Long Term Modulation and Control of Neuronal Firing in Excitable Tissue Using Optogenetics - Foundations on Natural and Artificial Computation - page 266

waveform is to avoid damage to the electrodes and surrounding tissue. However,
even if the stimulus pulse is charge balanced an electrode may still be polarized
during delivery resulting in the subsequent damage of the electrode and tissue
[6]. Similarly an unwanted effect can arise through the electrolysis of surrounding
water with subsequent changes to pH and gas formation [7] [8]. Thus alternative
methods of cellular activation which avoid these issues are essential if chronic
stimulations are to be used in therapies. In this work we propose the use of
the light activated cation channel, Channelrhodopsin-2 (ChR2) as an alterna-
tive means to modulate and control neural activity. Long term photoactivation
of ChR2 expressing hippocampal cultures [9] and slices [10] has already been
demonstrated using LEDs and we propose this as a possible alternative method
of chronically activating neural tissue for therapeutic purposes circumventing
some of the issues faced by electrical stimulation.
2 Channelrhodopsin-2
Extracellular electrodes have limitations in their spatial resolution and intra-
cellular electrodes rely on mechanical stability, are invasive and ultimately lead
to cell death. However, the use of light activated channels permits to manip-
ulate neural activity within targeted cells with high temporal precision and in
a non-invasive manner [11]. Moreover, the use of light for chronic stimulation
can also be projected onto tissue with great spatial accuracy and can be used
to stimulate specific neuronal regions such as the axons or dendrites [12]. The
first attempts to convey photoexcitabilty in non photoresponsive neural cells was
accomplished by taking advantage of the invertebrate rather than the vertebrate
phototransduction system as it differs in the polarity of current it produces. In
invertebrates light mobilizes a class of G protein (Gq/11) which activates phos-
pholipase C (PLC). This results in non-specific cation channels in the plasma
membrane opening which in turn depolarize the cell [13]. The potential of this
invertebrate light cascade to depolarise other cells was recognized and it was
subsequently determined that the minimum protein requirement for conveying
phototransduction was the G protein couple receptor NinaE, a G protein aq
subunit and arrestin, otherwise referred to as chARGe [14]. However, even with
this cohort of proteins present, light responses are slow and small in magni-
tude and require the exogenous expression of three different genes. Since then,
two distant relatives of rhodopsin, Channelrhodopsin 1 and 2, were identified
in the unicellular green algae Chlamydomonas reinhadrtii . Both these channels
are gated by visible light and are thought to enable phototaxis by coupling to
specific transducers. ChR1 has been shown to be selective towards protons [15].
ChR2 differs in that it is a rapidly gated light-sensitive non-selective cation
channel with conductance for H + > Na + > K + > Ca 2+ . Within the algae the
photoreceptors normally are located in the cytoplasmic membrane and situated
in the eyespot region [16]. Amino acids 1-315 of ChR2 have been established
to convey photoresponsiveness [17]. As photocurrents are produced within <
50ms of a light flash in the algae it was understood that the photoreceptor and