The Involvement Of Endothelin Pathway In Chronic Psychological Stress-Induced Interstitial Cystitis Through C-Fiber Afferents

Introductions and objective: Interstitial cystitis/bladder pain syndrome(IC/BPS)is a poorly understood chronic disorder characterized by bladder-related pain,which is commonly seen in middle-aged women.Chronic psychological stress plays a role in the exacerbation and development of IC/BPS via unclear mechanisms.Endothelin-1(ET-1)was found to induce detrusor overactivity and regulate the sensitivity of peripheral nervous system.The aim of this study was to investigate the role of endothelin 1(ET-1)and its receptors(ETAR)in the development of WAS-induced bladder dysfunction.Methods:1.22 Wistar-Kyoto rats were divided into chronic water avoidance stress group(WAS,n=11)and control group(Sham,n=11).WAS rats were exposed to chronic(10 days)water avoidance,while Sham rats did not apply stress.After conducting urodynamic studies under saline infusion,the bladder was emptied and performed ET-1 infusion.Cystometrogram(CMG)and visceromotor response(VMR)electromyograph were recoded simultaneously.Several parameters were used to evalutate urodynamic signals results,including pressure threshold(PT),intercontraction duration(ICI),contraction duration(CD),bladder capacity,the ratio of VMR threshold/intravesical maximal bladder pressure(VMR threshold/IVPmax).VMR was analyzed by calculating VMR duaration,VMR amplitude and area under the curve(AUC).2.After capsaicin pretreatment,22 Wistar-Kyoto rats were divided into chronic water avoidance stress group(WAS-Cap)and control group(Sham-Cap).For the above four groups(WAS,Sham,WAS-Cap,Sham-Cap),cystometrograms(CMG)were obtained with visceromotor response(VMR)simultaneously during intravesical saline and ET-1 infusion.After integrating the data of WAS and Sham group,the urodynamic parameters were analyzed to evaluate the role of ETAR in WAS-induced bladder dysfunction through capsaicin-sensitive C-fibers.3.After CMG and VMR signals of the above four groups were collected,the bladder,L6 DRGs and S1 DRGs were harvested,endothelin receptor type A(ETAR)expression was examined by Western blot and q RT-PCR in different tissues.Doublelabeling immunofluorescence(IF)staining was to identify the location of ETAR and CGRP,respectively.Toluidine blue staining was used to determine the number of mast cells in bladder tissue,aimed at checking the bladder inflammation.Results: 1.During saline infusion,WAS rats elicited significant decreases in PT(8.6±1.5 vs 10.9±3.6 cm H2 O,P<0.05)and in the ratio of VMR threshold/ IVPmax(45.2±7.6 vs 57.1±16.8%,P<0.05),and significant increases in VMR duration(21.7±9.6 vs 14.9±5.8 s,P<0.05)and AUC(0.6±0.3 vs 0.3±0.2 m V-s,P<0.05).WAS could induce bladder hypersensitivity and enhance nociceptive responses.Lower bladder pressure can stimulate micturition reaction in rats,and the activity intensity of pain reaction is further increased.ET-1 infusion induced similar alternations in WAS rats,but further significantly diminished the pressure to trigger PT(7.9±1.1 vs 10.2±3.5 cm H2 O,P<0.05)and VMR(30.2±22.8vs 14.8±6.3s,P<0.05),together with a more forceful(1.0±0.7 vs0.4±0.3m V-second,P<0.05)and longer VMR,indicating that endothelin receptor activation can enhance WAS-induced bladder hyperalgesia.2.Whether saline infusion or ET-1 infusion,after capsaicin pretreatment,no significant difference in the parameters of CMG and VMR electromyography between Sham-Cap group and WAS-Cap group was found.However,when compared with WAS and Sham,capsaicin treatment(WAS-Cap and Sham-Cap)can increase the ratio of VMR threshold / IVPmax,indicating that capsaicin pretreatment can reduce the sensitivity of bladder.The sole effect of WAS exposure or ET-1administration on the micturition reflex could be suppressed by capsaicin pretreatment.The activation of ETAR enhanced WAS-induced bladder hyperalgesia through C-fibers afferents.3.WAS exposure likely increased expressions of ETAR in the bladder and DRGs,while capsaicin pretreatment possibly antagonized them.IF staining indicated the colocalization of ETAR and calcitonin gene-related peptides in both bladder and DRGs.After toluidine blue staining,the mast cell count in WAS group was higher than Sham group,and capsaicin pretreatment could reduce this difference.Conclusion: WAS could induce bladder hypersensitivity and hyperalgesia,which may possibly relate to sensitized C-fibers and mechanoreceptors.The activation of ETAR may enhance WAS-induced bladder hypersensitivity and hyperalgesia through capsaicin-sensitive Cfiber afferents.WAS related bladder inflammation may lead to the activation of endothelin pathway.Targeting the endothelin pathway may have therapeutic value for IC/BPS.