Viral Vector-Based Techniques for Optogenetic Modulation In Vivo - Viral Vector Approaches in Neurobiology and Brain Diseases - page 279

proteins to encode both light sensation and effector function [ 7 ].
A major advantage of microbial opsin-based optogenetic tools is
that they are essentially cofactor free. The endogenous presence of
the organic cofactor all-trans-retinal (ATR) in vertebrate tissues
enables the use of these genes as single-component tools, allowing
for spatially and temporally precise functional modulation of the
intact mammalian nervous system. Neurons transduced with
microbial opsin genes become sensitized to light with the physio-
logical effect of illumination dictated by the type of optogenetic
tool being used. For example, expression of channelrhodopsin-2
(ChR2), a light-gated cation channel [ 3 , 4 ], renders cells excitable
by light. Light-gated ion pumps such as halorhodopsin (e.g.,
NpHR) or archaerhodopsin (e.g., Arch) can be used to hyperpo-
larize and inhibit the electrical activity of neurons [ 8 , 9 ]. In recent
years, the optogenetic toolbox has greatly expanded and now
includes multiple tools for excitation, inhibition, and modulation
of signal-transduction pathways in neuronal and non-neuronal
cells [ 6 ]. We and others have reviewed the criteria for selection of
optogenetic tools for particular experiment types [ 7 , 10 - 12 ]. The
choice of opsin for a given experiment will depend on a multitude
of factors, including the desired physiological effect, the type of
neuron targeted, the spatiotemporal extent of modulation, and the
light-delivery system utilized [ 13 ]. The use of viral vectors, there-
fore, provides cost effective and fl exible means of optimizing
experiment-specifi c tools for each application.
When applied in the adult brain, optogenetics requires genetic
modifi cation of post-mitotic neurons to induce expression of the
light-gated channels or pumps. Genetically engineered viruses are
by far the most popular means of delivering optogenetic tools.
Lentiviral vectors (LV) [ 14 ] and adeno-associated viral vectors
(AAV) [ 15 ] have been widely used to introduce opsin genes into
mouse, rat, and primate neural tissues [ 11 ]. These vectors allow
high expression levels over long periods of time with little or no
reported adverse effects [ 15 ]. AAV-based expression vectors pos-
sess lower immunogenicity and offer the advantage of larger trans-
duced tissue volumes compared with LV due to their high viral
titers and diffusion properties. Additionally, AAV is considered
safer than LV as the currently available strains do not broadly inte-
grate into the host genome and are thus rated as biosafety level
(BSL) 2 agents. Both LV and AAV vectors can be used in conjunc-
tion with cell type-specifi c promoters (e.g., [ 14 , 16 , 17 ], see [ 10 ]
for more examples), and both vector families support pseudotyp-
ing techniques, which in principle enable a wide range of cell-type
tropisms and transduction mechanisms [ 18 , 19 ]. Finally, Cre-
dependent vectors, in which Cre recombination can activate the
expression of transgenes to achieve cell type-specifi c control [ 20 ,
21 ], are typically made with AAV-based vectors.