Patch Clamp Techniques for Plant Cells

  • J. Theo M. Elzenga


The study of electrophysiological processes at the molecular level, with exquisite sensitivity and full control over the experimental conditions, is possible with the patch clamp technique. A concise overview is given of the different configurations that are used with the technique. The patch clamp technique critically depends on the formation of a tight, gigaOhm, seal between the glass tip of the measuring electrode and the cell membrane, which for plant cells means that the cell wall has to be removed. Methods that have been developed specifically to gain access to the membrane, such as laser microsurgery and protoplast release, are discussed. This chapter also provides a review of the factors that influence the interaction between the glass tip of the electrode and cell membrane.


Guard Cell Bath Solution Patch Clamp Pipette Solution Patch Clamp Technique 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.


  1. Arend M, Stinzing A, Wind C, Langer K, Latz A, Ache P, Fromm J, Hedrich R (2005) Polar-localised poplar K+ channel capable of controlling electrical properties of wood-forming cells. Planta 223:140–148PubMedCrossRefGoogle Scholar
  2. Auerbach A, Sachs F (1985) High resolution patch-clamp techniques. In: Smith TG, Lecar H, Redman SJ, Gage PW (eds) Voltage and patch clamping with microelectrodes. American Physiological Society, Baltimore, pp 121–149Google Scholar
  3. Blom-Zandstra M, Koot HTM, van Hattum J, Vogelzang SA (1995) Isolation of protoplasts for patch-clamping experiments: an improved method requiring minimal amounts of adult leaf or root tissue from monocotyledonous or dicotyledonous plants. Protoplasma 185:1–6CrossRefGoogle Scholar
  4. Boehle T, Benndorf K (1994) Facilitated giga-seal formation with a just originated glass surface. Pflugers Arch 427:487–491CrossRefGoogle Scholar
  5. Brownlee C (1994) Microelectrode techniques in plant cells and microorganisms. In: Ogden DC (ed) Microelectrode techniques, the plymouth workshop handbook, 2nd edn. Company of Biologists, Cambridge, pp 347–359Google Scholar
  6. Brudern A, Thiel G (1999) Effect of cell-wall digesting enzymes on physiological state and competence of maize coleoptile cells. Protoplasma 209:246–255CrossRefGoogle Scholar
  7. Bush DS, Hedrich R, Schroeder JI, Jones RL (1988) Channel-mediated K+ flux in barley aleurone protoplasts. Planta 176:368–377CrossRefGoogle Scholar
  8. Colquhoun D, Hawkes AG (1995) The principles of the stochastic interpretation of ion-channel mechanisms. In: Sakmann B, Neher E (eds) Single-channel recording, 2nd edn. Plenum Press, New York, pp 397–482Google Scholar
  9. Colquhoun D, Sigworth FJ (1995) Fitting and statistical analysis of single-channel records. In: Sakmann B, Neher E (eds) Single-channel recording, 2nd edn. Plenum Press, New York, pp 483–587Google Scholar
  10. Davey MR, Anthony P, Power JB, Lowe KC (2005) Plant protoplasts: status and biotechnological perspectives. Biotechnol Adv 23:131–171PubMedCrossRefGoogle Scholar
  11. Davies JM, Poole RJ, Rea PA, Sanders D (1992) Potassium transport into plant vacuoles energized directly by a proton-pumping inorganic pyrophosphatase. Proc Nat Acad Sci USA 89:11701–11705PubMedCrossRefGoogle Scholar
  12. Demidchik V, Sokolik A, Yurin V (2006) Electrophysiological characterization of plant cation channels. In: Volkov A (ed) Plant electrophysiology: theory and methods. Springer, Berlin, pp 173–183CrossRefGoogle Scholar
