In our lab, we are trying to understand how early life adversity affects brain development and later risk for mental disorders, like depression and addiction. We are particularly interested in how microglia interacting with stress sensitive neurons in multiple brain regions may be the mechanism by which the circuit rewiring occurs. We previously found that this model for ELA provokes aberrant stress responses in adulthood.
This was caused by dysfunctional microglial synaptic pruning of excitatory synapses onto corticotropin releasing hormone expressing neurons in the paraventricular nucleus of the hypothalamus and thus hyperactivation of this stress responsive brain region. Although we know adverse early life experiences increase the risk of developing all kinds of health problems in humans, the limited bedding and nesting model allows us to prove the mechanisms for how these occurs in laboratory rodents. And there has never been a video protocol of this model published before.
The limited bedding and nesting model is very ecologically and translationally relevant because the low resource environment causes unpredictable maternal care, a phenomenon that can also be studied in humans. It also requires minimal experimental intervention and has been successfully implemented by multiple laboratories around the world. To begin, take a mesh divider cut according to the dimensions of a mouse cage, leaving an excess of three centimeters on the longer sides.
Fold the edges to create a platform that fits the cage edges perfectly with 2.5 centimeters above the floor. After setting up the cameras, check the video manager software interface for continuous recording. Next, pair one to four females with a single house male breeder mouse.
After 17 days of gestation or once visibly pregnant, separate the females into their own standard plexiglass cages. Place a cotton nestlet into the cage to allow the females to build a nest. After a couple of days, when the females give birth, record the date of birth.
On postnatal day two, transfer the pups into a clean new cage. Separate the males and the females into groups and count. Record the number of males and females in each litter.
To set up a control condition, fill a standard size mouse shoebox cage with about 220 grams of corn cob bedding. Add one standard square cotton nestlet. To help the mice acclimate to this unfamiliar environment, place one of their fecal pellets from their old cage in each corner of the new cage.
Transfer a small amount of used cotton nestlet from the previous cage over the new nestlet. To set up the LBN, or limited bedding and nesting condition, weigh 110 grams of corn cob bedding. Then, fill a standard shoebox cage with the corn cob and place the prepared mesh divider into the cage.
Finally, add half of a cotton nestlet square and repeat the same acclimation steps as in control cage. Now add the pups to their respective cages, placing them on top of the nestlet. Place the dam in the cage facing the pups.
Position a camera on a tripod in front of the cage with a clear side view of the dam and her pups. On postnatal day 10, transfer the dam first to standard caging conditions. Then, weigh the pups as they are transferred one by one to the standard cage.
Pups that were reared in LBN conditions were significantly smaller at postnatal day 10 than the control pups.
