Background: Microglia activation is an early response in ischemic brain injury, which initiates neuroinflammation. Microglia express Sigma-1 receptors (Sig-1R) at the mitochondria-associated membranes (MAM) of the endoplasmic reticulum(ER), which have been implicated in cellular stress responses. Here we set out to evaluate whether the Sig-1R agonist N,N-dimethyltryptamine (DMT) modulates microglia activation. Materials and methods: Isolated microglia cultures were prepared from the cortex of neonatal Sprague Dawley rats. On day 6, the cells were treated with lipopolysaccharide (LPS;20ng/ml) or DMT (Sig-1R agonist) alone (5-10-20-50μM), or in combination with LPS for 24 h. Microglial activation was evaluated by Iba1 immunolabeling (transformation index - TI) and Western blot analysis, and the visualization of phagocytotic activity with fluorescent microbeads. Results: Microglia displayed decreased area, perimeter and TI in response to the LPS challenge, indicative of amoeboid transformation and activation. When the LPS-challenged cell cultures were treated with DMT, significantly more ramified cells were seen. Increased Iba1 signal intensity in Western blot analysis confirmed microglial activation due to LPS treatment, which was increased particularly by DMT. The proportion of phagocytic microglia in the culture was increased by the addition of LPS, which was counterbalanced by DMT treatment.Conclusion: DMT reduced microglia activation effectively. We assume that DMT exerted its protective effect by modifying the stress response of the ER and the Ca2+ homeostasis of microglia. The assumption is substantiated by previous findings that microglia activation is associated with increased intracellular Ca2+ concentration, and Sig-1R in the MAM regulates Ca2+ transport between the ER and mitochondria.