Preparation and optical characteristics of hemoglobin-free isolated perfused rat head in situ

Using a new cannulation method, which completely prevented interruption of circulation to brain tissue during surgical operations, we could prepare a functionally intact hemoglobin-free isolated perfused rat head in situ. The frequency distribution (8-25 Hz) and amplitude of a spontaneous electroencephalogram, EEG (50 microV), were kept within the normal ranges for up to 4 h after the perfusion was started. Our preparation gave characteristic flash-evoked EEG responses through the eyes. The administration of bicuculline elicited an epileptic seizure similar to that in normal rats. Near-infrared difference spectra of the rat head obtained on an aerobic to anaerobic transition were similar to those of cytochrome oxidase in isolated mitochondria as well as in the purified state. Under the aerobic perfusion conditions, both heme a+a3 and copper in cytochrome oxidase were in the fully oxidized states, showing that normoxia was maintained in the brain tissue. The redox behaviors of these two chromophores in the brain were identical to those observed in isolated mitochondria. The usefulness of brain perfusion for bridging in vitro and in vivo studies is well documented.