Subcutaneous B16 melanoma impairs intrinsic pressure generation and relaxation of the heart, which are not restored by short-term voluntary exercise in mice.

The aim of this study was to investigate whether subcutaneous melanoma impairs intrinsic cardiac function and hypoxia tolerance in mice. In addition, it was investigated whether these changes could be prevented by voluntary wheel-running exercise. The roles of different molecular pathways were also analyzed. Male mice (C57Bl/6NCrl) were divided into unexercised tumor-free group, unexercised melanoma group, and exercised melanoma group. The experiment lasted 2.7 ± 0.1 wk (determined by the tumor size) after which the heart function was measured in different oxygen levels ex vivo using Langendorff method. All the melanoma mice had lower pressure amplitude (50.3%), rate of pressure production (54.1%), and decline (52.5%) in hearts ex vivo when compared with tumor-free group. There were no functional differences between the two melanoma groups. All the groups had similar weight changes, heart weights, cardiomyocyte sizes, levels of Ca2+ channels, energy metabolism enzyme activities, lipid peroxidation, and reactive oxygen species in their cardiac tissue homogenates. However, all the melanoma mice had 7.4% lower superoxidase dismutase activity compared with the control animals, which might reduce the ability of the heart to react to changes in oxidative stress. The exercising melanoma group had a 28.6% higher average heart capillary density compared with the unexercised melanoma group. Short-term wheel running did not affect the tumor growth. In conclusion, subcutaneous melanoma seems to impair intrinsic heart function even before cachexia, and these functional alterations were not caused by any of the measured molecular markers. Short-term voluntary wheel-running exercise was insufficient to alleviate the intrinsic cardiac impairments caused by melanoma.NEW & NOTEWORTHY Melanoma has been shown to induce cardiac atrophy and impair cardiac function in vivo, however, it has not been investigated how melanoma affects the intrinsic heart function. Here, we showed that subcutaneous melanoma can impair intrinsic heart function in noncachectic mice, decreasing the heart's pressure production and relaxation. In addition, we investigated whether short-term voluntary wheel-running exercise could attenuate the impairment of intrinsic cardiac function. However, our results do not seem to support this hypothesis.

The Translational Role of Animal Models for Estrogen-Related Functional Bladder Outlet Obstruction and Prostatic Inflammation.

The prevalence of LUTS and prostatic diseases increases with age both in humans and companion animals, suggesting that a common underlying cause of these conditions may be age-associated alterations in the balance of sex hormones. The symptoms are present with different and variable micturition dysfunctions and can be assigned to different clinical conditions including bladder outlet obstruction (BOO). LUTS may also be linked to chronic non-bacterial prostatitis/chronic pelvic pain syndrome (CP/CPPS), but the relationship between these conditions is unknown. This review summarizes the preclinical data that supports a role for excessive estrogen action in the development of obstructive voiding and nonbacterial prostatic inflammation. Preclinical studies that are emphasized in this review have unequivocally indicated that estrogens can induce functional and structural changes resembling those seen in human diseases. Recognizing excessive estrogen action as a possible hormonal basis for the effects observed at multiple sites in the LUT may inspire the development of innovative treatment options for human and animal patients with LUTS associated with functional BOO and CP/CPPS.

Short-term exercise affects cardiac function ex vivo partially via changes in calcium channel levels, without influencing hypoxia sensitivity.

Exercise is known to improve cardiac recovery following coronary occlusion. However, whether short-term exercise can improve cardiac function and hypoxia tolerance ex vivo independent of reperfusion injury and the possible role of calcium channels in improved hypoxia tolerance remains unknown. Therefore, in the current study, heart function was measured ex vivo using the Langendorff method at different oxygen levels after a 4-week voluntary wheel-running regimen in trained and untrained male mice (C57Bl/6NCrl). The levels of cardiac Ca2+-channels: L-type Ca2+-channel (CACNA1C), ryanodine receptor (RyR-2), sarco(endo)plasmic reticulum Ca2+-ATPase (SERCA2), and sodium-calcium exchanger were measured using western blot. Trained mice displayed lower cardiac afterload pressure generation capacity (rate and amplitude), but unaltered hypoxia tolerance when compared to untrained mice with similar heart rates. The level of CACNA1C positively correlated with the pressure generation rate and amplitude. Furthermore, the CACNA1C-RYR-2 ratio also positively correlated with the pressure generation rate. While the 4-week training period was not enough to alter the intrinsic cardiac hypoxia tolerance, interestingly it decreased pressure generation capacity and slowed pressure decreasing capacity in the mouse hearts ex vivo. This reduction in pressure generation rate could be linked to the level of channel proteins in sarcolemmal Ca2+-cycling in trained mice. However, the Ca2+-channel levels did not differ significantly between the groups, and thus, the level of calcium channels cannot fully explain all the functional alterations, despite the detected correlations. Therefore, additional studies are warranted to reveal further mechanisms that contribute to the reduced intrinsic capacity for pressure production in trained mouse hearts.

