Endometriosis is an enigmatic painful disorder whose pain symptoms remain difficult to alleviate in large part because the disorder is defined by extrauteral endometrial growths whose contribution to pain is poorly understood. A rat model (ENDO) involves autotransplanting on abdominal arteries uterine segments that grow into vascularized cysts that become innervated with sensory and sympathetic fibers. ENDO rats exhibit vaginal hyperalgesia. We used behavioral, physiological, and immunohistochemical methods to test the hypothesis that cyst innervation contributes to the development of this hyperalgesia after transplant. Rudimentary sensory and sympathetic innervation appeared in the cysts at two weeks, sprouted further and more densely into the cyst wall by four weeks, and matured by six weeks post-transplant. Sensory fibers became abnormally functionally active between two and three weeks post-transplant, remaining active thereafter. Vaginal hyperalgesia became significant between four and five weeks post-transplant, and stabilized after six to eight weeks. Removing cysts before they acquired functional innervation prevented vaginal hyperalgesia from developing, whereas sham cyst removal did not. Thus, abnormally-active innervation of ectopic growths occurs before hyperalgesia develops, supporting the hypothesis. These findings suggest that painful endometriosis can be classified as a mixed inflammatory/neuropathic pain condition, which opens new avenues for pain relief. The findings also have implications beyond endometriosis by suggesting that functionality of any transplanted tissue can be influenced by the innervation it acquires.

Endometriosis (ENDO) is a painful disorder defined by extrauteral endometrial growths. It is created in rats by autotransplanting pieces of uterus (which form cysts), or, for shamENDO, fat (no cysts). ENDO induces vaginal hyperalgesia, likely via central sensitization. The severity of this hyperalgesia correlates with estradiol levels during the estrous cycle, suggesting the hyperalgesia is estradiolmodulated. If so, then hyperalgesic severity should track estradiol changes during reproductive senescence (estropause) when estradiol levels initially decrease, then increase. Using psychophysical methods to assess vaginal nociception, we found that the severity of ENDO-induced hyperalgesia paralleled estradiol changes during estropause: hyperalgesia first decreased, then returned. Furthermore, the return occurred regardless of the presence of the cysts (excised in some rats). This finding provides further support for ENDO's likely centrally-mediated effects. Additionally, the results suggest that elimination of estradiol via ovariectomy (OVX) should alleviate ENDO-induced hyperalgesia and estradiol replacement should restore it. However, in healthy and shamENDO rats, OVX produces a vaginal hyperalgesia that is alleviated by estradiol, likely via estradiol's peripheral influences on the vagina. Hence, we tested the hypothesis that OVX in ENDO rats would trigger a different type of vaginal hyperalgesia dependent on the loss of estradiol. We predicted that the opposing influences of estradiol on ENDO-and OVX-induced hyperalgesia would cancel each other. As predicted, OVX had no effect on ENDO-induced hyperalgesia and estradiol replacement alleviated it. These results suggest that, in intact rats, ENDO-induced vaginal hyperalgesia is exacerbated by estradiol, and that different mechanisms underlie ENDO-induced versus OVXinduced vaginal hyperalgesia.

Endometriosis is a painful disorder defined by extrauteral endometrial growths whose contribution to pain symptoms is poorly understood. Endometriosis is created in rats by autotransplanting on abdominal arteries pieces of either uterus (ENDO), which form cysts, or fat (shamENDO), which do not form cysts. ENDO, but not shamENDO induces vaginal hyperalgesia. We tested the hypothesis that the cysts are necessary to maintain vaginal hyperalgesia by assessing the effect of surgically removing them. Complete cyst removal eliminated ENDO-induced vaginal hyperalgesia up to four months post-operatively. Sham-cyst-removal in ENDO rats, in which cysts were not removed, or partial cyst-removal increased the ENDO-induced hyperalgesia. The decreases and increases both took three to six weeks to develop. Changes in ENDO-induced hyperalgesia did not occur in a control group of ENDO rats who had no surgery after ENDO. In a double-surgery control group, neither shamENDO surgery nor a subsequent sham surgery that mimicked "removal" of non-existent cysts influenced vaginal nociception. In a no-surgery control group, vaginal nociception remained stable for > six months. The increases in ENDO-induced hyperalgesia produced by the sham-cyst-removal surgery were smaller in proestrus than in other estrous stages. During the other stages (but not during proestrus), sympathetic innervation of the cysts increased. These results suggest that maintenance of ENDO-induced vaginal hyperalgesia requires continued presence of at least some ectopic endometrial tissue, and that surgical treatment that fails to remove ectopic endometrial tissue can exacerbate the hyperalgesia, possibly due in part to an increase in the cysts' sympathetic innervation.