Neurourology and Urodynamics 31:148–155 (2012) Effects of TRPV4 Cation Channel Activation on the Primary Bladder Afferent Activities of the Rat Naoki Aizawa, 1 Jean-Jacques Wyndaele, 2 Yukio Homma, 3 and Yasuhiko Igawa 1 * 1 Department of Continence Medicine, The University of Tokyo Graduate School of Medicine, Tokyo, Japan 2 Faculty of Medicine, Department of Urology, University of Antwerp, Antwerp, Belgium 3 Department of Urology, The University of Tokyo Graduate School of Medicine, Tokyo, Japan Aims: Transient receptor potential vanilloid 4 (TRPV4) may affect afferent pathways innervating the bladder. We investigated the effects of GSK1016790A (GSK) and RN1734, a TRPV4 agonist and antagonist, respectively, and P2X- purinoceptor antagonists (TNP-ATP and PPADS) on cystometry (CMG), and the effect of GSK on single afferent fiber activities (SAAs) of the rat bladder and its relationship with capsaicin (Cap)-sensitivity. Methods: Conscious female Sprague–Dawley rats were used for CMG measurements. In SAA measurements, under urethane anesthesia, SAA was identified by electrical stimulation of the pelvic nerve and by bladder distention. Cystometric parameters were mea- sured before and after intravesical drug instillation. In SAA measurements, response with saline instillation served as baseline. Then, GSK was instilled three times, and finally Cap was instilled to investigate the relationship with Cap- sensitivity. Results: Intravesical GSK-instillation transiently decreased bladder capacity and voided volume, which were counteracted by RN1734, TNP-ATP, and PPADS. In SAA measurements, Ad-fibers (n ¼ 7) were not affected by either GSK or Cap. Based on the Cap-sensitivity, C-fibers could be divided into two subtypes: Cap-insensitive (n ¼ 14) and Cap-sensitive (n ¼ 8). In the Cap-insensitive C-fibers, GSK significantly increased the SAAs during the first instil- lation, but the increase attenuated with time, whereas GSK did not significantly affect the Cap-sensitive C-fibers. Conclusions: The present results suggest that activation of TRPV4 in the bladder, probably urothelium, facilitates the micturition reflex by activation of the mechanosensitive, Cap-insensitive C-fibers of the primary bladder afferents in rats. Neurourol. Urodynam. 31:148–155, 2012. ß 2011 Wiley Periodicals, Inc. Key words: afferent nerves; desensitization; rats; transient receptor potential (TRP); urinary bladder INTRODUCTION The transient receptor potential vanilloid subfamily (TRPV) contains six proteins in mammals, and they are commonly divided into two subgroups based on sequence homology, functional similarities, and Ca 2þ -selectivity; TRPV1–V4 and V5/6. 1 The subgroup of TRPV1–V4 members are weakly Ca 2þ - selective cation channels, modulated by various intracellular signals and activated by temperature. 2,3 Expression of the TRPV1, V2, and V4 has been reported in human and rat/mouse urinary bladders. 4–10 Moreover, TRPV1 has been exploited clin- ically to desensitize bladder afferents and reduce bladder over- activity. 11 On the other hand, TRPV4 is sensitive to osmotic and mechanical stimuli, such as cell stretching or fluid flow. 12 Some previous studies show that TRPV4 may be modulated by calmodulin (CaM) and adenosine triphosphate (ATP), C-termi- nal CaM binding potentiating the current and Ca 2þ -dependent CaM binding to the N-terminal desensitizing the current. 13–16 Several researchers reported that TRPV4 is implicated in the regulation of urothelial ATP release that modulates the sensi- tivity of bladder afferent nerves. 7,8,17–19 In our previous study, the activation of the bladder mechanosensitive afferents in- duced by exogenous ATP was mainly through capsaicin (Cap)- insensitive (probably TRPV1-independent) C-fibers in the rat. 20 Therefore, it is conceivable that TRPV1 and TRPV4 have a role in the bladder afferent transduction via a different pathway. In the present study, we focused on the afferent function of TRPV4, and investigated the effects of intravesical administra- tion of GSK1016790A (GSK), a TRPV4 agonist, which has at least 300-fold greater potency for activating TRPV4 than 4a- PDD, 21 on single fiber activities of the primary bladder mecha- nosensitive afferent nerves. MATERIALS AND METHODS Animals Forty-eight adult female Sprague–Dawley rats weighing 180–234 g were used. The rats were maintained under stan- dard laboratory conditions with a 12:12 h light:dark cycle, and free access to food pellets and tap water. The protocol was approved by Animal Ethics Committees of The University of Tokyo Graduate School of Medicine and in line with NIH guidelines for the care and use of experimental animals. Cystometry (CMG) Measurements Rats were anesthetized with 30 mg/kg intraperitoneal pen- tobarbital sodium. A polyethylene catheter (Clay-Adams PE- 50; Parsippany, NJ) was inserted in the bladder through the dome, and secured. After the operation, each rat was housed single in a cage. Lori Birder led the review process. Conflict of interest: none. Grant sponsor: Ministry of Education, Culture, Sport, Science and Technology of the Japanese Government; Grant numbers: 40159588. 80595257. *Correspondence to: Yasuhiko Igawa, M.D., Ph.D., Professor and Chairman, Depart- ment of Continence Medicine, The University of Tokyo Graduate School of Medi- cine, 7-3-1, Hongo, Bunkyo-ku, Tokyo 113-8655, Japan. E-mail: yigawa-jua@umin.ac.jp Received 13 June 2011; Accepted 29 July 2011 Published online 28 October 2011 in Wiley Online Library (wileyonlinelibrary.com). DOI 10.1002/nau.21212 ß 2011 Wiley Periodicals, Inc.