Jan 10, 2024
Version 1
Stereotaxic Injections and Implants V.1
- Jonathan Tang1,2,3
- 1Department of Neuroscience, Zuckerman Mind Brain Behavior Institute, Columbia University, New York, NY 10027, USA;
- 2Seattle Children’s Research Institute, Center for Integrative Brain Research, Seattle, WA, USA;
- 3University of Washington School of Medicine, Department of Pediatrics, Seattle, WA, USA
Protocol Citation: Jonathan Tang 2024. Stereotaxic Injections and Implants . protocols.io https://dx.doi.org/10.17504/protocols.io.8epv5xdw4g1b/v1Version created by Christine Weber-Schmidt
Manuscript citation:
Tang, J.C.Y., Paixao, V., Carvalho, F. et al. Dynamic behaviour restructuring mediates dopamine-dependent credit assignment. Nature (2023). https://doi.org/10.1038/s41586-023-06941-5
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 21, 2023
Last Modified: May 31, 2024
Protocol Integer ID: 92645
Keywords: ASAPCRN, mouse, brain, AAV
Funders Acknowledgement:
Life Sciences Research Fellowship
NINDS K99/R00 Award
Grant ID: 1K99NS112575
NIH
Grant ID: 5U19NS104649
Aligning Science Across Parkinson’s (ASAP)
Grant ID: ASAP-020551
Abstract
Methods for Stereotaxic injections and implants from Tang et al 2023.
Materials
Please see references for similar materials to what was used.
VTA AAV Viral Steriotaxic Injections
VTA AAV Viral Steriotaxic Injections
Using a standard mouse surgical suite (see references for example), 750nl of rAAV.EF1a.DIO.hChR2(H134R).eYFP or rAAV.EF1a.DIO.eYFP (3-4 x 10^12 vg/ml,
AAV5, University of North Carolina Vector Core; 1-2 x 10^13 vg/ml, AAV1,
Addgene, 27056-AAV1 and 20298-AAV1) were injected into each hemisphere of the
VTA of 3–4-month-old DAT-Cre mice.
For viral injections, the coordinates are AP - 3.52 mm, ML - +/- 0.35 mm, DV – 4.3
mm. Injections were made at 0.2 Hz pulses. Each pulse injects 4.6 nl volume.
Injected needles were kept in place in the injection site for ~15 minutes before withdrawal.
Dual Optic Fiber Cannula Implant
Dual Optic Fiber Cannula Implant
For each mouse, a dual optic fiber cannula (200/240 μm diameter, 6 mm length, 0.7 mm center-to-center FLT, 0.22 NA; Doric, DFC_200/240-0.22_6mm_DF0.7_FLT) was placed 200 μm above
the injection site and fixed to the skull.
Next, a 4-position receptacle connector (Harwin Inc., M52-5000445) was fixed anteriorly to the dual optic fiber cannula, with its posterior edge set at -0.6 mm.
Skull implants are then fixed with dental cement.
A 4-position connector (Harwin Inc., M52-040023V0445) with pins removed from one end was used to cap the receptacle connector.
Photometry Experiments
Photometry Experiments
The conditions used for VTA injections and implants were as above, and 3–5-month-oldDAT-Cre males were used.
Additionally, 1 μl and 500 nl of AAV9-hSyn-GRAB-rDA1m (2 x 10^13 vg/ml; RRID:Addgene_140556) were injected into the dorsal striatum (AP 0.5 mm, ML +2.1 (right), DV 2.3 (from brain surface)) and ventral striatum (AP 1.15mm, ML +1.65 (right),DV 4.2 (from Bregma)), respectively.
For photometry fiber implants, mono fiberoptic cannula were used (400/430 μm diameter, 4 mm length (dorsal striatum) and 6 mm length(ventral striatum), 0.37 NA, 1.25 diameter ferrule, flat; Doric, MFC_400/430-0.37_6mm_MF1.25_FLT (ventral striatum) and MFC_400/430-0.37_4mm_MF1.25_FLT
(dorsal striatum)). Implants were inserted at a 22 degrees angle.
Protocol references
Paxinos, George & Franklin, K. B. J. The mouse brain in stereotaxic coordinates / George
1431 Paxinos, Keith B.J. Franklin. (2001).
Allen Institute for Brain Science 2023. Stereotaxic Injection by Nanoject Protocol. protocols.iohttps://dx.doi.org/10.17504/protocols.io.bp2l6nr7kgqe/v4Version created by Allen Institute