Mice and viral expression in CeA regions
Adult male C57BL/6 mice (6-8 week-old, 20-25 g, Vital River Inc.,
Beijing, China) were used in this study. Mice were housed 6 per cage in
the animal facility and free to food and water. Surgeries including
virus and nanoheaters injection were conducted using a standard
stereotaxic apparatus under aseptic conditions. The mice were
anesthetized through an intraperitoneal injection of pentobarbital
sodium (40 mg/kg). Before in vivo magnetothermal neurostimulation, AAV5
virus encoding TRPV1His-mCherry was packaged and
injected into CeA region for getting
TRPV1His-expressing mice. The stereotaxic coordinate
for the CeA was as follows: – 1.25 AP, – 2.75 ML, – 4.30 DV. A small
hole was drilled in the skull at the coordinate of CeA and a 32-gauge
needle was lowered into the hole. 300 nL of AAV5 virus were injected
into the CeA region at 0.1 μL/min using a microsyrine pump. After lifted
by 0.1 mm, the syringe was remained for 10 minutes within the brain
before slow withdrawal. At least 4 weeks postviral expression, the
anti-His-FVIOs in PBS (2 mg/mL) was injected into the same CeA region.
The skin tissue was closed with sutures. TRPV1-expressing and
FVIOs-loaded mice were prehoused in a closed box with a diameter of 12
cm under a reverse 12-hour light/dark cycle for 3 days as a habituation
stage. Animal husbandry and all experimental manipulation of the animals
were performed with the approval of the Animal Care and Use Committee of
the Chinese Academy of Military Medical Science and Northwest
University.