To assess whether premenopausal and postmenopausal vestibulodynia have different histologic features.We conducted a retrospective analysis of vestibulectomy specimens from 21 women with postmenopausal vestibulodynia and compared them with 88 premenopausal patients (42 primary, 46 secondary). Women with primary vestibulodynia experienced pain at first introital touch and women with secondary vestibulodynia experienced pain after an interval of painless intercourse. Clinical records established the type of vestibulodynia, duration of symptoms, and hormone status. Tissues were stained for inflammation, nerves, mast cells, estrogen receptor α, and progesterone receptor. Histologic findings in the postmenopausal patients were compared with primary and secondary premenopausal patients using proportional odds logistic regression and analysis of variance.Seventy-one percent (15/21) of postmenopausal women reported vestibular dyspareunia related to a drop in estrogen either with menopause (13/21) or previously, postpartum (2/21). Eighty-six percent (18/21) of postmenopausal patients were using local or systemic estrogen but pain persisted. Compared with premenopausal primary and secondary vestibular biopsies, postmenopausal tissues had more lymphocytes (unadjusted odds ratio [OR] 9.0, 95% confidence interval [CI] 2.8-33.3; adjusted OR for parity and duration of symptoms 9.1, 95% CI 2.6-31.9; unadjusted OR 6.2, 95% CI 1.9-20.0; adjusted OR 6.6, 95% CI 2.0-21.9, respectively) and mast cells (mean 36 compared with 28 and 36 compared with 26, respectively). There was significantly less neural hyperplasia and progesterone receptor expression in postmenopausal biopsies compared with primary cases but less progesterone receptor and similar neural hyperplasia compared with premenopausal secondary cases. Estrogen receptor α did not vary among groups.Premenopausal and postmenopausal vestibulodynia share histologic features of neurogenic inflammation but differ strikingly in degree. When estrogen supplement does not alleviate symptoms of postmenopausal dyspareunia, vestibulodynia should be considered.: II.