Skip to main content
SpringerLink
Log in

Ca-dependent K channels in smooth muscle cells permeabilized by β-escin recorded using the cell-attached patch-clamp technique

  • Published:
Pflügers Archiv Aims and scope Submit manuscript

Abstract

Using the cell-attached patch-clamp technique, the activity of single, Ca-dependent K channels was recorded in single smooth muscle cells permeabilized by β-escin. The conductance and the relationship between the open probability of the channels and pCa recorded in permeabilized cells were very similar to those obtained in excised inside-out patches. At pCa 7, application of 30 μM acetylcholine (ACh) or 0.1 μM substance P (SP) together with 1 mM guanosine 5′-trisphosphate to permeabilized cells elicited transient bursts of channel openings similar to those which occur in intact cells. Transient activation was also observed when 2–30 μM inositol trisphosphate (IP3) was applied to permeabilized cells. This single channel activity was inhibited by pretreatment with low-molecular-weight heparin at 50–100 μg/ml. Channel activity at pCa 7.0 was greatly enhanced by 200 μM cyclic adenosine monophosphate. These results provide direct evidence that single Ca-dependent K channel activity is regulated by the transmitters ACh and SP, as well as a second messenger, IP3, via the release of intracellular Ca from intracellular sites which are blocked by heparin. This novel approach is valuable in elucidating second messenger mechanisms involved in the regulation of single channel activity by transmitters and autacoids, since permeabilization by β-escin preserves the entire system of receptor-operated signal transduction and allows intracellular application of second messengers at fixed concentrations.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Similar content being viewed by others

References

  • Akabarali H, Nakajima T, Wyse DG, Giles W (1990) Ca2+-activated K+ currents in smooth muscle. Can J Physiol Pharmacol 68:1489–1494

    Google Scholar 

  • Benham CD, Bolton TB (1986) Spontaneous transient outward currents in single visceral and vascular smooth muscle cells of the rabbit. J Physiol (Lond) 381:385–406

    Google Scholar 

  • Benham CD, Bolton TB, Lang RJ, Takewaki T (1986) Calcium-activated potassium channels in single smooth muscle cells of rabbit jejunum and guinea-pig mesenteric artery. J Physiol (Lond) 371:45–67

    Google Scholar 

  • Berridge MJ, Downes CP, Hanley MR (1982) Lithium amplifies agonist-dependent phosphatidylinositol responses in brain and salivary glands. Biochem J 206:587–595

    Google Scholar 

  • Best PM (1983) Cardiac muscle function: results from skinned fiber preparations. Am J Phsiol 244:H167-H177

    Google Scholar 

  • Bolotina V, Omelyanenko V, Ryan U, Bregestovski P (1989) Variations of membrane cholesterol alter the kinetics of Ca2+-dependent K+ channels and membrane fluidity in vascular smooth muscle cells. Pflügers Arch 415:262–268

    Google Scholar 

  • Bolton TB, Lim SP (1989) Properties of calcium stores and transient outward currents in single smooth muscle cells from rabbit intestine. J Physiol (Lond) 409:385–401

    Google Scholar 

  • Brami BA, Lei U, Hauser G (1991) Influence of lithium on second messenger accumulation in NG108-15 cells. Biochem Biophys Res Commun 174:606–612

    Google Scholar 

  • Bregestovski PD, Bolotina VM, Serebryakov VN (1989) Fatty acid modifies Ca2+-dependent potassium channel activity in smooth muscle cells from the human aorta. Proc R Soc Lond B 237:259–266

    Google Scholar 

  • Chilvers ER, Challiss RAJ, Willcocks AL, Potter BVL, Braners PJ, Nahorski SR (1990) Characterization of stereospecific binding sites for inositol 1,4,5-trisphosphate in airway smooth muscle. Br J Pharmacol 99:297–302

    Google Scholar 

  • Cullen PJ, Comerford JG, Dawson AP (1988) Heparin inhibits the inositol 1,4,5-trisphosphate-induced Ca2+ release from rat liver microsomes. FEBS Lett 228:57–59

    Google Scholar 

  • Doyle VM, Rüegg UT (1985) Vasopressin induced production of inositol trisphosphate and calcium efflux in a smooth muscle cell line. Biochem Biophys Res Commun 131:469–476

    Google Scholar 

  • Drummond AH, Raeburn CA (1984) The interaction of lithium with thyrotropin-releasing hormone-stimulated lipid metabolism in GH3 pituiatry tumor cells. Biochem J 224:129–136

    Google Scholar 

  • Ehrlich BE, Watras J (1988) Inositol 1,4,5-triphosphate activates a channel from smooth muscle sarcoplasmic reticulum. Nature 336:583–586

    Google Scholar 

  • Endo M, Iino M (1980) Specific perforation of muscle cell membranes with preserved SR functions by saponin treatment. J Muscle Res Cell Motil 1:89–100

    Google Scholar 

  • Fabiato A (1981) Myoplasmic free calcium concentration reached during the twitch of an intact isolated cardiac cell and during calcium-induced release of calcium from the sarcoplasmic reticulum of a skinned cardiac cell from the adult rat or rabbit ventricle. J Gen Physiol 78:457–497

