the center of the field, showing that microglia-depleted mice did not have increased anxiety in this task. Additionally, no changes in the distance traveled or velocity were seen, indicating that there were no locomotor differences between groups. Microglial elimination throughout the CNS had no effects on learning and memory, as determined via acquisition and probe trial of the Barnes maze or on locomotion, as tested with the accelerating rotarod. Given these results in mice depleted of microglia for 21 days, we then set out to determine if further long-term depletion of microglia would alter cognition or behavior, as perhaps purchase LGX818 deficits would take longer than 3 weeks to manifest. To that end, wild-type mice were treated with PLX3397 or vehicle for 2 months, and then cognition and behavior assessed. A third group was administered the cholinergic antagonist scopolamine on testing days to induce cognitive deficits as a positive control. With long-term microglial elimination, no changes in elevated plus maze were observed. Likewise, no changes were seen in open field or accelerating rotarod. Intriguingly, training on the Barnes maze revealed that microglia-depleted mice were able to learn the task significantly better than microglia-intact animals, as shown by shorter escape latencies on days 2 and 3 of training, as well as an overall reduction in average escape latency across all training days. Mice treated with scopolamine were unable to learn the task, as evidenced by an inability to escape the maze more quickly on subsequent days. No differences were found in the probe trial. We then performed contextual fear conditioning, an additional hippocampal-dependent learning and memory task. No significant differences were found between microglia-depleted mice and microglia-intact mice, while mice treated with scopolamine performed significantly worse, and therefore showed a cognitive deficit as Neuron. Author manuscript; available in PMC 2015 April 16. NIH-PA Author Manuscript NIH-PA Author Manuscript NIH-PA Author Manuscript Elmore et al. Page 6 expected. Thus, mice depleted of microglia for either 21 days or 2 months show no deficits in learning, memory, motor function, or behavior, and surprisingly, mice chronically depleted of microglia showed some evidence of enhanced learning. Immune profiling of the microglia-depleted brain To explore how the microglia-depleted brain responds to immune challenges, we treated 2 month-old wild-type mice for 7 days with PLX3397 to deplete their microglia. We then administered either PBS or LPS and sacrificed the animals 6 hours later. mRNA was extracted from whole brains, converted to cDNA, and then analyzed against a panel of 86 immune-related genes. Overall, depletion of microglia leads to robust reductions in the expression of many inflammatory genes, including TNF- and other cytokines. Microglia-expressed genes are also robustly reduced, including CD4, CD68, CD86, H2-Eb1, and PTPRC, reinforcing the finding that microglia are absent from these treated brains. Responses to LPS are 
dampened for many genes, but chemokine responses are mixed. Additionally, these results demonstrate that microglial elimination is not accompanied by PubMed ID:http://www.ncbi.nlm.nih.gov/pubmed/19845972 an inflammatory response by the remaining cells in the CNS, which is an important feature of the approach to microglial-depletion shown here. Rapid restoration of CNS microglia following drug removal 18 month-old wild-type mice were treated with PLX3397 for 28 days to eliminate mic
