Advertisement

Pflügers Archiv

, Volume 428, Issue 1, pp 30–38 | Cite as

Dual-frequency method for synchronous measurement of cell capacitance, membrane conductance and access resistance on single cells

  • V. Rohlicek
  • A. Schmid
Molecular and Cellular Physiology

Abstracts

A dual-frequency method was developed to monitor changes of membrane capacitance, membrane conductance and serial resistance between patch pipette and cytoplasm of the cell in the whole-cell configuration. Measurement of real and imaginary components of cell admittance during excitation with two superimposed sinusoidal voltages of different frequencies provides mathematical solutions for all three variables. The validity of the method was verified with experiments on mast cells and exocrine pancreatic acinar cells. During degranulation of mast cells, induced by GTPγS in the pipette solution, a stepwise increase in membrane capacitance could be observed, indicating that the resolution of the method is below 10 fF. Precalibration of the setup allows all calculated data to be expressed as absolute values. The capacitance measurement proved to be rather independent of changes in the access resistance and in the cell membrane resistance over a wide range. The huge changes in membrane conductance of mouse pancreatic acinar cells during hormonal stimulation with acetylcholine produced a relative error of less than 0.3% in the capacitance trace. This allows a clear distinction between changes of membrane conductance and cell capacitance. The method therefore offers great advantages in the study of exocytosis as well as endocytosis in cell types, such as exocrine gland cells, with major changes in membrane conductance during hormonal stimulation.

Key words

Patch-clamp Cell membrane capacitance Cell membrane conductance Exocytosis Mast cell Exocrine pancreatic acinar cell 

Preview

Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.

References

  1. 1.
    Almers W, Neher E (1985) Gradual and stepwise changes in the membrane capacitance of rat peritoneal mast cells. J Physiol (Lond) 386:205–217Google Scholar
  2. 2.
    Clausen C, Fernandez JM (1981) A low-cost method for rapid transfer function measurements with direct application to biological impedance analysis. Pflügers Arch 390:290–295Google Scholar
  3. 3.
    Fernandez JM, Neher E, Gomperts BD (1984) Capacitance measurements reveal stepwise fusion events in degranulating mast cells. Nature 312:453–455Google Scholar
  4. 4.
    Fidler N, Fernandez JM (1989) Phase tracking: an improved phase detection technique for cell membrane capacitance measurements. Biophys J 56:1153–1162Google Scholar
  5. 5.
    Gillespie JI (1979) The effect of repetitive stimulation on the passive electrical properties of the presynaptic terminal of the squid giant synapse. Proc R Soc Lond [Biol] 206:293–306Google Scholar
  6. 6.
    Gillis KD, Misler S (1992) Single cell assay of exocytosis from pancreatic islet B cells. Pflügers Arch 420:121–123Google Scholar
  7. 7.
    Gögelein H, Dahlem D, Englert HC, Lang HJ (1990) Flufenamic acid, mefenamic acid and niflumic acid inhibit single non-selective cation channels in rat exocrine pancreas. FEBS Lett 268:79–82Google Scholar
  8. 8.
    Greger R, Allert N, Fröbe U, Normann C (1993) Increase in cytosolic Ca2+ regulates exocytosis and Cl conductances in HT29 cells. Pflügers Arch 424:329–334Google Scholar
  9. 9.
    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 391:85–100Google Scholar
  10. 10.
    Jaffe LA, Hagiwara S, Kado RT (1978) The time course of cortical vesicle fusion in sea urchin eggs observed as membrane capacitance changes. Dev Biol 67:243–248Google Scholar
  11. 11.
    Joshi C, Fernandez JM (1988) Capacitance measurements. An analysis of the phase detector technique used to study exocytosis and endocytosis. Biophys J 53:885–892Google Scholar
  12. 12.
    Lindau M, Fernandez JM (1986) A patch-clamp study of histamine-secreting cells. J Gen Physiol 88:349–368Google Scholar
  13. 13.
    Lindau M, Neher E (1988) Patch-clamp techniques for time-resolved capacitance measurements. Pflügers Arch 411:137–146Google Scholar
  14. 14.
    Maruyama Y (1986) Ca2+-induced excess capacitance fluctuation studied by phase-sensitive detection method in exocrine pancreatic cells. Pflügers Arch 407:561–563Google Scholar
  15. 15.
    Maruyama Y (1988) Agonist-induced changes in cell membrane capacitance and conductance in dialysed pancreatic acinar cells of rat. J Physiol (Lond) 406:299–313Google Scholar
  16. 16.
    Maruyama Y, Inooka G, Li YX, Miyashita Y, Kasai H (1993) Agonist-induced localized Ca2+ spikes directly triggering exocytotic secretion in exocrine pancreas. EMBO J 12:3017–3022Google Scholar
  17. 17.
    Neher E, Marty A (1982) Discrete changes of cell membrane capacitance observed under conditions of enhanced secretion in bovine adrenal chromaffin cells. Proc Natl Acad Sci USA 79:6712–6716Google Scholar
  18. 18.
    Okada Y, Hazama A, Hashimoto A, Maruyama Y, Kubo M (1992) Exocytosis upon osmotic swelling in human epithelial cells. Biochim Biophys Acta 1107:201–205Google Scholar
  19. 19.
    Osipchuk Y, Wakui M, Yule DI, Gallacher DV, Petersen OH (1990) Cytoplasmic Ca2+ oscillations evoked by receptor stimulation, G protein activation, internal application of inositol trisphosphate or Ca2+: simultaneous microfluorimetry and Ca2+-dependent chloride current recording in single pancreatic acinar cells. EMBO J 9:697–704Google Scholar
  20. 20.
    Petersen OH (1992) Stimulus-secretion coupling: cytoplasmic calcium signals and the control of ion channels in exocrine acinar cells. J Physiol (Lond) 448:1–51Google Scholar
  21. 21.
    Poronnik P, Cook DI, Allen DG, Young JA (1991) Diphenylamine-2-carboxylate (DPC) reduces calcium influx in a mouse mandibular cell line (ST885). Cell Calcium 12:441–447Google Scholar
  22. 22.
    Rohlicek V, Rohlicek J (1993) Measurement of membrane capacitance and resistance of single cells using two frequencies. Physiol Res 42:423–428Google Scholar
  23. 23.
    Thorn P, Petersen OH (1992) Activation of nonselective cation channels by physiological cholecystokinin concentrations in mouse pancreatic acinar cells. J Gen Physiol 100:11–25Google Scholar
  24. 24.
    Tse A, Tse FW, Almers W, Hille B (1993) Rhythmic exocytosis stimulated by GnRH-induced calcium oscillations in rat gonadotropes. Science 260:82–84Google Scholar

Copyright information

© Springer-Verlag 1994

Authors and Affiliations

  • V. Rohlicek
    • 1
  • A. Schmid
    • 2
  1. 1.Czech Academy of SciencePragueCzech Republic
  2. 2.Physiologisches InstitutUniversität des SaarlandesHomburg/SaarGermany

Personalised recommendations