Characterization a model of prostatic diseases and obstructive voiding induced by sex hormone imbalance in the Wistar and Noble rats.

BACKGROUND: Chronic nonbacterial prostatitis associated with lower urinary tract symptoms (LUTS) is a prevalent condition in men. One potential pathophysiological factor is change in sex hormone, testosterone and estrogen, balance. Inflammation, cancer and obstructive voiding has been induced in the Noble rat strain by altering levels of sex hormones. We evaluated if imbalance of sex hormones could induce comparable diseases also in a less estrogen sensitive Wistar strain rats. METHODS: Subcutaneous testosterone (830 µg/day) and 17ß-estradiol (83 µg/day) hormone pellets were used in male Wistar and Noble strain rats to induce prostatic diseases. The rats were followed for 13 and 18 weeks. Urodynamical measurements were performed at the end of the study under anesthesia. Prostates were collected for further histological analysis. A panel of cytokines were measured from collected serum samples. RESULTS: Noble rats exhibited stromal and glandular inflammation after 13 weeks that progressed into more severe forms after 18 weeks of hormonal treatment. CD68-positive macrophages were observed in the stromal areas and inside the inflamed acini. CD163-positive macrophages were present in the stromal compartment but absent inside inflammatory foci or prostate acini. Thirteen-week hormonal treatment in Noble rats induced obstructive voiding, which progressed to urinary retention after 18-weeks treatment. In the Wistar rats 18-week treatment was comparable to the 13-week-treated Noble rats judged by progression of prostatic inflammation, being also evident for obstructive voiding. Incidence of PIN-like lesions and carcinomas in the periurethal area in Noble rats were high (100%) but lower (57%) and with smaller lesions in Wistar rats. Serum cytokines leptin, CCL5, and VEGF concentrations showed a decrease in the hormone-treated rats compared to placebo-treated rats. CONCLUSIONS: Prostate inflammation and obstructive voiding developed also in the Wistar rats but more slowly than in Noble rats. Male non-castrated Wistar strain rats may thus be suitable to use in studies of pathophysiology and hormone-dependent prostate inflammation and obstructive voiding.

Intravesical treatment with cis-urocanic acid improves bladder function in rat model of acute bladder inflammation.

AIMS: The aim was to study the effect of intravesically instilled cis-urocanic acid (cis-UCA) on bladder function in an experimental rat model of acute bladder inflammation. Hyaluronic acid (HA) was used as a comparator compound. METHODS: Bladder irritation was induced in female rats by intravesical hydrochloric acid (HCl) infusion. Vehicle, 0.5% HA, or 2% cis-UCA solutions were infused intravesically twice a day for three consequent days. On the fourth day, urodynamical measurements were performed, the animals were sacrificed, and the bladders were removed for histopathological assessment. RESULTS: HCl treatment caused significant impairment of bladder function indicated by decreased micturition intervals and voided urine volumes and induced severe voiding dysfunction observed as occurrence of overflow incontinence. These functional changes were accompanied by increased bladder weight, hemorrhage, and infiltration of inflammatory cells into the urothelium. Intravesical cis-UCA treatment recovered bladder function by significantly prolonging the micturition interval, increasing the voided volume, and reducing the occurrence of overflow incontinence. All these changes were comparable to the effects of HA. CONCLUSIONS: Intravesical administration of cis-UCA was able to partially recover bladder function impaired by chemical irritation. Cis-UCA may offer a novel intravesical treatment option in some inflammatory conditions of the bladder. Neurourol. Urodynam. 35:786-791, 2016. © 2015 Wiley Periodicals, Inc.

Potentiation of Glibenclamide Hypoglycaemia in Mice by MK-467, a Peripherally Acting Alpha2-Adrenoceptor Antagonist.

