Acclimation Protocols for Nociceptive Assays - How long and how often?

There is general agreement that animals (mice) need to be acclimated to the testing room and apparatus before performing tests. But the duration of acclimation (both intraday and interday) is variable between labs.

What are people doing for:

  • Number of acclimation days before testing?
  • How long acclimation per day?
  • Duration of acclimation during the day of testing
  1. Von Frey
  2. Hargreaves/Cold Plantar
  3. Rotarod

@runDRG @CandlerPaige @tberta @thicunha @gcorder @ShanTan

3 days of 30-min habituation (general approach) and then 30-min on test day, but it varies slightly depending on the apparatus. We also have fully switched over to reverse-light, and now doing more operant tasks; here we use separate “home-away-from-home” cages living in the test room on a seperate rack, each cage here is for an individual mouse where we keep the bedding the same across hab and the experimental length (many days). We sometimes do vF still, and no longer do Hargreaves… the question we still ask ourselves is why do so much hab? Traditionally relfex assays need the animal to be still (light phase) to poke it, but the animal is technically “sleepy” which is a different internal state than wakefullness in the dark (so what are we modeling?), and now we’re disrupting sleep cycles further possibly causing more stress (major confound). Also repeat hab is likely a stressor as well in some cases, such as an animal sitting on a 1-2 foot floating platform that was continually handled by a large primate.

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We do von Frey and Hargreaves on mice.

For von Frey, 3-4 days of acclimation before testing and usually 3 hours each day. At the end of three hours, most of the mice would be sitting still, either grooming or just sitting there. Then on testing days, I would still acclimate them until they are mostly sitting still. Most times it would be somewhere between 1.5-2 hours, but sometimes still 3 hours.

I have tried to sit in the room with the mice for the the last hour of acclimation, in order to avoid their reaction to me as a novel object entering the room.

We usually do Hargreaves after von Frey in the same room on the same day. So we also acclimate the mice to hargreaves once or twice before testing days. 30-45 min acclimation on Hargreaves at 30 degrees Celsius usually is sufficient. Occasionally, in the very beginning of acclimation phase, it could take longer than an hour for mice to settle on Hargreaves. On testing days, I gave the mice 30 min for acclimating on Hargreaves.

They do fall asleep on the 30 degrees platform. To avoid stimulating them in sleep, I try to make some tiny noises or movements to see if their ears or whiskers move. If they don’t move, then I wouldn’t stimulate them.

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