111. [DOI] [PubMed] [Google Scholar]
  6. Henry P. D. Comparative pharmacology of calcium antagonists: nifedipine, verapamil and diltiazem. Am J Cardiol. 1980 Dec 1;46(6):1047–1058. doi: 10.1016/0002-9149(80)90366-5. [DOI] [PubMed] [Google Scholar]
  7. Hescheler J., Pelzer D., Trube G., Trautwein W. Does the organic calcium channel blocker D600 act from inside or outside on the cardiac cell membrane? Pflugers Arch. 1982 Jun;393(4):287–291. doi: 10.1007/BF00581411. [DOI] [PubMed] [Google Scholar]
  8. Hille B. Local anesthetics: hydrophilic and hydrophobic pathways for the drug-receptor reaction. J Gen Physiol. 1977 Apr;69(4):497–515. doi: 10.1085/jgp.69.4.497. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Latorre R., Miller C. Conduction and selectivity in potassium channels. J Membr Biol. 1983;71(1-2):11–30. doi: 10.1007/BF01870671. [DOI] [PubMed] [Google Scholar]
  10. Latorre R., Vergara C., Hidalgo C. Reconstitution in planar lipid bilayers of a Ca2+-dependent K+ channel from transverse tubule membranes isolated from rabbit skeletal muscle. Proc Natl Acad Sci U S A. 1982 Feb;79(3):805–809. doi: 10.1073/pnas.79.3.805. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Lee K. S., Tsien R. W. High selectivity of calcium channels in single dialysed heart cells of the guinea-pig. J Physiol. 1984 Sep;354:253–272. doi: 10.1113/jphysiol.1984.sp015374. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Lee K. S., Tsien R. W. Mechanism of calcium channel blockade by verapamil, D600, diltiazem and nitrendipine in single dialysed heart cells. Nature. 1983 Apr 28;302(5911):790–794. doi: 10.1038/302790a0. [DOI] [PubMed] [Google Scholar]
  13. Moczydlowski E. G., Latorre R. Saxitoxin and ouabain binding activity of isolated skeletal muscle membrane as indicators of surface origin and purity. Biochim Biophys Acta. 1983 Jul 27;732(2):412–420. doi: 10.1016/0005-2736(83)90058-5. [DOI] [PubMed] [Google Scholar]
  14. Rosemblatt M., Hidalgo C., Vergara C., Ikemoto N. Immunological and biochemical properties of transverse tubule membranes isolated from rabbit skeletal muscle. J Biol Chem. 1981 Aug 10;256(15):8140–8148. [PubMed] [Google Scholar]
  15. Tsien R. W. Calcium channels in excitable cell membranes. Annu Rev Physiol. 1983;45:341–358. doi: 10.1146/annurev.ph.45.030183.002013. [DOI] [PubMed] [Google Scholar]

Articles from Biophysical Journal are provided here courtesy of The Biophysical Society

RESOURCES