Functional imaging studies typically give prominence to positive responses. However, negative changes that accompany activation-induced positive responses are not yet clearly understood. The authors investigated the origin of sustained negative CBF responses that accompanied positive CBF changes. Measurements were made in the rat somatosensory cortex in response to whisker stimulation using laser-Doppler imaging. Flux images indicative of CBF were obtained at rest and during whisker stimulation with a spatial resolution of 200 microm. Large and intermediate blood vessels in the cortical surface exhibiting high flux values were clearly resolved. This greatly reduced the contamination of the tissue pixel volume with macroscopic blood vessels. Regions that responded positively to whisker stimulation were from areas with intermediate to low baseline flux and distinctly away from high flux areas. Stimulation-induced change in signal intensity was the largest in pixels with low baseline flux, presumably from tissue and microvessels. Simultaneously, a sustained decrease in signal intensity was observed in regions with high baseline flux values. The temporal coherence, macrovascular origin, lesser trial-to trial variability, and complete absence of the negative CBF response in the microvascular regions suggest that it may be purely hemodynamic in nature.