The adverse side effects of opioids, especially antinociceptive tolerance, limit their clinical application. A recent study reported that platelet-derived growth factor receptor beta (PDGFR beta) blockage selectively inhibited morphine tolerance. Autophagy has been reported to contribute to the cellular and behavioral responses to morphine. However, little is known about the relationship between PDGFR beta and autophagy in the mechanisms of morphine tolerance. In this study, rats were intrathecally administered with morphine twice daily for 7 days to induce antinociceptive tolerance, which was evaluated using a tail-flick latency test. By administration autophagy inhibitor 3-Methyladenine, PDGFR beta inhibitor imatinib, p38 mitogen-activated protein kinase (MAPK) inhibitor SB203580 hydrochloride and minocycline hydrochloride, western blot, immunofluorescence, and transmission electron microscopy techniques were used to elucidate the roles of PDGFR beta, autophagy, and related signaling pathways in morphine tolerance. This study demonstrated for the first time that spinal PDGFR beta in microglia promotes autophagy in gamma-aminobutyric acid (GABA) interneurons through activating p38 MAPK pathway during the development of morphine tolerance, which suggest a potential strategy for preventing the development of morphine tolerance clinically, thereby improving the use of opioids in pain management.