Long Term Modulation and Control of Neuronal Firing in Excitable Tissue Using Optogenetics - Foundations on Natural and Artificial Computation - page 267

the channel form a complex or are one and the same. ChR2 is activated with
blue light (480nm) in which the all-trans form of retinal isomerizes into its cis
configuration opening the channel (Fig.1). Under normal conditions the inward
flood of cations depolarizes the cell resulting in the generation of action poten-
tials. The channel can return to the all-trans ground state without the necessity
for other enzymes, making it ideal for stimulating neurons with millisecond pre-
cision. Recovery from inactivation is also dependent on extracellular pH with it
being slower in high pH and faster in low pH. Pulses of H + given during the dark
periods accelerates the recovery of the channel from desensitization suggesting
that it is dependent also on the protonation of certain amino acids situated
on the extracellular side. It has been suggested that the amino acid Glu-123 is
the main culprit as it is predicted to be exposed on the extracellular side and
mutating it to Gln abolished peak photocurrents [17].
Fig. 1. Photostimulation of Channelrhodopsin-2. A: Illumination of ChR2 with blue
light (bandwidth 450-500nm) activates the channel allowing cations to pass through
resulting in depolarization of excitable cells. B: Action spectra of ChR2 indicating
maximum activation between 470 and 490nm in the blue spectrum.
Since its discovery, ChR2 has been successfully expressed in mammalian cells
[17] [18], Drosophila [19], C. Elegans [20] as well as in vivo in rodent inner
retinal cells [21], motor cortex [22], spinal chord and hindbrain [23] to name a
few. Other advantages of this tool are also the apparent lack of toxicity to cells
that have been genetically targeted with ChR2 [22]. Importantly; no significant
differences have been reported between the electrical properties of ChR2 +ive
and -ive cells confirming that the expression of protein does not affect the resting
properties and action potentials generation in transgenic mice [24]. As ChR2
is encoded by a single gene less than 1kb long it can be targeted with relative
ease to distinct cell populations through genetic techniques. ChR2 has been
tagged to various fluorophore markers through molecular engineering techniques
making it possible to visually identify neurons expressing the channel. One slight
disadvantage is that visualization of ChR2+ive neurons with GFP-YFP is not
possible without concurrently activating the channel.