The feasibility of gradient echo echo-planar imaging sequence (GE-EPI) for the accurate detection of stimulation-specific BOLD activation contrast in the rat brain at 16.4 T was investigated. An experimental protocol for longitudinal fMRI studies with extensive monitoring of the animal’s physiological status was employed. It was found that parameter optimized single-shot GE-EPI detects high quality images and is suitable for fMRI studies, provided motion effects during the timeseries can be compensated by data processing. The first specific BOLD activation maps at 16.4 T are presented and methodical details are discussed