Dimethyltryptamine (DMT), an endogenous ligand of sigma-1 receptors (Sig-1Rs), acts against systemic hypoxia, but whether DMT may prevent cerebral ischemic injury is unexplored. Here global forebrain ischemia was created in anesthetized rats and aggravated with the induction of spreading depolarizations (SDs) and subse­ quent short hypoxia before reperfusion. Drugs (DMT, the selective Sig-1R agonist PRE-084, the Sig-1R antagonist NE-100, or the serotonin receptor antagonist asenapine) were administered intravenously alone or in combi­ nation while physiological variables and local field potential from the cerebral cortex was recorded. Neuro­ protection and the cellular localization of Sig-1R were evaluated with immunocytochemistry. Plasma and brain DMT content was measured by 2D-LC-HRMS/MS. The affinity of drugs for cerebral Sig-1R was evaluated with a radioligand binding assay. Both DMT and PRE-084 mitigated SDs, counteracted with NE-100. Further, DMT attenuated SD when co-administered with asenapine, compared to asenapine alone. DMT reduced the number of apoptotic and ferroptotic cells and supported astrocyte survival. The binding affinity of DMT to Sig-1R matched previously reported values. Sig-1Rs were associated with the perinuclear cytoplasm of neurons, astrocytes and microglia, and with glial processes. According to these data, DMT may be considered as adjuvant pharmaco­ logical therapy in the management of acute cerebral ischemia.