  13. Dutta AK, Korchev YE, Shevchuk AI, Hayashi S, Okada Y, Sabirov RZ (2008) Spatial distribution of maxi-anion channel on cardiomyocytes detected by smart-patch technique. Biophys J 94:1646–1655PubMedCrossRefGoogle Scholar
  14. Elzenga JTM, Van Volkenburgh E (1997) Characterization of a light-controlled anion channel in the plasma membrane of mesophyll cells of pea. Plant Physiol 113:1419–1426PubMedGoogle Scholar
  15. Elzenga JTM, Keller CP, Van Volkenburgh E (1991) Patch clamping protoplasts from vascular plants: method for the quick isolation of protoplasts having a high success rate of gigaseal formation 97:1573–1575Google Scholar
  16. Fairley KA, Walker NA (1989) Patch clamping corn protoplasts. Gigaseal frequency is not improved by Congo red inhibition of cell wall regeneration. Protoplasma 153:111–116CrossRefGoogle Scholar
  17. Gadsby DC (2009) Ion channels versus ion pumps: the principal difference, in principle. Nat Rev Mol Cell Biol 10:344–352PubMedCrossRefGoogle Scholar
  18. Hafke JB, Furch ACU, Reitz MU, van Bel AJE (2007) Functional sieve element protoplasts. Plant Physiol 145:703–711PubMedCrossRefGoogle Scholar
  19. 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(2):85–100PubMedCrossRefGoogle Scholar
  20. Hedrich R, Schroeder JI (1989) The physiology of ion channels and electrogenic pumps in higher plants. Annu Rev Plant Physiol Plant Mol Biol 40:539–569CrossRefGoogle Scholar
  21. Henriksen GH, Assmann SM (1997) Laser-assisted patch clamping: a methodology. Pfluegers Arch 433:832–841CrossRefGoogle Scholar
  22. Henriksen GH, Taylor AR, Brownlee C, Assmann SM (1996) Laser microsurgery of higher plant cell walls permits patch-clamp access. Plant Physiol 110:1063–1068PubMedGoogle Scholar
  23. Hille B (1992) Ion channels of excitable membranes. Sinaur Associates, Sunderland, p 607Google Scholar
  24. Ivashikina N, Deeken R, Sche P, Kranz E, Pommerrenig B, Sauer N, Hedrich R (2003) Isolation of AtSUC2 promoter-GFP-marked companion cells for patch-clamp studies and expression profiling. Plant J 36:931–945PubMedCrossRefGoogle Scholar
  25. Kennedy BF, De Filippis LF (2004) Tissue degradation and enzymatic activity observed during protoplast isolation in two ornamental Grevillea species. In Vitro Cell Dev Biol 40:119–125Google Scholar
  26. Klercker JAf (1892) Eine Methode zur Isolierung lebender Protoplasten. Ofversigt af kong vetenskaps-akademiens forhandlingar 9:463–474Google Scholar
  27. Korchev YE, Negulyaev YA, Edwards CRW, Vodyanoy I, Lab MJ (2000) Functional localization of single active ion channels on the surface of a living cell. Nat Cell Biol 2:616–619PubMedCrossRefGoogle Scholar
  28. Lapointe J-Y, Szabo G (1987) A novel holder allowing internal perfusion of patch-clamp pipettes. Pfluegers Arch Eur J Physiol 410:212–216CrossRefGoogle Scholar
  29. Latorre R, Miller C (1983) Conduction and selectivity in potassium channels. J Membr Biol 71:11–30PubMedCrossRefGoogle Scholar
  30. Lebaudy A, Vavasseur A, Hosy E, Dreyer I, Leonhardt N, Thibaud J-B, Very AA, Simonneau T, Sentenac H (2008) Plant adaptation to fluctuating environment and biomass production are strongly dependent on guard cell potassium channels. PNAS 105:5271–5276PubMedCrossRefGoogle Scholar
  31. Levchenko V, Guinot DR, Klein M, Roelfsema MRG, Hedrich R, Dietrich P (2008) Stringent control of cytoplasmic Ca2+ in guard cells of intact plants compared to their counterparts in epidermal strips or guard cell protoplasts. Protoplasma 233:61–72PubMedCrossRefGoogle Scholar