Pharmacological antagonism and genetic depletion of pancreatic α2A-adrenoceptors increase insulin secretion in mice and enhance the insulinotropic action of glibenclamide, a representative of the sulphonylurea class of insulin secretagogues used in the therapy of type 2 diabetes. Antagonism of α2-adrenoceptors in the central nervous system (CNS) causes tachycardia and hypertension, making generalized α2-adrenoceptor blockade unfavourable for clinical use despite its potential to decrease blood glucose levels. The purpose of this study was to test the acute effects of the peripherally acting α2-adrenoceptor antagonist MK-467 alone and in combination with glibenclamide in non-diabetic C57BL/6N mice. Cardiovascular safety was assessed in freely moving mice with radiotelemetry. Dose-dependent decreases in blood glucose and increases in plasma insulin concentrations were seen with the combination of MK-467 and glibenclamide; the combinations were much more potent than glibenclamide or MK-467 alone. Furthermore, MK-467 had no effect on mean arterial pressure or heart rate in freely moving mice and did not prevent the centrally mediated hypotensive effect of the α2-adrenoceptor agonist medetomidine. Thus, peripheral blockade of α2-adrenoceptors does not evoke the same cardiovascular adverse effects as antagonism of CNS α2-adrenoceptors. The current results indicate that the combined use of small doses of a peripherally acting α2-adrenoceptor antagonist with a sulphonylurea drug could provide a novel option for the treatment of type 2 diabetes, especially in patients with increased tonic α2-adrenoceptor-mediated inhibition of insulin secretion.

Comparison of simultaneous measurement of mouse locomotor activity by radiotelemetry and photobeam methods.

INTRODUCTION: Locomotor activity recordings are widely used in different physiological and pharmacological studies. There are two mainly used methods - radiotelemetry and photobeam recording systems. To our knowledge, these methods have not previously been directly and simultaneously compared. METHODS: The current study consisted of a comparison of locomotor activity data gained simultaneously from radiotelemetry and photobeam recordings, firstly from a robotic device and secondly from an animal experiment performed with mice. RESULTS: Data gained from the animal study showed relatively high variation, but overall agreement between the methods was good. DISCUSSION: The two methods were cross-validated in the current study. The data gained from both methods were in good general agreement. However, in an animal experiment, e.g. when sedative drugs or other behavior-modifying interventions are used, one should interpret the results with caution as alterations in animal behavior (e.g. in grooming) may possibly not be picked up similarly by the two methods.

Is there a peripheral site of action contributing to the voiding effects of α2-adrenoceptor agonists and antagonists?

PURPOSE: Since it has not been established whether there is an effect on voiding exerted by direct stimulation or blockade of α2-adrenoceptors in the bladder and urethra, MK-467, a peripherally acting α2-adrenoceptor antagonist not penetrating into the CNS, was used to test whether part of the voiding effects of systemically given α2-adrenoceptor agonists is peripheral. METHODS: Urodynamic recordings from 27 conscious male adult C57/Bl J-strain mice were performed. After vehicle (saline) administration, two groups of animals were treated first with the selective α2-adrenoceptor agonist dexmedetomidine (Dex) and then with the selective α2-adrenoceptor antagonists atipamezole (Ati) or MK-467. Two other groups were first treated with Ati or MK-467 and then with Dex. RESULTS: Treatment with vehicle or α2-adrenoceptor antagonists alone did not affect micturition parameters. All animals treated first with Dex-developed overflow incontinence. Treatment with Ati after Dex reversed almost totally the effects of Dex on all voiding parameters, but treatment with MK-467 after Dex showed no detectable improvement. Treatment with Dex after Ati had no effect on any voiding parameter except maximal pressure. When mice were treated with Dex after MK-467, overflow incontinence was produced in seven of eight animals studied. CONCLUSIONS: The absence of functionally relevant peripheral effects on voiding mediated via α2-adrenoceptors is supported by the finding that neither Ati nor MK-467 alone had any effect on micturition parameters and by the inability of MK-467 to inhibit the effects of Dex, suggesting that the relevant Dex effects were exerted within the CNS.

Hemodynamic actions and mechanisms of systemically administered α-MSH analogs in mice.

