Patch-Clamp Biosensor Method

  • Seiji Hayashi
  • Makoto Tominaga
Part of the Springer Protocols Handbooks book series (SPH)


Cells or membranes with ligand-gated ion channels endogenously expressed in cultured cells or heterologously expressed in HEK293 cells can be used as biosensors with a patch-clamp technique to detect small amounts of locally released bioactive substances that are known to activate ionotropic receptors with high affinity. We describe a detailed procedure for a patch-clamp biosensor method by focusing on the detection of ATP using undifferentiated PC12 and HEK293 cells expressing P2X2 ion channels.


PC12 Cell Sensor Cell Bioactive Substance Inlet Tube Patch Membrane 
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. 1.
    Hume RI, Role LW, Fischbach GD (1983) Acetylcholine release from growth cones detected with patches of acetylcholine receptor-rich membranes. Nature 305:632–634PubMedCrossRefGoogle Scholar
  2. 2.
    Young SH, Poo MM (1983) Spontaneous release of transmitter from growth cones of embryonic neurones. Nature 305:634–637PubMedCrossRefGoogle Scholar
  3. 3.
    Schwartz EA (1987) Depolarization without calcium can release gamma-aminobutyric acid from a retinal neuron. Science 238:350–355PubMedCrossRefGoogle Scholar
  4. 4.
    Kataoka Y, Ohmori H (1994) Activation of glutamate receptors in response to membrane depolarization of hair cells isolated from chick cochlea. J Physiol 477:403–414PubMedGoogle Scholar
  5. 5.
    Bell PD, Lapointe J-Y, Sabirov R, Hayashi S, Peti-Peterdi J, Manabe K, Kovacs G, Okada Y (2003) Macula densa cell signaling involves ATP release through a maxi anion channel. Proc Natl Acad Sci USA 100:4322–4327PubMedCrossRefGoogle Scholar
  6. 6.
    Hazama A, Hayashi S, Okada Y (1998) Cell surface measurements of ATP release from single pancreatic beta cells using a novel biosensor technique. Pflugers Arch 437:31–35PubMedCrossRefGoogle Scholar
  7. 7.
    Hazama A, Shimizu T, Ando-Akatsuka Y, Hayashi S, Tanaka S, Maeno E, Okada Y (1999) Swelling-induced, CFTR-independent ATP release from a human epithelial cell line: lack of correlation with volume-sensitive Cl channels. J Gen Physiol 114:525–533PubMedCrossRefGoogle Scholar
  8. 8.
    Hayashi S, Hazama A, Dutta AK, Sabirov RZ, Okada Y (2004) Detecting ATP release by a biosensor method. Sci STKE 2004:pl14CrossRefGoogle Scholar
  9. 9.
    Dutta AK, Sabirov RZ, Uramoto H, Okada Y (2004) Role of ATP-conductive anion channel in ATP release from neonatal rat cardiomyocytes in ischaemic or hypoxic conditions. J Physiol 559:799–812PubMedGoogle Scholar
  10. 10.
    Mandadi S, Sokabe T, Shibasaki K, Katanosaka K, Mizuno A, Moqrich A, Patapoutian A, Fukumi-Tominaga T, Mizumura K, Tominaga M (2009) TRPV3 in keratinocytes transmits temperature information to sensory neurons via ATP. Pflugers Arch 458:1093–1102PubMedCrossRefGoogle Scholar
  11. 11.
    Beigi R, Kobatake E, Aizawa M, Dubyak GR (1999) Detection of local ATP release from activated platelets using cell surface-attached firefly luciferase. Am J Physiol 276:C267–C278PubMedGoogle Scholar
  12. 12.
    Schneider SW, Egan M, Jena BP, Guggino WB, Oberleithner H, Geibel JP (1999) Continuous detection of extracellular ATP on living cells by using atomic force microscopy. Proc Natl Acad Sci USA 96:12180–12185PubMedCrossRefGoogle Scholar
  13. 13.
    Arslan G, Filipeanu CM, Irenius E, Kull B, Clementi E, Allgaier C, Erlinge D, Fredholm BB (2000) P2Y receptors contribute to ATP-induced increases in intracellular calcium in differentiated but not undifferentiated PC12 cells. Neuropharmacology 39:482–496PubMedCrossRefGoogle Scholar

Copyright information

© Springer 2012

Authors and Affiliations

  1. 1.Discovery Research LaboratoryNippon Shinyaku Co., LtdKyotoJapan
  2. 2.Division of Cell SignalingOkazaki Institute for Integrative Bioscience (National Institute for Physiological Sciences), National Institutes of Natural SciencesOkazakiJapan
  3. 3.Department of Physiological SciencesThe Graduate University for Advanced StudiesOkazakiJapan

Personalised recommendations