    Google Scholar 

  • Fabiato A (1985) Rapid ionic modifications during the aequorin-detected calcium transient in a skinned canine cardiac Purkinje cell. J Gen Physiol 85:189–246

    Google Scholar 

  • Ganitkevich V, Isenberg G (1990) Isolated guinea pig coronary smooth muscle cells; acetylcholine induces hyperpolarization due to sarcoplasmic reticulum calcium release activating potassium channels. Circ Res 67:525–528

    Google Scholar 

  • Gennis RB (1989) Biomembranes. In: Cantor CR (ed) Springer advanced texts in chemistry. Springer, New York Berlin Heidelberg, pp 419–510

    Google Scholar 

  • Ghosh TK, Eis PS, Mullaney JM, Ebert CL, Gill DL (1988) Competitive, reversible, and potent antagonism of inositol 1,4,5-trisphosphate-activated calcium release by heparin. J Biol Chem 263: 11075–11079

    Google Scholar 

  • Gordon AR (1978) Contraction of detergent-treated smooth muscle. Proc Natl Acad Sci USA 75:3527–3530

    Google Scholar 

  • Hamill OP, Marty A, Neher E, Sakmann B, Sigworth FJ (1981) Improved patch-clamp techniques for high-resolution current recording from cells and cell-free membrane patches. Pflügers Arch 391:85–100

    Google Scholar 

  • Hilger GA, Gratzl M (1988) Controlled manipulation of the cell interior by pore-forming proteins. Trends Pharmacol Sci 9:195–197

    Google Scholar 

  • Iino M (1981) Tension responses of chemically skinned fibre bundles of the guinea-pig taenia caeci under varied ionic environments. J Physiol (Lond) 320:449–467

    Google Scholar 

  • Iino M (1989) Calcium-induced calcium release mechanism in guinea pig taenia caeci. J Gen Physiol 94:363–383

    Google Scholar 

  • Imaizumi Y, Muraki K, Takeda M, Watanabe M (1989) Measurement and simulation of noninactivating Ca currents in smooth muscle cells. Am J Physiol 256:C880-C885

    Google Scholar 

  • Imaizumi Y, Muraki K, Watanabe M (1990) Characteristics of transient outward currents in smooth muscle cells from the ureter of the guinea-pig. J Physiol (Lond) 427:301–324

    Google Scholar 

  • Ito H, Takikawa R, Iguchi M, Hamada E, Sugimoto T, Kurachi Y (1990) Heparin uncouples the muscarinic receptors from GK protein in the atrial cell membrane of the guinea-pig heart. Pflügers Arch 417:126–128

    Google Scholar 

  • Izumi K, Tasaka K (1989) Histamine release from beta-escin-permeabilized rat peritoneal mast cells and its inhibition by intracellular Ca2+ blockers, calmodulin inhibitors and cAMP. Immunopharmacology 18:177–186

    Google Scholar 

  • Kaibara M, Kameyama M (1988) Inhibition of the calcium channel by intracellular protons in single ventricular myocyte of the guinea-pig. J Physiol (Lond) 403:621–640

    Google Scholar 

  • Kakei M, Noma A, Shibasaki T (1985) Properties of adenosine-triphosphate-regulated potassium channels in guinea-pig ventricular cells. J Physiol (Lond) 363:441–462

    Google Scholar 

  • Kitazawa T, Kobayashi S, Horiuchi K, Somlyo AV, Somlyo AP (1989) Receptor-coupled, permeabilized smooth muscle. J Biol Chem 264:5339–5342

    Google Scholar 

  • Klöckner U, Isenberg G (1985) Action potentials and net membrane currents of isolated smooth muscle cells (urinary bladder of the guinea-pig). Pflügers Arch 405:329–339

    Google Scholar 

  • Kobayashi S, Somlyo AV, Somylo AP (1988) Heparin inhibits the inositol 1,4,5-triphosphate-dependent, but not the independent, calcium release induced by guanine nucleotide in vascular smooth muscle. Biochem Biophys Res Commun 153:625–631

    Google Scholar 

  • Kobabayashi S, Kitazawa T, Somlyo AV, Somlyo AP (1989) Cytosolic heparin inhibits muscarinic and -adrenergic Ca2+ release in smooth muscle. J Biol Chem 264:17997–18004

    Google Scholar 

  • Kobayashi S, Gong MC, Somlyo AV, Somlyo AP (1991) Ca2+ channel blockers distinguish between GE protein-coupled pharmacomechanical Ca2+ release and Ca2+ sensitization. Am J Physiol 260:C364-C370

    Google Scholar 

  • Komori S, Bolton TB (1989) Actions of guanine nucleotides and cyclic nucleotides on calcium stores in single patch-clamped smooth muscle cells from rabbit portal vein. Br J Pharmacol 97:973–982

    Google Scholar 

  • Komori S, Bolton TB (1990) Roles of G-proteins in muscarinic receptors inward current and outward currents in rabbit jejunum smooth muscle. J Physiol (Lond) 427:395–419