α-Melanocyte-stimulating hormone (α-MSH) regulates important physiological functions including energy homeostasis and inflammation. Potent analogs of α-MSH, [Nle(4), D-Phe(7)]-α-MSH (NDP-α-MSH) and melanotan-II (MT-II), are widely used in pharmacological studies, but the hemodynamic effects associated with their systemic administration have not been thoroughly examined. Therefore, we investigated the hemodynamic actions of these compounds in anesthetized and conscious C57Bl/6N mice using peripheral routes of administration. NDP-α-MSH and MT-II induced mild changes in blood pressure and heart rate in anesthetized mice compared to the effects observed in conscious mice, suggesting that anesthesia distorts the hemodynamic actions of α-MSH analogs. In conscious mice, NDP-α-MSH and MT-II increased blood pressure and heart rate in a dose-dependent manner, but the tachycardic effect was more prominent than the pressor effect. Pretreatment with the melanocortin (MC) 3/4 receptor antagonist SHU9119 abolished these hemodynamic effects. Furthermore, the blockade of ß(1)-adrenoceptors with metoprolol prevented the pressor effect and partly the tachycardic action of α-MSH analogs, while the ganglionic blocker hexamethonium abrogated completely the difference in heart rate between vehicle and α-MSH treatments. These findings suggest that the pressor effect is primarily caused by augmentation of cardiac sympathetic activity, but the tachycardic effect seems to involve withdrawal of vagal tone in addition to sympathetic activation. In conclusion, the present results indicate that systemic administration of α-MSH analogs elevates blood pressure and heart rate via activation of MC(3/4) receptor pathways. These effects and the consequent increase in cardiac workload should be taken into account when using α-MSH analogs via peripheral routes of administration.

Effects on bladder function of combining elocalcitol and tolterodine in rats with outflow obstruction.

UNLABELLED: It has previously been shown that elocalcitol might protect bladder contractile function in experimental models and that elocalcitol has beneficial effects in patients with LUTS. In humans, elocalcitol was demonstrated with a very good safety profile but only exhibited limited efficacy on LUTS in patients with BPH and overactive bladder (OAB). Recent reports show that therapies with antimuscarinics, when combined with other drugs in clinical use, might perform better than a monotherapy in managing LUTS. It is not known how a combination of elocalcitol and an antimuscarinic performs on bladder dysfunction. The present study suggests that concomitant use of secosteroids and antimuscarinics has additive beneficial effects on obstruction-related functional changes in an experimental model. If confirmed also in a clinical setting, this could allow for individual dose adjustments to improve efficacy in obstruction-related LUTS, and possibly reduce unwanted adverse activities by antimuscarinic therapy. OBJECTIVE: To evaluate the effects of tolterodine on urodynamics in elocalcitol- or vehicle-treated rats with partial urethral obstruction (PUO). MATERIALS AND METHODS: After ethical approval, 20 female Sprague-Dawley rats were subjected to PUO and treated (gavage) for 14 days (once daily) with elocalcitol (75 µg/kg) or vehicle. Cystometries were performed on day 15 in awake rats before and after i.v. administration of tolterodine (1, 10 and 100 µg/kg). RESULTS: No differences in bladder weights or body/bladder weight ratios were noted between groups. Tolterodine dose-dependently increased micturition intervals and volumes and bladder capacity in both elocalcitol- (n = 11) and vehicle-treated rats (n = 9). In elocalcitol-treated rats, flow pressure (FP) was dose-dependently reduced (12-20%) by tolterodine, whereas no effect on FP was noted in vehicle-treated animals (P < 0.05). Flow compliance (FC) was increased by tolterodine by 21-54% in vehicle-treated rats, and by 47-131% (P < 0.05 vs vehicle) in elocalcitol-treated animals. Maximal tension vs bladder weight was improved in elocalcitol-treated rats in comparison to vehicle (P < 0.05). The area under the curve (AUC) was reduced by tolterodine with 11-16% in vehicle-treated rats and 26-30% in elocalcitol -treated rats (P < 0.05). CONCLUSIONS: Elocalcitol-treatment improved the effects of tolterodine on bladder compliance at the start of flow. The effects of tolterodine on AUC suggest that elocalcitol exerts additional beneficial actions on PUO-induced functional changes during the filling phase of micturition. The reduction of FP and increase in FC by elocalcitol and tolterodine could have translational value and, if valid in humans, support combined therapy in benign prostatic obstruction (BPO)-related lower urinary tract symptoms (LUTS).

Cannabinor, a selective cannabinoid-2 receptor agonist, improves bladder emptying in rats with partial urethral obstruction.

