Intrathecal Injections

Does any body have a protocol or advice for doing intrathecal injections in mice that doesn’t require surgery. Is there a way to inject directly into the lumbar enlargement (or other regions) just using an insulin syringe?

If so how reliable is it do people find?

Apologies if this has been discussed previously


Can you clarify what you’re trying to do? If by intrathecal, you mean into the CSF in a region where there isn’t any spinal cord (L4/5), then yes, people do this all the time. You just use an insulin syringe.

You just take 10 ul of injectate, find the interspace between L4/L5 based on landmarks (illiac crest), insert the needle between the interspace, and then inject. If you’re in the right place, you usually see a tail flick. You can do this under anesthesia, or, if you’re good and fast, even awake. @tonellor from @tberta lab does them awake, as does @zhzhj131421. I do it both ways.

Now lumbar enlargement indicates something different. That sounds to me like you want to inject directly into the substance of the spinal cord itself. That is an entirely different procedure. Is that what you’re trying to do? For that, you do need to expose the cord directly and use totally different tools and approaches.

The least invasive is this approach, which I use:

We’ve discussed this here before:

Yes, we do this all the time. The method was developed by George Wilcox. We used it in these papers:

Walwyn, W, W Chen, H Kim, A Minasyan, H Ennes, J A McRoberts, J C Marvizon. Sustained suppression of hyperalgesia during latent sensitization by µ, δ and κ opioid receptors and α2A adrenergic receptors - role of constitutive activity. J Neurosci 36: 204-221 (2016). PMC4701961

Severino, A, W Chen, J K Hakimian, B L Kieffer, C Gaveriaux-Ruff, W Walwyn, J C G Marvizon. Mu-opioid receptors in nociceptive afferents produce a sustained suppression of hyperalgesia in chronic pain. Pain 159: 1607-1620 (2018). PMC6053329

The right volume for an intrathecal injection in mice is 5 ul, with a 5 ul chaser. Otherwise, you risk the injectate spilling over into the brainstem.

The procedure is not easy because you have to get a feel of when the tip of the syringe is under the dura but still out of the spinal cord. The angle of the needle is crucial. The way my investigators train for this is by injecting morphine. If the injection is correct, the mouse will develop a Straub tail: the tail is erected vertically.

We do it without anesthesia. One person holds the mouse inside a cloth while the other does the injection.

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@achamess @marvizon

Apologies for the delay in replying. This is very helpful. I was thinking direct injection into the CSF region using an insulin syringe. Does this reliably result in diffusion or drug/virus etc into the spinal tissue? Based on what you described above regarding the lumbar enlargement procedure, I probably got my language a little mixed up, my goal primarily is to inject into the CSF and let the substance diffuse into the spinal tissue if possible.

Can you tell us more about what you’re trying to achieve? Are you delivering a drug or a virus? If a drug, I do think it penetrates. Will also depend on PK properties of drugs. Note that IT drugs will also affect DRGs.

If virus, from what I’ve seen, intrathecal virus does not produce high efficiency transduction of spinal cells.

Yes I’m delivering a drug to target specific neuronal sub types in spinal cord. I also want to target neurons in the DRG too so IT admin will be a benefit in this case too.


Great. That should work. Just it won’t go too many levels up. So you’ll be targeting mostly lumbar. Depends on drug. Some will definitely move up through CNS but will be most concentrated at lumbar DRG.