Benzodiazepines are one of the most frequently prescribed drugs due to their anxiolytic properties. The aim of this study was to evaluate the effects of diazepam on lipopolysaccharide-induced peritoneal acute inflammatory responses. Swiss mice were treated with diazepam in a single dose of 1 or 10 mg/kg- subcutaneously 1 h before an intraperitoneal injection of lipopolysaccharide or sterile saline solution. The mice were killed 16 h after and the cells were washed from the peritoneal cavity to determine the total number of cells and the mononuclear and polimorfonuclear subpopulations, as well as the TNF-alpha activity and percentage of spread macrophages. Our results showed that the diazepam treatment (1 and 10 mg/kg) induced a significant reduction in the LPS-induced macrophage stimulation and TNF-α activity. Diazepam (10 mg/kg) also reduced the inflammatory cellular migration when compared to the control. It can be concluded that the diazepam treatment in a single dose is able to influence the inflammatory cellular influx, macrophage stimulation and TNF-α activity in the acute inflammatory response in mice, having possible implications on the anti-infectious response efficiency.Departamento de Morfologia e Patologia Universidade Federal de São CarlosDepartamento de Análises Clínicas Faculdade de Ciências Farmacêuticas Universidade Estadual Paulista Júlio de Mesquita FilhoDepartamento de Morfologia e Patologia Centro de Ciências Biológicas e da Saúde Universidade Federal de São Carlos, Rodovia Washington Luiz, km 235, 13565-905 - São Carlos - SPDepartamento de Análises Clínicas Faculdade de Ciências Farmacêuticas Universidade Estadual Paulista Júlio de Mesquita Filho