Identification of transient microglial cell colonies in the forebrain white matter of developing rats

Herein, we describe the existence of distinct colonies of transient microglial cells that reside in well-defined zones of the forebrain white matter. Rats, aged at postnatal day (P) 0, P2, P5, P7, P10, P15 or adult, were anaesthetised with halothane gas, and various neural centres were injected unilaterally with the tracer biotinylated Dextran. The neural centres injected were cingulate or sensorimotor cortices, ventral nuclei of the dorsal thalamus, and the pontine reticular formation of the brainstem. Rats were allowed to survive to various stages, from 4 hours to 21 days, after the injection. They were then anaesthetised with sodium pentobarbitone, and their brains were aldehyde-fixed and processed by using standard methods. The following is a description of what is seen after injections at P0, P2, P5, P7, P10; we saw no labelled cells (described below) in the rats injected at P15 or adult. From 2 to 21 days after an injection of dextran into the above-mentioned centres, labelled microglial cell colonies, identified by using double-labelling with anti-OX-6 or Griffonia simplicifolia (Bandeiraea; isolectin B4), were seen in small isolated zones in the forebrain white matter. These colonies were in the corpus callosum, the dorsal and ventral regions of the external capsule, and the internal capsule. A striking feature of these labelled microglial cell colonies was that they were seen on both sides of the brain. Thus, regardless of the location of the injection site in either the cortex, thalamus, or brainstem, the same microglial cell colonies were labelled with dextran in the forebrain white matter. After injections of different coloured fluorescent dextrans into the cortex and into the brainstem of the same animal, many double-labelled cells in each of the colonies were seen. From our short-term survival cases (4 hours to 1 day), a rather strict sequence or progression of labelling of the colonies across the white matter from the injection site was seen; in general, the microglial cell colonies closest to the injection site became labelled well before (about a day) those further away. These results lead us to suggest that the microglial cells in each colony become labelled after a slow diffusion of the tracer through the extracellular space from the injection site.