Objectives: Opioids are widely used as potent pain relievers. However repeated use can induce not only series of side effects such as drowsiness, respiratory depression, but also increases in the dosage and patient's demand for analgesics due to the weakened effect resulting from the development of the tolerance. Acupuncture, one of the representative traditional therapies of the Eastern Asia, has been gaining interests in diverse fields including pain and drug addiction. In the previous studies, acupuncture showed to inhibit the morphine addiction. The present study examined if acupuncture could prevent opioid tolerance and a possible neuronal mechanism was also investigated. Methods: Male Sprague-Dawley rats weighing 270-300 g were subjected. Morphine hydrochloride (1.5 mg/ml), Naloxone (0.1 µg/0.25 µl), picrotoxin (0.1 µg/0.25 µl), 2-hydroxysaclofen (0.1 µg/0.25 µl) were used. For surgery rats were anesthetized by sodium pentobarbital (50 mg/kg), Their right jugular vein was cannulated for repeated intravenous morphine administration and they were placed in a stereotaxic device, an access hole was made by drill through the skull for the RMTg (AP: -6.9, ML: ±0.9, DV: -7.0) and the guide cannula was placed at the destination. 0.2 mL of saline was infused into the catheter once a day during recovery period, and heparin (30 U/mL) was contained to maintain patency. Acupuncture was treated bilaterally at SI5 or TE9 for 1 min. After vertical insertion to a depth of 2-3 mm needle, needle was twisted in both directions for 2 s at a frequency of 2 times per second when entering and exiting the acupoint. Before the experiment starts, rats were acclimated to the Von Frey test environment for 30 min. Mechanical stimulation was given with a von Frey filament (Stoelting, Wood Dale, IL), which was sequentially applied (increased or decreased) to the right behind plantar for a total of six applications. The rat's pain perception was recorded over 14 days after the baseline establishment for 2 days during which saline was given instead morphine. Results: According to the dose response, the optimal morphine dose was decided as 1.5 mg/kg. During 14 days, morphine induced less body weight gaining and the analgesic effect was diminished in the later half following the repeated administration. However, this diminish of analgesic effect was prevented by the acupuncture at SI5 but not at TE9. Also, the blood BDNF level increased later in the control group, indicating the decline of morphine analgesic effect, however was maintained at the similar level in the SI5 group, indicating that the tolerance of morphine analgesic effect was inhibited by SI5 acupuncture. In addition, this inhibitory effect of SI5 was blocked by the pretreatments of the Naloxone and Picrotoxin, but not by the 2-hydroxysaclofen. Conclusion: Results suggest that acupuncture at SI5 can be an option to prevent the tolerance of the morphine’s analgesic effect and this inhibition is mediated by the opioid receptor and GABAA receptor pathway in the RMTg.