Intratesticular Bradykinin Involvement in Rat Testicular Pain Models.

OBJECTIVES: To clarify the role of bradykinin in urogenital pain, we investigated bradykinin involvement in rat models of testicular pain. METHODS: Bradykinin (0.1, 0.3, 1, 3 and 10 mmol/L) or distilled water was injected into the testes of male Wistar rats, and induced pain behaviors in conscious rats were evaluated. The effect of pretreatment with bradykinin B2 receptor antagonist FK3657 on bradykinin-induced pain behavior was then examined. We also evaluated the analgesic effect of FK3657 in a rat acetic acid-induced testicular pain as well as changes in the intratesticular bradykinin concentration after testicular injection of acetic acid. RESULTS: An injection of bradykinin into the testes of conscious rats induced pain behaviors that were dose-proportionally reduced by prior administration of FK3657. In addition, FK3657 dose-dependently inhibited the pain responses induced by testicular injection of 1% acetic acid. An increase in intratesticular bradykinin concentration was detected after the testicular injection of 1% acetic acid. CONCLUSIONS: Here, we found that intratesticular bradykinin evokes pain behavior via stimulation of bradykinin B2 receptors and that intratesticular acetic acid injection induces intratesticular bradykinin synthesis, consequently leading to pain behavior. These findings suggest that the potential utility of bradykinin B2 receptor antagonists as a novel target for treating urogenital pain.

Induction of Bladder Overactivity by Nerve Growth Factor in Testes in Rats: Possible Neural Crosstalk Between the Testes and Urinary Bladder.

OBJECTIVES: To clarify the pathophysiological factor underlying neural crosstalk among pelvic organs, we investigated the possible role of nerve growth factor (NGF) in the neural crosstalk between the testes and urinary bladder. METHODS: Nerve growth factor (10, 30, and 100 µg/mL) or saline was injected into the testes of male Wistar rats. The change in bladder capacity via cystometry and duration of spontaneous scratching behavior induced by NGF in conscious rats was measured. The effects of pretreatment with capsaicin on NGF-induced changes in bladder capacity and behavior were examined. Further, we evaluated the effect of analgesics, indomethacin and morphine, and pretreatment with compound 48/80 on NGF-induced scratching behavior to elucidate the mechanism of the behavior. RESULTS: Injection of saline into the testes had no effect on bladder capacity or behavior. However, an injection of NGF (30 and 100 µg/mL) reduced bladder capacity, which was regarded as bladder overactivity, and evoked scratching behavior in a dose-dependent manner. Pretreatment with capsaicin inhibited NGF-induced bladder overactivity and scratching behavior. Neither indomethacin nor pretreatment with compound 48/80 affected the scratching behavior, but morphine inhibited the behavior. CONCLUSIONS: The present study provides evidence of a possible new role of NGF in the testes regarding the activation of testicular primary afferent neurons mediated by capsaicin-sensitive C-fibers, which evokes bladder overactivity via neural crosstalk between the testes and the urinary bladder as well as testicular pain.

A role for peripheral 5-HT2 receptors in serotonin-induced facilitation of the expulsion phase of ejaculation in male rats.

INTRODUCTION: The urethrogenital reflex (UGR) is used as a physiological animal model of the autonomic and somatic activity that accompanies ejaculatory-like reflexes (ELRs). Serotonin (5-HT) plays an important role in regulating ejaculation. AIM: To examine the effects of intraurethral 5-HT on ELRs and to examine the effects of various 5-HT receptor subtypes on the 5-HT-induced changes in the ELRs. METHODS: The effects of intraurethral infusion of 5-HT on ELRs were examined by monitoring the urethrogenital reflex in male rats. The effects of various 5-HT receptor-specific antagonists on the 5-HT-induced responses were examined. MAIN OUTCOME MEASURES: Main outcome measures were urethral pressure threshold required to elicit the UGR and bulbospongiosus activity or ELRs. RESULTS: Intraurethral infusion of 5-HT (10-1,000 µM) produced a dose-dependent facilitation of the UGR, i.e., decrease in threshold urethral perfusion pressure and an increase in number of ELRs. The 5-HT3 receptor antagonists tropisetron (1 and 3 mg/kg, i.v.) and ramosetron (0.1 and 1 mg/kg, i.v.), the 5-HT7 receptor antagonist SB269970 (3 mg/kg, i.v.), and the 5-HT1 A receptor antagonist WAY-100635 (1 mg/kg, i.v.) all failed to inhibit 5-HT-induced facilitation of the UGR. However, ritanserin (1 mg/kg, i.v.), a nonselective 5-HT2 receptor antagonist, and xylamidine (0.01-1 mg/kg, i.v.), a peripherally restricted nonselective 5-HT2 receptor antagonist, significantly inhibited both the decrease in urethral pressure threshold and the increase in number of ELRs induced by intraurethral infusion of 5-HT. CONCLUSION: These results suggest that in the male rat urethra, peripheral 5-HT2 receptors are involved in the 5-HT-induced facilitation of the expulsion phase of ejaculation.