PURPOSE: We studied the effects of chronic treatment with the novel selective cannabinoid 2 receptor agonist cannabinor (Procter & Gamble Pharmaceuticals, Cincinnati, Ohio) on bladder function in conscious rats with partial urethral obstruction and on the functional properties of isolated detrusor muscle. MATERIALS AND METHODS: A total of 24 female Sprague-Dawley® rats with surgically created partial urethral obstruction received daily intraperitoneal injections of 3 mg/kg cannabinor (12) or saline as controls (12) for 2 weeks. Cystometry was done, the rats were sacrificed and the bladders were prepared for in vitro studies. RESULTS: Mean ± SEM bladder weight was 0.97 ± 0.15 gm in controls and 0.53 ± 0.08 gm in cannabinor treated rats (p <0.05). There was no difference between the groups in the mean micturition interval, or mean baseline, threshold, flow or maximum pressure. In controls and cannabinor treated rats mean post-void residual volume was 0.28 ± 0.07 and 0.06 ± 0.02 ml, mean micturition compliance was 0.032 ± 0.006 and 0.069 ± 0.016 ml/cm H(2)O, and mean bladder wall force at the start of flow was 950 ± 280 and 1,647 ± 325 mN/gm, respectively (each p <0.05). Nonvoiding contractions were significantly less frequent in cannabinor treated rats than in controls. We noted no difference in carbachol (Sigma®) half maximum concentration between the groups but the carbachol maximum response in detrusor strips from cannabinor treated rats was significantly higher than that in control strips. CONCLUSIONS: In rats with partial urethral obstruction treated daily for 14 days with cannabinor bladder weight was lower, the ability to empty the bladder was preserved and nonvoiding contraction frequency was low compared to those in controls. Detrusor preparations from cannabinor treated rats showed a higher response to nerve stimulation than those from controls. Selective cannabinoid 2 receptor activation may be a novel principle to enable improved bladder function after partial urethral obstruction.

TRPA1 receptor induced relaxation of the human urethra involves TRPV1 and cannabinoid receptor mediated signals, and cyclooxygenase activation.

PURPOSE: We studied whether TRPA1 agonists interact with sensory and inflammatory signals to relax human urethral smooth muscle. MATERIALS AND METHODS: Urethral specimens were obtained perioperatively from 19 patients, and prepared for immunohistochemistry and functional experiments. The effects of allyl isothiocyanate, cinnamaldehyde and NaHS were studied in phenylephrine activated preparations combined with capsaicin, capsazepine, N omega-nitro-L-arginine, indomethacin or CP55940. RESULTS: TRPA1, cannabinoid 1 and cannabinoid 2 immunoreactivity was colocalized in nerve fibers of the human urethra. All TRPA1 agonists produced relaxation of phenylephrine contracted urethral preparations. Capsaicin increased relaxant responses to all TRPA1 agonists. It increased the mean +/- SEM -logIC50 of cinnamaldehyde and NaHS from 4.91 +/- 0.26 to 5.15 +/- 0.22 and 3.27 +/- 0.14 to 3.79 +/- 0.35, and the -logIC30 of allyl isothiocyanate from 3.11 +/- 0.24 to 3.41 +/- 0.26 (each p <0.05). Capsazepine in 5 preparations, indomethacin in 6 and CP55940 in 5 decreased cinnamaldehyde mediated relaxation by up to 39%, 88% and 89%, respectively. Nomega-nitro-L-arginine and urothelial removal had no effect on relaxation by cinnamaldehyde in 5 preparations. CONCLUSIONS: Relaxation to TRPA1 agonists in human urethral preparations seem to work in cooperation with TRPV1 mediated signals, are negatively coupled via cannabinoid receptor activation and involve cyclooxygenase products. Urothelial TRPA1 signals may not be important to regulate normal human urethral smooth muscle tone. This does not exclude a role in the initiation of afferent activity normally and in disease states.

Effects of cannabinor, a novel selective cannabinoid 2 receptor agonist, on bladder function in normal rats.

