Brush stimulation in SNI mice to look at c-Fos - Best Practices?

I have some SNI mice. I want to stimulate the sural territory of the ipsilateral hind paw with a stimulus to induce c-Fos. Mechanical hypersensitivity/allodynia is a behavioral feature of this model. We see it both with VF and dynamic brush.

Groups would be:

  1. Naive
  2. SNI
  3. SNI + Brush
  4. Naive + Brush

As in this paper:

I’ve seen in previous literature that people will do a prolonged brush stimulus of 5-10 minutes. The question is, what is the best way to do this? All methods have pros/cons.

  1. Under isoflurane anesthesia: Put out the mouse, brush the paw back and forth for 10 mins. Pro: Mouse is well controlled by the anesthesia: Cons: Isoflurane itself activates some neurons and may suppress some relevant pain-related neurons.
  2. Awake, using a restraint: As in this paper: Touch and tactile neuropathic pain sensitivity are set by corticospinal projections | Nature. Pros: Mouse is better controlled and you can do focused, fairly uniform stimulation of the hindpaw. Cons: The stress of the restraint itself may induce stress-induced analgesia and may cause Fos activation in stress neurons that are not pain-related. Of course, one can see this with proper controls but it’d be good to avoid. If you were to restrain, how much acclimatization to the restraint would you do? 2-3 days?
  3. Awake, in the von frey rack: In this case, we would try to stimulate the mouse’s paw as it is awake in it’s cage. Pros: No concern for restraint-related stress. Cons: The mice invariably will move and try to avoid the stimulus. This will result in variable number of strokes between mice. Also, the mice might be stressed by being chased around, which makes it not much better than restrained. Also, it is more difficult to precisely stimulate the sural territory if the mouse is moving. And the moving around could induce Fos in motor circuits that are unrelated to pain.

None of these is perfect. If you were doing this experiment, or you were the reviewer, which would you pick?

@tberta @thicunha @liz @jmogil @gcorder @marvizon @adgaudet @RPSeal @alex_naka

Some responses on Twitter:

In this paper we did the brushing under isoflurane anesthesia.

Marvizon, J C, W Chen, W Fu, B K Taylor. Neuropeptide Y release in the rat spinal cord measured with Y1 receptor internalization is increased after nerve injury. Neuropharmacology 158: 107732 (2019).

But we had one group of rats that underwent hindpaw clamp, and that had to be done under anesthesia for ethical reasons. So all groups of rats were anesthetized so they could be compared.

You could use a beaker upside-down to restrain the mouse to a small space in the von Frey cage. The mouse would not be completely immobilized, so hopefully that would reduce the stress. You can try different sizes of beakers to find one that lets the mouse move, but not escape the brush.

Juan Carlos

1 Like

Thank you Juan Carlos!