  32. Lew RR (1991) Substrate regulation of single potassium and chloride ion channels in Arabidopsis plasma membrane. Plant Physiol 95:642–647PubMedCrossRefGoogle Scholar
  33. Maathuis FJM, Prins HBA (1990) Patch clamp studies on root cell vacuoles of a salt-tolerant and a salt-sensitive Plantago species. Plant Physiol 92:23–28PubMedCrossRefGoogle Scholar
  34. Maathuis FJM, Prins HBA (1991) Inhibition of inward rectifying tonoplast channels by a vacuolar factor: physiological and kinetic implications. J Membr Biol 122:251–258PubMedCrossRefGoogle Scholar
  35. Maathuis FJM, Taylor AR, Assmann SM, Sanders D (1997) Seal-promoting solutions and pipette perfusion for patch clamping plant cells. Plant J 11:891–896PubMedCrossRefGoogle Scholar
  36. Maathuis FJM, May ST, Graham NS, Bowen HC, Jelitto TC, Trimmer P, Bennett MJ, Sanders D, White PJ (1998) Cell marking in Arabidopsis thaliana and its application to patch-clamp studies. Plant J 15:843–851PubMedCrossRefGoogle Scholar
  37. Malhoubi M, Ostadi H, Wang S, Gu Y, Jiang K (2009) Effects of the surface morphology of pipette tip on giga-seal formation. Eng Lett 17:4Google Scholar
  38. Miedema H, Henriksen GH, Assmann SM (1999) A laser microsurgical method of cell wall removal allows detection of large-conductance ion channels in the guard cell plasma membrane. Protoplasma 209:58–67PubMedCrossRefGoogle Scholar
  39. Milton RL, Caldwell JH (1990) How do patch clamp seals form? A lipid bleb model. Pfluegers Arch Eur J Physiol 416:758–765CrossRefGoogle Scholar
  40. Moran N, Ehrenstein G, Iwasa K, Bare C, Mischke C (1984) Ion channels in plasmalemma of wheat protoplasts. Science 226:835–883PubMedCrossRefGoogle Scholar
  41. Ogden D (ed) (1994) Microelectrodes Techniques. The Plymouth Workshop Handbook. The Company of Biologists Ltd, Cambridge, p 448Google Scholar
  42. Qian YC, Nguyen T, Murphy TM (1993) Effect of washing on the plasma membrane and on stress reactions of cultured rose cells. Plant Cell, Tissue Organ Cult 35:245–252CrossRefGoogle Scholar
  43. Schauf CL, Wilson KJ (1987a) Effects of abscisic acid on K+ channels in Vicia faba guard cell protoplasts. Biochem Biophys Res Commun 145:284–290PubMedCrossRefGoogle Scholar
  44. Schauf CL, Wilson KJ (1987b) Properties of single K+ and Cl- channels in Asclepias tuberosa protoplasts. Plant Physiol 85:413–418PubMedCrossRefGoogle Scholar
  45. Schroeder JI, Hedrich R, Fernandez JM (1984) Potassium-selective single channels in guard cell protoplasts of Vicia faba. Nature 312:361–362CrossRefGoogle Scholar
  46. Suchyna TM, Markin VS, Sachs F (2009) Biophysics and structure of the patch and the gigaseal. Biophys J 97:738–747PubMedCrossRefGoogle Scholar
  47. Vogelzang SA, Prins HBA (1992) Plasmalemma patch clamp experiments in plant root cells: procedure for fast isolation of protoplasts with minimal exposure to cell wall degrading enzymes. Protoplasma 171:104–109CrossRefGoogle Scholar
  48. Ward JM (1997) Patch-clamping and other molecular approaches for the study of plasma membrane transporters delmystified. Plant Physiol 114:1151–1159PubMedCrossRefGoogle Scholar
  49. Ward JM, Mäser P et al (2009) Plant ion channels: gene families, physiology, and functional genomics analyses. Annu Rev Physiol 71:59–82PubMedCrossRefGoogle Scholar

Copyright information

© Springer-Verlag Berlin Heidelberg 2012

Authors and Affiliations

  1. 1.Plant EcophysiologyUniversity of GroningenGroningenThe Netherlands

Personalised recommendations