AbstractInflammatory insult to the central nervous system (CNS) can lead to development of depression, and subsequently depression is the most frequent psychiatric comorbidity following ischemic stroke, often limiting recovery and rehabilitation in patients. The initiators of inflammatory pathways in the CNS are microglia activated in response to acute ischemic stress, and anti‐depressants have been shown to have anti‐inflammatory effects in the CNS, promoting neuronal survival following ischemic insult. We have previously shown that the selective serotonin reuptake inhibitors (SSRIs) fluoxetine and citalopram promote neuronal survival after oxygen‐glucose deprivation, an in vitro model of ischemia, by attenuating the release of glutamate and D‐serine from activated microglia. Interestingly, we found that fluoxetine‐treated microglial cultures contained fewer numbers of cells compared to other groups and hypothesized that fluoxetine and citalopram attenuated the release of glutamate and D‐serine by promoting the apoptosis of microglia. The present study aimed to test and compare antidepressants from three distinct classes (tricyclics, monoamine oxidase inhibitors, and SSRIs) on microglial apoptosis. Primary microglia were treated with 1 μg/mL lipopolysaccharide and/or 10 μM antidepressants, and various apoptotic markers were assayed. Fluoxetine and its metabolite norfluoxetine decreased protein levels in cell lysates, decreased cell viability of microglia, and increased the expression of the apoptotic marker cleaved‐caspase 3 in microglia. Live/dead nuclear staining also showed that fluoxetine‐ or norfluoxetine‐treated cultures contained greater numbers of dying microglial cells compared to vehicle‐treated cultures. Cultures treated with citalopram, phenelzine, or imipramine showed no evidence of inducing microglial apoptosis. Our results demonstrate that fluoxetine and norfluoxetine induce the apoptotic death of microglia, which may serve as a mechanism to attenuate the release of glutamate and D‐serine from activated microglia.Open Science BadgesThis article has received a badge for *Open Materials* because it provided all relevant information to reproduce the study in the manuscript. The complete Open Science Disclosure form for this article can be found at the end of the article. More information about the Open Practices badges can be found at https://cos.io/our-services/open-science-badges/. image