title: Electrical stimulation of cultured neurons using a simply patterned indium-tin-oxide (ITO) glass electrode.
abstract: BACKGROUND
  Indium-tin-oxide (ITO) glass electrodes possess the properties of optical transparency and high electrical conductivity, which enables the electrical stimulation of cultured cells to be performed whilst also measuring the responses with fluorescent imaging techniques. However, the quantitative relationship between the intensity of the stimulating current and the cell response is unclear when using conventional methods that employ a separated configuration of counter and stimulation electrodes.
  
  
  METHODS
  A quantitative electrical current stimulation device without the use of a counter electrode was fabricated.
  
  
  RESULTS
  Nerve growth factor (NGF)-induced differentiated PC12 cells were cultured on an ITO single glass electrode, and the Ca(2+) response to electrical stimuli was measured using fluorescent Ca(2+) imaging. ITO electrode devices with a width less than 0.1mm were found to evoke a Ca(2+) response in the PC12 cells. Subsequent variation in the length of the device in the range of 2-10mm was found to have little influence on the efficiency of the electric stimulus. We found that the stimulation of the cells was dependent on the electrical current, when greater than 60 μA, rather than on the Joule heat, regardless of the width and length of the conductive area.
  
  
  CONCLUSIONS
  Because of the cells directly in contact with the electrode, our device enables to stimulate the cells specifically, comparing with previous devices with the counter electrode.
  
  
  CONCLUSIONS
  The ITO device without the use of a counter electrode is a useful tool for evaluating the quantitative neural excitability of cultured neurons.
authors.affiliation: Department of Bioscience and Informatics, Faculty of Science and Technology, Keio University, 3-14-1 Hiyoshi, Kohoku-ku, Yokohama, 223-8522, Japan.
authors.forename: Ryo
authors.initials: R
authors.lastname: Tanamoto
authors.affiliation: Department of Bioscience and Informatics, Faculty of Science and Technology, Keio University, 3-14-1 Hiyoshi, Kohoku-ku, Yokohama, 223-8522, Japan.
authors.forename: Yutaka
authors.initials: Y
authors.lastname: Shindo
authors.affiliation: Department of Mechanical Engineering, Faculty of Science and Technology, Keio University, 3-14-1 Hiyoshi, Kohoku-ku, Yokohama, 223-8522, Japan.
authors.forename: Norihisa
authors.initials: N
authors.lastname: Miki
authors.affiliation: Department of Applied Physics and Physico-Informatics, Faculty of Science and Technology, Keio University, 3-14-1 Hiyoshi, Kohoku-ku, Yokohama, 223-8522, Japan.
authors.forename: Yoshinori
authors.initials: Y
authors.lastname: Matsumoto
authors.affiliation: Department of Bioscience and Informatics, Faculty of Science and Technology, Keio University, 3-14-1 Hiyoshi, Kohoku-ku, Yokohama, 223-8522, Japan.
authors.forename: Kohji
authors.initials: K
authors.lastname: Hotta
authors.affiliation: Department of Bioscience and Informatics, Faculty of Science and Technology, Keio University, 3-14-1 Hiyoshi, Kohoku-ku, Yokohama, 223-8522, Japan. Electronic address: oka@bio.keio.ac.jp.
authors.forename: Kotaro
authors.initials: K
authors.lastname: Oka
mesh_terms: D000814:Aniline Compounds / Q000378:metabolism; D000818:Animals; D001703:Biophysics; D002118:Calcium / Q000378:metabolism; D002478:Cells, Cultured; D004558:Electric Stimulation / Q000379:methods*; D006624:Hippocampus / Q000166:cytology; D008564:Membrane Potentials / Q000502:physiology; D008839:Microelectrodes; D020932:Nerve Growth Factor / Q000494:pharmacology; D009474:Neurons / Q000187:drug effects / Q000502:physiology*; D016716:PC12 Cells / Q000187:drug effects; D051381:Rats; D017208:Rats, Wistar; D013696:Temperature; D017971:Tin Compounds; D014966:Xanthenes / Q000378:metabolism
chemical_list: D000814:Aniline Compounds; C409648:Fluo 4; D017971:Tin Compounds; D014966:Xanthenes; C109984:indium tin oxide; D020932:Nerve Growth Factor; D002118:Calcium
keywords: Calcium imaging; Counter electrode; Hippocampal neurons; Indium-tin-oxide; NGF-differentiated PC12 cell; Patterning