RGS12 controls pain/mechanical sensitivity in a sex-dependent manner via regulating the estrogen/GPER-calcium signaling pathway.

Sex differences in pain sensitivity and response to analgesics are well-documented, with females often experiencing higher pain sensitivity and different responses to pain treatments compared to males. While Regulator of G-protein Signaling 12 (RGS12) is crucial in the central nervous system, its role in the peripheral nervous system and pain modulation remains unclear. Putative nociceptor-specific RGS12 conditional knockout (Rgs12-cKO) mice were generated by crossing Rgs12 floxed (Ctrl) mice with Trpv1-Cre transgenic mice. Behavioral assays, including the von Frey microfilament, hot plate, running wheel, and conditioned place preference, were conducted to assess pain sensitivity. Dorsal root ganglion (DRG) neurons from both sexes were dissected and cultured to examine neurite outgrowth and relative Ca²⁺ fluorescence (ΔF/F₀). Immunofluorescence, Ca2+ imaging, quantitative real-time PCR, and Western blot analyses were performed to investigate molecular and cellular mechanisms. Female Rgs12-cKO mice exhibited decreased pain sensitivity to both mechanical and thermal stimuli and reduced neurite outgrowth in DRG neurons. In contrast, male Rgs12-cKO mice did not show significant differences compared to Ctrl mice. The expression of estrogen receptors, particularly Gper, dramatically increased in Rgs12-cKO females but not males. GPER-specific agonist G1 and GPER-specific inhibitor CIMBA stimulation differentially increased KCl-induced relative Ca²⁺ fluorescence in DRG neurons from male and female mice, highlighting a sex-dependent response. This study demonstrates that RGS12 plays a critical role in modulating pain sensitivity in a sex-dependent manner. The interplay between RGS12 and estrogen/GPER signaling pathways in DRG neurons underscores the importance of RGS12 in considering sex as a critical biological variable in pain research. PERSPECTIVE: This study reveals RGS12 as a key regulator of sex-dependent pain modulation through estrogen/GPER-mediated calcium signaling in sensory neurons. The findings provide mechanistic insight into sex differences in nociception and suggest RGS12 as a potential therapeutic target for female-predominant pain disorders.