Unit

UNIT 7.11 Measurement of Cation Movement in Primary Cultures Using Fluorescent Dyes

  1. Ian J. Reynolds

Published Online: 1 MAY 2001

DOI: 10.1002/0471142301.ns0711s04

Current Protocols in Neuroscience

Current Protocols in Neuroscience

How to Cite

Reynolds, I. J. 2001. Measurement of Cation Movement in Primary Cultures Using Fluorescent Dyes. Current Protocols in Neuroscience. 4:7.11:7.11.1–7.11.17.

Author Information

  1. University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania

Publication History

  1. Published Online: 1 MAY 2001
  2. Published Print: AUG 1998

Abstract

Ca2+, Na+, K+, and Mg2+ have a central role in neuronal excitability. The concentration of these cations in the cytoplasm of neurons (generically termed [ion+]i) provides a marker of the excitation state of the neurons, and may also illuminate the activity of specific signaling mechanisms that involve Ca2+- or Mg2+-activated enzymes. The measurement of [ion+]i in cultured neurons is achieved with the use of an ion-sensitive fluorescent dye in combination with equipment designed to quantitatively measure fluorescence. Specificity is obtained by choosing dyes with the appropriate selectivity for the ion of interest. Measurements of steady state ion concentrations can be made, as well as measurements of the net difference between ion movement into the cytoplasm (in response to a stimulus) and the physiological buffering of that ion. The procedures in this unit for loading and recording from dyes are broadly similar for each ion when ratiometric dyes are used as described, and can readily be modified for use with single-wavelength dyes. Support protocols are provided for calibration of individual dyes, which can be more problematic.