Behavioral and urological evaluation of a testicular pain model.

OBJECTIVES: To develop an animal model of testicular pain to examine the hypothesis that neural crosstalk between testicular nociceptors and bladder reflex pathways may underlie bladder overactivity. In chronic pelvic pain disorders, neural crosstalk is thought to underlie referred pain and functional interaction in pelvic organs, and patients with chronic prostatitis/chronic pelvic pain syndrome (CP/CPPS) suffer from pain in multiple organs, including the testes and perineum, as well as increased urinary frequency. METHODS: In male Wistar rats, acetic acid was injected into the testes, and behaviors and bladder functions with conscious cystometry were examined. The effects of indomethacin and capsaicin pretreatment on both behaviors and bladder functional changes induced by acetic acid injection were examined. The weight of the testes and bladder after the testicular injection were measured. RESULTS: Injection of acetic acid (1% and 3%) induced pain behaviors and bladder overactivity proportional to the concentration. Indomethacin reduced, and capsaicin pretreatment almost completely abolished, both pain behavior and bladder overactivity induced by acetic acid injection. Administration of acetic acid increased testis weight and blanched the tissue, but no apparent changes were observed in the bladder. CONCLUSIONS: Injection of dilute acetic acid into the testes produces a reproducible testicular pain model involving testicular inflammation and activation of primary afferent C fibers and suggests a neural pathway for interaction between testicular pain and bladder overactivity. This study may provide a simple method to evaluate testicular pain, related bladder overactivity, and insight into the pathophysiology of bladder overactivity in patients with CP/CPPS.

Key roles of Phe1112 and Ser1115 in the pore-forming IIIS5-S6 linker of L-type Ca2+ channel alpha1C subunit (CaV 1.2) in binding of dihydropyridines and action of Ca2+ channel agonists.

Voltage-dependent L-type Ca2+ channels are modulated by the binding of Ca2+ channel antagonists and agonists to the pore-forming alpha1c subunit (CaV 1.2). We recently identified Ser1115 in IIIS5-S6 linker of alpha1C subunit as a critical determinant of the action of 1,4-dihydropyridine agonists. In this study, we applied alanine-scanning mutational analysis in IIIS5-S6 linker of rat brain alpha1C subunit (rbCII) to illustrate the role of pore-forming IIIS5-S6 linker in the action of Ca2+ channel modulators. Ca2+ channel currents through wild-type (rbCII) or mutated alpha1C subunits, transiently expressed in BHK6 cells with beta1a and alpha2/delta subunits, were analyzed. The replacement of Phe1112 by Ala (F1112A) significantly impaired the sensitivity to Ca2+ channel agonists (S)-(-)-Bay k 8644 and FPL-64176, and modestly to 1,4-dihydropyridine (DHP) antagonists. The low sensitivity of F1112A and S1115A to DHP antagonists was consistent with the reduced binding affinity for [3H](+)PN200-110. The replacement of Phe1112 by Tyr, but not by Ala, restored the long openings produced by FPL-64176, thus indicating the critical role of aromatic ring of Phe1112 in the Ca2+ channel agonist action. Interestingly, double-mutant Ca2+ channel (F1112A/S1115A) failed to discriminate between Ca2+ channel agonist (S)-(-)-1,4-dihydro-2,6-dimethyl-5-nitro-4-(2-[trifluoromethyl] phenyl)-3-pyridine carboxylic acid methyl ester (Bay k 8644) and antagonist (R)-(+)-Bay k 8644 and was blocked by the two enantiomers in an identical manner. These results indicate that both Phe1112 and Ser1115 in linker IIIS5-S6 are required for the action of Ca2+ channel agonists. A model of the DHP receptor is proposed to visualize possible interactions of Phe1112, Ser1115, and other DHP-sensing residues with a typical DHP ligand nifedipine.