BackgroundLow‐intensity pulsed ultrasound (LIPUS) combined with acupoint can promote gastric motility of diabetic rats. The switch of gastric smooth muscle cell (GSMCs) phenotype was related to the diabetes‐induced gastric dysfunction, but the mechanism is not clearly elucidated. This study was aimed at exploring the underlying mechanism of LIPUS stimulation application in diabetic gastroparesis rats.MethodsIn this study, Sprague–Dawley male rats were divided into three groups: control group (CON), diabetic gastroparesis group (DGP), and LIPUS‐treated group (LIPUS). LIPUS irradiation was performed bilaterally at ST36 for 20 min per day for 4 weeks. The gastric emptying rate was measured by ultrasound examination. Contraction ability of GSMCs was assessed by muscle strip experiment. The expression of related proteins or mRNAs including α‐SMA, SM22α, MHC, RhoA, Rock2, p‐MYPT1, MYPT1, p‐MLC, MLC, MALAT1, miR‐449a, and DLL1 was detected by different methods such as western blotting, RT‐qPCR, immunohistochemistry, and immunofluorescence staining, as appropriate.Key Results(a) LIPUS stimulation at ST36 could improve the gastric motility dysfunction of diabetic rats. (b) LIPUS increased RhoA, Rock2, p‐MYPT1, and p‐MLC expression level. (c) MALAT1 and DLL1 contents were decreased, but the level of miR‐449a was increased in the LIPUS group.Conclusions & InferencesLIPUS may affect the contractile marker expression of gastric smooth muscle through the RhoA/Rock and MALAT1/miR‐449a/DLL1 pathway to ameliorate DGP.