Jan 16, 2024
Striatal injection of 6-OHDA or Saline and post-operative care (Mendonça et al 2024)
- Marcelo D Mendonça1,2,3
- 1Champalimaud Research, Champalimaud Foundation, Lisbon 1400-038, Portugal;
- 2Champalimaud Clinical Centre, Champalimaud Foundation, Lisbon 1400-038, Portugal;
- 3NOVA Medical School | Faculdade de Ciências Médicas, Universidade Nova de Lisboa, Lisbon 1169-056, Portugal
Protocol Citation: Marcelo D Mendonça 2024. Striatal injection of 6-OHDA or Saline and post-operative care (Mendonça et al 2024). protocols.io https://dx.doi.org/10.17504/protocols.io.x54v9p4pzg3e/v1
Manuscript citation:
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: January 12, 2024
Last Modified: May 31, 2024
Protocol Integer ID: 93470
Keywords: ASAPCRN
Abstract
Striatal injection of 6-OHDA or Saline and post-operative care procedure from Mendonça et al 2024
Guidelines
All procedures were done in aseptic conditions.
Experiments were done on male mice-C57BL/6 or B6.SJL-Slc6a3tm1.1(Cre)Bkmn/J (Jax labs stock #006660).
Materials
-Stereotaxic surgery set up required (this protocol uses Kopf insturments)
-6-Hydroxydopamine hydrochloride (Sigma Aldrich AB, Sweden) was dissolved at a fixed concentration of 3.2 μg/μl free-base in 0.02% ice-cold ascorbic acid/saline and used within 2 h
Before start
Mice were kept in deep anesthesia using 1-3% isoflurane and oxygen (at 1L/min flow rate).
Stereotaxic Surgery
Stereotaxic Surgery
Place the anesthetized mouse in the stereotaxic apparatus then stabilize the head.
Make a skin incision to expose the skull.
Carefully remove any connective and muscle tissue from the exposed area.
Level the skull surface to be at less than 0.05mm by comparing the height of bregma and lambda, and also in medial-lateral directions.
6-OHDA or Saline Injection
6-OHDA or Saline Injection
Find the injection coordinates and mark area for drilling: dorsolateral striatum +0.5 mm anteroposterior and ±2.5 mm lateral from bregma and 3.0 mm deep
Drill Burr-holes in marked areas carefully.
Load glass pipette into the pipette holder (Nanojet II (Drummond Scientific)) and affix to the stereotaxic frame.
Set rate of injection to 4.6 nL per 5 seconds.
Inject 2 ul of 6-OHDA or saline.
After the injection finishes, leave the pipette in place for 10-15 minutes.
Post-operative Care
Post-operative Care
After surgery each animal had access to up 4 mg of food pellets/day (some mice were on food restriction pre-surgery)
Mice that showed weight loss were hand-fed (i.e. they were presented with the food while being held by the hands of the investigator) and DietGel Boost was placed in their boxed up to 4 days after surgery. In order to avoid competition for the food, weaker mice were placed in cages other than those containing unimpaired mice. The postoperative survival rate was 100%.