Dec 22, 2025
Stereotaxic virus injection and fiber optic probe implantation
- Asa Mackenzie1
- 1Lund University
Protocol Citation: Asa Mackenzie 2025. Stereotaxic virus injection and fiber optic probe implantation. protocols.io https://dx.doi.org/10.17504/protocols.io.eq2ly5dpqvx9/v1
Manuscript citation:
Serra GP, Guillaumin A, Vlcek B, Delgado-Zabalza L, Ricci A, Rubino E, Dumas S, Baufreton J, Georges F, Wallén-Mackenzie Å. A role for the subthalamic nucleus in aversive learning. Cell Rep. 2023 Nov 28;42(11):113328. doi: 10.1016/j.celrep.2023.113328. Epub 2023 Nov 4. PMID: 37925641.
License: This is an open access protocol distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited
Protocol status: Working
We use this protocol and it's working
Created: December 22, 2025
Last Modified: December 23, 2025
Protocol Integer ID: 235671
Keywords: ASAPCRN, fiber optic probe implantation stereotaxic virus injection, stereotaxic virus injection, fiber optic probe implantation
Funders Acknowledgements:
ASAP
Grant ID: ASAP-020600
Abstract
Stereotaxic virus injection and fiber optic probe implantation
Troubleshooting
Stereotaxic injections
Stereotaxic injections were performed in mice under isofluorane anesthesia (4% for induction and maintained at 1–1.5% air mix v/v).
After being placed in a stereotaxic apparatus, mice received subcutaneous injection of
analgesic and anti-inflammatory drugs (buprenorphine, 0.1 mg/kg and Carprofen, 5 mg/kg) as well as a local analgesic (lidocaine,
7 mg/kg) before the incision of the skin.
Mice were bilaterally injected in the STN/pSTN with a virus containing either a Cre-dependent
Channelrhodopsin (ChR2) construct coupled with an eYFP reporter (rAAV2/EF1a-DIO-hChR2(H134R)-eYFP), or only the eYFP reporter (rAAV2/EF1a-DIO-eYFP), respectively, 3.8 3 1012 virus molecules/mL and 4.6 3 1012 virus molecules/mL (viruses purchased
from UNC Vector Core, Chapel Hill, NC, USA) at the following mouse brain coordinates (from Paxinos and Franklin, 2013): anteroposterior (AP) = 1.90 mm, mediolateral (ML) = +/ 1.70 mm from the midline and 250 nL of virus was injected with a NanoFil syringe
(World Precision Instruments, Sarasota, FL, USA) at two dorsoventral levels (DV) = 4.65 mm and 4.25 mm from the dura matter at
100 nL.min-1.
Optic cannula implantation
Optic cannulas (Doric Lenses) were implanted directly after completion of virus injections
in mice preparated for behavior analysis.
Two skull screws were implanted in the skull to hold the optic cannula-cement-skull complex.
Optibond FL Prime 1 and 2 Adhesive (Kerr) were then applied and harden using Elipar LED polymerisation lamp (3M Espe).
Optic cannulas were implanted bilaterally above the STN (coordinates: AP = 1.90 mm, ML = +/1.70 mm from the midline DV = 4.30 mm) or the VP (coordinates: AP = +0.45 mm, ML = +/1.55 mm from the midline, DV = 4.00 mm), and fixed with dental cement.
1 mL of saline was injected subcutaneously at the end of the surgery.
For ex vivo electrophysiology experiments
Mice were injected into the LHb and the STN.
A glass pipette (30–50 mm tip
diameter) filled with rgAAV2-hSyn-DIO-EGFP and was lowered at the LHb coordinates: AP: 1.55 mm; ML: 0.50 mm; DV: 2.65 mm depth and 120 nL were injected using a picospritzer III (Parker).
Similarly, a glass pipette filled with AAV2-EF1a-DIOChR2(H134R)-mCherry was lowered at the STN coordinates: AP: 1.90 mm; ML: 1.60 mm; DV: 4.65/-4.30 mm and 2 times
120 nL were injected.
5 min after the last injection, the glass pipette was withdrawn
After stitching, mice were transferred to a
cage placed on a heating pad until waking up.