Mechanism of electroacupuncture at sensitized “Sanyinjiao” (SP6) in alleviating primary dysmenorrhea in rats based on Piezo1 ion channel

To investigate the mechanism of electroacupuncture (EA) at sensitized “Sanyinjiao” (SP6) in improving primary dysmenorrhea (PDM), with a focus on the role of Piezo1 protein in EA effects.

Sixty female rats were randomly divided into a blank group (n=10) and a model establishment group (n=50). Mechanical pain thresholds at the left SP6 acupoint were measured using a von Frey electronic algometer. According to the change rate of mechanical pain threshold, the model establishment group was further divided into model high-sensitization group, model low-sensitization group, EA high-sensitization group, and EA low-sensitization group (n=10 each). The PDM rat model was established by combined injection of estradiol benzoate and oxytocin. The two EA groups received EA at SP6 (50 Hz, dense wave, 20 min/d) for 5 consecutive days. Writhing responses were recorded to evaluate abdominal pain severity; uterine blood flow was observed by laser speckle imaging; uterine histopathology was examined by HE staining; mast cell degranulation rate at the SP6 region was detected by toluidine blue staining; ultrastructural changes of uterine tissue were observed by transmission electron microscopy; levels of prostaglandin E₂ (PGE₂), prostaglandin F₂α (PGF₂α) in serum and uterine homogenate, and serum tumor necrosis factor-α (TNF-α) were measured by ELISA; protein expressions of Piezo1, calcitonin gene-related peptide (CGRP), and Tryptase in the SP6 region, as well as Piezo1 in the uterus, were detected by Western blot; immunofluorescence was used to detect CGRP and Tryptase expressions at SP6; RT-qPCR was performed to measure mRNA expressions of Piezo1 and cyclooxygenase-2 (COX-2) in the uterus.

Compared with the blank group, the model high-sensitization and model low-sensitization groups showed increased writhing times, writhing scores and writhing latencies (P<0.01), decreased uterine blood perfusion (P<0.01), elevated uterine pathological scores (P<0.01), and increased mast cell degranulation rate (P<0.01). Serum and uterine PGF₂α contents, serum TNF-α content and PGF₂α/PGE₂ ratio were significantly increased (P<0.01), while serum and uterine PGE₂ contents were significantly decreased (P<0.01). Protein expressions of Piezo1, CGRP, Tryptase, and positive expressions of CGRP and Tryptase at the acupoint, mRNA and protein expressions of Piezo1, and COX-2 mRNA in the uterus were up-regulated (P<0.01), accompanied by ultrastructural damage in uterine tissue. Compared with the corresponding model groups, the writhing latencies were increased further, and the other indicators mentioned above were significantly reversed in the two EA groups (P<0.01, P<0.05). Moreover, the EA high-sensitization group exhibited more obvious improvements than the EA low-sensitization group (P<0.01).

EA at SP6 with different sensitization states effectively alleviates pain in PDM rats, with a better effect in the high-sensitization state. The mechanism is dependent on acupoint sensitization, which may involve regulating Piezo1 expression at the acupoint, inhibiting mast cell degranulation and neurogenic inflammation, and down-regulating the Piezo1/COX-2/PGs pathway in the uterus to reduce inflammatory factor release, thereby producing analgesic effects.