Biomedical Engineering Reference
In-Depth Information
Key words:
Two-photon microscopy, one-photon (wide-field) microscopy, voltage-sensitive dyes,
calcium-sensitive dyes, neural networks.
1. Introduction
An optical measurement of brain activity using a molecular probe
can be beneficial in a variety of circumstances. An obvious advan-
tage of any imaging method is the possibility of simultaneous
measurements from many locations. This is especially important
in the study of the nervous system where many parts of an
individual cell, or many cells, or many regions, are simultaneously
active. In addition, optical recording offers the possibility of
recording from processes which are too small or fragile for
electrode recording.
An important advantage of optical methods for measuring
activity is their relatively high spatial resolution. In one of the
examples presented below, the resolution is on the order of a few
microns; in another, it is easy to distinguish the activity of indi-
vidual cells. However, optical recordings are generally limited to
areas of the brain that can be visualized and, in these areas, limited
to a recording depth of about 0.5 mm.
Using a molecular probe has advantages (and disadvantages)
over intrinsic signals. For optical imaging of membrane poten-
tial, the use of molecular probes has resulted in greatly increased
signal size in comparison to intrinsic signals. Furthermore, with
the introduction of protein activity sensors, there is now the pos-
sibility for cell type specificity. In addition, many of the molec-
ular probes are fast with a time resolution in microseconds
or milliseconds but others, especially the protein-based biosen-
sors, are substantially slower. Finally, there are novel probes
that are sensitive to various ions and metabolic intermediates.
With these advantages comes the responsibility for demonstrat-
ing that pharmacological and photodynamic effects are minimal
(see below).
1.1. Voltage-Sensitive
Dyes
Several different optical properties of membrane-bound organic
dyes are sensitive to membrane potential including fluorescence,
absorption, dichroism, birefringence, Forster resonance energy
transfer (FRET), non-linear second harmonic generation, and
resonance Raman absorption. Similarly, another group of organic
dyes are sensitive to ion concentrations. However, because
the vast majority of applications have involved fluorescence or
absorption measurements, these will be the only subjects of this
review.
