Imidacloprid is a systematic neonicotinoid widely used to combat piercing-sucking insects; however, neonicotinoids, despite having low effects on vertebrates, showed high adverse effects on honeybees (Apis mellifera L.). Glutathione-S-transferases (GST) are an important constituent of the defense system in detoxifying invading chemicals. This work examines the effects of imidacloprid on GST activity in vivo and in vitro conditions. Results revealed that there are two responses of GST activity toward IMI treatment; first, stimulation of enzyme activity to combat and detoxify the insecticide. Second, direct inhibition of GST, which is confirmed by the in vitro inhibition with IC50 887.42 ppm. At a short exposure time to IMI (2 hours) at an IMI concentration of 0.35 ppm, the enzyme was stimulated up to 113% while increasing exposure time or IMI concentration, the inhibition effect dominates.