Electrophysiological and metabolic activities in brain tissue preparations maintained in vitro may be influenced by the persistent effect of anesthetic drugs utilized during tissue dissection. In order to clarify this issue, we studied elimination kinetics of the barbiturate thiopental from the brain parenchyma in the isolated guinea pig brain maintained in vitro, arterially perfused with a protein-free saline solution [M. de Curtis, G. Biella, C. Buccellati, G. Folco, Simultaneous investigation of the neuronal and vascular compartments in the guinea pig brain isolated in vitro, Brain Res. Protoc. 3 (1998) 21-28]. At the onset of anesthesia induced by a single i.p. injection of 125 mg/kg thiopental, the brain concentration of the drug, measured by high-performance liquid chromatographic assay, was 44.22+/-5.1 mg/L (mean+/-S.E.; n=7). After 30 min of arterial perfusion in vitro with a thiopental-free solution, the cerebral levels of the barbiturate decreased to 2.03+/-0.56 mg/L (n=3), and reached values close to zero within 1 h. No significant changes in thiopental elimination curve were observed when in vitro perfusion rate was either increased or decreased. The study demonstrates that thiopental is rapidly eliminated from the brain tissue with a mono-exponential kinetic. It can be concluded that barbiturate anesthesia utilized during brain dissection is not likely to influence activities recorded from the in vitro isolated brain preparation.