    Google Scholar 

  • Komori S, Bolton TB (1991) Calcium release induced by inositol 1,4,5-trisphosphate in single rabbit intestinal smooth muscle cells. J Physiol (Lond) 433:495–517

    Google Scholar 

  • Kume H, Takai A, Tokuno H, Tomita T (1989) Regulation of Ca2+-dependent K+-channel activity in tracheal myocytes by phosphorylation. Nature 341:152–154

    Google Scholar 

  • Levitan ES, Kramer RH (1990) Neuropeptide modulation of single calcium and potassium channels detected with a new patch clamp configuration. Nature 348:545–547

    Google Scholar 

  • Loirand G, Pacaud P, Baron A, Mironneau C, Mironneau J (1991) Large conductance calcium-activated non-selective cation channel in smooth muscle cells isolated from rat portal vein. J Physiol (Lond) 437:461–475

    Google Scholar 

  • Mayer EA, Loo DD, Snape WJ Jr, Sachs G (1990a) The activation of calcium and calcium-activated potassium channels in mammalian colonic smooth muscle by substance P. J Physiol (London) 420:47–71

    Google Scholar 

  • Mayer EA, Loo DD, Kodner A, Reddy SN (1990b) Differential modulation of Ca2+-activated K+ channels by substance P. Am J Physiol 257:G887-G987

    Google Scholar 

  • Muraki K, Imaizumi Y, Kojima T, Kawai T, Watanabe M (1990) Effects of tetraethylammonium and 4-aminopyridine on outward currents and excitability in canine tracheal smooth muscle cells. Br J Pharmacol 100:507–515

    Google Scholar 

  • Natori R (1954) The property and contraction process of isolated myofibrils. Jikeikai Med J 1:119–126

    Google Scholar 

  • Nishimura J, Kolber M, Breemen C van (1989) Norepinephrine and GTP-r-S increase myofilament Ca2+ sensitivity in alpha-toxin permeabilized arterial smooth muscle. Biochem Biophys Res Commun 157:677–683

    Google Scholar 

  • Ohya Y, Sperelakis N (1989) Modulation of single slow (L-type) calcium channels by intracellular ATP in vascular smooth muscle cells. Pflügers Arch 414:257–264

    Google Scholar 

  • Sadoshima J, Akaike N, Kanaide H, Nakamura M (1988) Cyclic AMP modulates Ca-activated K channel in cultured smooth muscle cells of rat aortas. Am J Physiol 255:H754-H759

    Google Scholar 

  • Saida K (1982) Intracellular Ca release in skinned smooth muscle. J Gen Physiol 80:191–202

    Google Scholar 

  • Sims SM, Vivaudou MB, Hillemeier C, Biancani P, Walsh JV, Singer JJ (1990) Membrane currents and cholinergic regulation of K+ current in esophageal smooth muscle cells. Am J Physiol 258:G794-G802

    Google Scholar 

  • Streb H, Irvine RF, Berridge MJ, Schulz I (1983) Release of Ca2+ from a non-mitochondrial intracellular store of pancreatic acinar cells by inositol 1,4,5-trisphosphate. Nature 306:67–69

    Google Scholar 

  • Takeda M, Imaizumi Y, Watanabe M (1991) Effects of noradrenaline and heparin on outward current in single smooth muscle cells of the guinea-pig vas deferens. Eur J Pharmacol 193:375–378

    Google Scholar 

  • Thomas AP, Alexander J, Williamson JR (1984) Relationship between inositol polyphosphate production and the increase of cytosolic free Ca2+ induced by vasopressin in isolated hepatocytes. J Biol Chem 259:5574–5584

    Google Scholar 

  • Willuweit B, Aktories K (1988) Heparin uncouples alpha 2-adrenoceptors from the Gi-protein in membranes of human platelets. Biochem J 249:857–863

    Google Scholar 

  • Winegrad S (1971) Studies of cardiac muscle with a high permeability to calcium produced by treatment with ethylenediaminetetraacetic acid. J Gen Physiol 58:71–93

    Google Scholar 

  • Yamamoto H, Kanaide H, Nakamura M (1990) Heparin specifically inhibits the inositol 1,4,5-trisphosphate-induced Ca2+ release from skinned rat aortic smooth muscle cells in primary culture. Naunyn Schmiedebergs Arch Pharmacol 141:273–278

    Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Department of Chemical Pharmacology, Faculty of Pharmaceutical Sciences, Nagoya City University, 3-1 Tanabedori Mizuhoku, 467, Nagoya, Japan

    Katsuhiko Muraki, Yuji Imaizumi & Minoru Watanabe

Authors
  1. Katsuhiko Muraki
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Yuji Imaizumi
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Minoru Watanabe
    View author publications

    You can also search for this author in PubMed Google Scholar

Rights and permissions

Reprints and permissions

About this article

Cite this article

Muraki, K., Imaizumi, Y. & Watanabe, M. Ca-dependent K channels in smooth muscle cells permeabilized by β-escin recorded using the cell-attached patch-clamp technique. Pflugers Arch. 420, 461–469 (1992). https://doi.org/10.1007/BF00374620

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Issue Date:

  • DOI: https://doi.org/10.1007/BF00374620

Key words

Search

Navigation