Biomedical Engineering Reference
In-Depth Information
Chapter 3
Wide-Field and Two-Photon Imaging of Brain Activity
with Voltage- and Calcium-Sensitive Dyes
Ryota Homma, Bradley J. Baker, Lei Jin, Olga Garaschuk, Arthur
Konnerth, Lawrence B. Cohen, Chun X. Bleau, Marco Canepari,
Maja Djurisic and Dejan Zecevic
Abstract
This chapter presents three examples of imaging brain activity with voltage- or calcium-sensitive dyes.
Because experimental measurements are limited by low sensitivity, the chapter then discusses the method-
ological aspects that are critical for optimal signal-to-noise ratio. Two of the examples use wide-field
(1-photon) imaging and the third uses two-photon scanning microscopy. These methods have relatively
high temporal resolution ranging from 10 to 10,000 Hz.
The three examples are the following: (1) Internally injected voltage-sensitive dye can be used to mon-
itor membrane potential in the dendrites of invertebrate and vertebrate neurons in in vitro preparations.
These experiments are directed at understanding how individual neurons convert the complex input
synaptic activity into the output spike train. (2) Recently developed methods for staining many indi-
vidual cells in the mammalian brain with calcium-sensitive dyes together with two-photon microscopy
made it possible to follow the spike activity of many neurons simultaneously while in vivo preparations
are responding to stimulation. (3) Calcium-sensitive dyes that are internalized into olfactory receptor
neurons in the nose will, after several days, be transported to the nerve terminals of these cells in the
olfactory bulb glomeruli. There, the population signals can be used as a measure of the input from the
nose to the bulb.
Three kinds of noise in measuring light intensity are discussed: (1) Shot noise from the random emis-
sion of photons from the preparation. (2) Extraneous (technical) noise from external sources. (3) Noise
that occurs in the absence of light, the dark noise. In addition, we briefly discuss the light sources, the
optics, and the detectors and cameras.
The commonly used organic voltage and ion sensitive dyes stain all of the cell types in the preparation
indiscriminately. A major effort is underway to find methods for staining individual cell types in the brain
selectively. Most of these efforts center around fluorescent protein activity sensors because transgenic
methods can be used to express them in individual cell types.
