Biology Reference
In-Depth Information
disadvantage of this viral system is its adverse effect on cell and
animal health over a course of days and weeks.
Viral vector-based expression is a versatile and widely used method
for delivering optogenetic tools in the brain. However, animal-to-
animal variation in the effi ciency of viral transduction is also a
potential major source of variability in optogenetic experiments.
Although electrophysiology and immunohistochemistry can allow
researchers to correlate the behavioral or physiological effect with
the effi ciency of genetic expression of the optogenetic tool, optimi-
zation of the method for viral transduction and light delivery to
the targeted cells can reduce this inherent variability and allow
more robust, reproducible results. In the protocol below, we focus
on the procedures developed for direct intracerebral delivery of
high-titer virus for expression of optogenetic tools in defi ned cell
populations. We also describe the procedure for chronic implanta-
tion of fi beroptic connectors for robust, reproducible optical mod-
ulation of virally transduced neurons in vivo.
1.3 Summary
2
Protocol
Iodine antiseptic (e.g., Betadine)
Ethanol, 70 %
Sterile saline
Sterile distilled, deionized water (DDW)
Anesthetics and analgesics
Lubricant eye ointment
Optional: hydrogen peroxide (H
2
O
2
)
2.1 Reagents
Surgical tools (e.g., curved sharp tip tweezers; fi ne, spring scissors;
surgical scalpel; dull-tip forceps; wide tipped bulldog clamps)
Hot bead tool sterilizer (
see
Note 1
)
Dissection stereomicroscope (Fig.
1
-a1)
Small animal stereotaxic apparatus with cannula holder, non-
rupture tip ear-bars, tooth bar, rodent face mask for isofl urane
circulation (Fig.
1
-a2, a3)
Temperature-controlled heating pad (Fig.
1
-a4, a5)
Isofl urane vaporizer (LEI Medical; Fig.
1
-a6)
Isofl urane induction box (Fig.
1
-a7)
Small heat lamp
Small hair trimmer
2.2 Equipment
2.2.1 Virus Injection
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