Chronic stress has been shown to have long-term effects on functional hemispheric asymmetries in both humans and non-human species. The short-term effects of acute stress exposure on functional hemispheric asymmetries are less well investigated. It has been suggested that acute stress can affect functional hemispheric asymmetries by modulating inhibitory function of the corpus callosum, the white matter pathway that connects the two hemispheres. On the molecular level, this modulation may be caused by a stress-related increase in cortisol, a major stress hormone. Therefore, it was the aim of the present study to investigate the acute effects of cortisol on functional hemispheric asymmetries. Overall, 60 participants were tested after administration of 20 mg hydrocortisone or a placebo tablet in a cross-over design. Both times, a verbal and an emotional dichotic listening task to assess language and emotional lateralization, as well as a Banich-Belger task to assess interhemispheric integration were applied. Lateralization quotients were determined for both reaction times and correctly identified syllables in both dichotic listening tasks. In the Banich-Belger task, across-field advantages were determined to quantify interhemispheric integration. While we could replicate previously reported findings for these tasks in the placebo session, we could not detect any differences in asymmetry between hydrocortisone and placebo treatment. This partially corroborates the results of a previous study we performed using social stress to induce cortisol increases. This suggests that an increase in cortisol does not influence dichotic listening performance on a behavioral level. As other studies reported an effect of stress hormones on functional hemispheric asymmetries on a neuro-functional level, future research using neuronal imaging methods would be helpful in the characterization of the relation of hemispheric asymmetries and stress hormones.