I want to (bilaterally) lesion VNAB in mice, however all studies I could find were done in rats, using 6-OHDA. I performed ink test to find VNAB coordinates in mice and then proceeded with pilot experiment, following a protocol from a study where they used 6-OHDA to lesion striatum area (PMID: 22370630).
Unfortunately, mice don't handle this surgery well. Most of them died, including vehicle injected group. One thing that is noticeably different, when I compare to injecting NTS, is that there is a lot more bleeding in most of them. We do give them saline but it doesn't seem to help.
I would greatly appreciate any feedback or suggestions if anyone has experience with this. Are there any alternative ways to lesion VNAB in mice or would it be better to try it in rats instead?
Hi Jovana,
from my experience with injections into the brain parenchyma of mice the key to a successfull procedure is to inject as slowly as possible (max. 1µl/min) to not increase the intracranial pressure that may cause the bleeding events. I hope this is a first starting point for optimization.
Hi Luisa,
Thank you for your response. I did it 0,1 µl/min. Bleeding usually occurs after drilling, but a lot of them died even when there was not much bleeding. We established Pargyline dose that was tolerated well and we have also tried a bit more frontal injection site to try to avoid injuring breathing center but still reach VNAB fibers, however no changes.
Hi Jovana Zlatkovic
I have a few queries,
* At what stage are you observing mortality? Immediately post surgery? Or from the next days?
* Have you tried the surgery with only vehicle injection? Was mortality observed there? which species of mice are you using?
* Initially during standardization of models, there is a high amount of mortality observed in stereotaxic surgery as there are lot of steps and the surgery itself needs some practice (especially the drilling part, slow injection, slow removal, suture and then comes the post-operative part and pre-operative (pain killers, antibiotics).
* In mice it is even more difficult, so if you are a beginner, I would advise you to use SD Rats instead of mice.
I have attached link to a protocol, please check.
1. Article Survivable Stereotaxic Surgery in Rodents
Thanks,
Hi Samir!
1. Around 80% die immediately, they start waking up from anesthesia but can't seem to recover all the way through. Few that were able to survive died within 1-3 days.
2. Yes, high mortality rate in that group as well. Just to add, we have tested pargyline (6-OHDA enhancer) doses as well and found the tolerable dose so we excluded that as a factor. C57BL/J.
3. We did take time to practice this procedure and I have a lot of experience in mice, I feel quite confident in the technical part now. We have been very gentle and careful, ensuring best possible chance of survival by reducing mechanical damage, as well as stimulating recovery as much as possible. The only thing we could add for post-operative care is some lighter form of food, instead of HFD. We usually leave some for them on the cage floor. But for that we still need them to be make it back to the animal room :D
*I found some studies mentioning that unilateral 6-OHDA injections in mice are better for reducing mortality (PMID: 34768962), however, this wouldn't lead to complete lesion. Considering this option at the moment as a last attempt to do this experiment in mice.
And thank you, I really appreciate that you took time to respond to my question!