Abstract: The objective of this study was to evaluate the effects of tramadol, a centrally acting synthetic opioid analgesic, on glutamate metabolism, without inducing pain. Male adult Wistar rats weighing 150 ± 20 g were used in the study. An effective dose of tramadol was injected subcutaneously into the rats at 0, 24, and 48 hours, and the changes in the levels of activities of glutamate dehydrogenase (GDH), glutamine synthetase (GS), glutaminase, aspartate (AAT) and alanine (AlAT) aminotransferases, and glutamine content, were recorded at 3, 6, 12, 24, 48 and 72 hours in different areas of the rat brain, viz. cerebral cortex, cerebellum, pons-medulla, hippocampus and hypothalamus. Aminotransferase activities were examined in serum also. Following the first administration of tramadol at zero hours, GDH activity showed positive deviations in all areas except pons-medulla, GS activity showed positive deviations in all areas, glutaminase activity showed negative deviations in all areas except pons-medulla, and glutamine content showed positive deviations in all areas except hippocampus, up to 12 hours. Aminotransferase activities showed differential deviations, with increases in some areas and decreases in the others. Peak deviations in all cases occurred either at 3 or at 6 hours. All parameters reverted towards near control levels by 24 hours. Following the second and third injections of tramadol at 24 and 48 hours respectively, the parameters recorded deviations at 48 and 72 hours that were slightly reverting from those at 24 hours. The results indicate differential tissue responses from different areas of the brain to the administered analgesic. Further, while the administration of opioids could affect the glutamate release vis-à-vis reuptake and the activation of N-methyl-D-aspartate (NMDA) receptors, these changes could presumably be associated with alterations in the levels of other parameters related to glutamate metabolism. This could be another facet of the analgesic effects of tramadol.