BACKGROUND: Cannabinoid (CB) receptors may be involved in the control of bladder function; the role of CB receptor subtypes in micturition has not been established. OBJECTIVES: Our aim was to evaluate the effects of cannabinor, a novel CB2 receptor agonist, on rat bladder function. DESIGN, SETTING, AND PARTICIPANTS: Sprague Dawley rats were used. Distribution of CB2 receptors in sensory and cholinergic nerves of the detrusor was studied. Selectivity of cannabinor for human and rat CB receptors was evaluated. Effects of cannabinor on rat detrusor and micturition were investigated. MEASUREMENTS: Immunohistochemistry, radioligand binding, tritium outflow assays, organ bath studies of isolated bladder tissue, and cystometry in awake rats were used. RESULTS AND LIMITATIONS: CB2 receptor immunoreactivity was expressed in the urothelium and in sensory and cholinergic bladder nerves. Cannabinor exhibited similar binding at human and rat CB2 receptors and a 321-fold functional selectivity for the CB2 receptor versus the CB1 receptor. Cannabinor had no effect on isolated detrusor muscle function. In vivo, cannabinor 3.0mg/kg increased micturition intervals and volumes by 52% (p<0.05) and 96% (p<0.01), respectively, and increased threshold and flow pressures by 73% (p<0.01) and 49% (p<0.001), respectively. Cannabinor 0.3 or 1.0mg/kg or vehicle did not affect urodynamic parameters. CONCLUSIONS: Considering that CB2 receptors are localized on sensory nerves and on the urothelium and that cannabinor had effects on "afferent" urodynamic parameters, peripheral CB2 receptors may be involved in sensory functions of rat micturition. Effects of cannabinor on cholinergic nerve activity in normal bladder tissue appear to be limited.

Voiding effects mediated by α2-adrenoceptors in the anaesthetized male rat.

OBJECTIVE: To study the role of α(2)-adrenoceptors (α(2)-AR) in micturition of anaesthetized male rats, with specific focus on the effects on the electrical activity (by electromyography, EMG) of the rhabdosphincter, and actual urinary flow rate, as the effects mediated by α(2)-ARs on sphincter activity and urethral pressures have not been established. MATERIALS AND METHODS: Adult anaesthetized male Noble rats were used; intravesical pressure, rhabdosphincter EMG and urinary flow rate from the distal urethra were recorded. After baseline recordings, an α(2)-AR agonist (dexmedetomidine, DEX) or α(2)-AR antagonist (atipamezole), were injected intravenously. RESULTS: DEX treatment significantly decreased the maximum bladder pressure and urinary flow rate, and the amplitude of rhabdosphincter EMG was significantly reduced. Intraluminal pressure high-frequency oscillations, usually observed during rat voiding were abolished. The effects of DEX were fully reversed within 31 min. Atipamezole treatment significantly increased actual urinary flow rates and rhabdosphincter EMG amplitude, but the number of times flow was interrupted was increased during the voiding cycle, leading to increased overall micturition time. CONCLUSION: Stimulation and blockade of α(2)-ARs have a significant effect on lower urinary tract function. If the data from this rat model are also valid in humans, a study of the effects of atipamezole on urethral sphincter activity and urethral pressures in humans would be of interest, and might show therapeutic potential of the drug.

Distribution and function of cannabinoid receptors 1 and 2 in the rat, monkey and human bladder.

PURPOSE: We investigated the distribution of cannabinoid receptor subtypes 1 and 2 in the detrusor of different species and studied the effects of cannabinoid receptor 1 and 2 agonists on bladder function. MATERIALS AND METHODS: Cannabinoid receptor 1 and 2 expression was studied with Western blot and immunohistochemistry in rat, monkey and human detrusors. Co-staining was done for markers of sensory nerves using calcitonin gene-related peptide (Euro-Diagnostica, Malmö, Sweden) and transient receptor potential vanilloid 1, and for cholinergic nerves using VAChT (Santa Cruz Biotechnology, Santa Cruz, California). Actions of the endogenous cannabinoid receptor-1 and 2 agonist anandamide (Sigma(R)), and the cannabinoid receptor 1 and 2 agonist CP55,940 (Sigma) on isolated detrusor and during cystometry in conscious rats were recorded. RESULTS: Higher expression of cannabinoid receptor 2 but not cannabinoid receptor 1 was noted in the mucosa than in the detrusor. Compared to the detrusor larger amounts of cannabinoid receptor 2 containing nerves that also expressed transient receptor potential vanilloid 1 or calcitonin gene-related peptide were observed in the suburothelium. Nerve fibers containing cannabinoid receptor 2 and VAChT were located in the detrusor. Neither anandamide nor CP55,940 affected isolated detrusor carbachol (Sigma) contractions. Nerve contractions were enhanced by 10 muM anandamide and decreased by 10 muM CP55,940 (p<0.05). In vivo CP55,940 increased the micturition interval by 46% and threshold pressure by 124% (p <0.05). Anandamide increased threshold pressure by 26% and decreased the micturition interval by 19% (p <0.05 and <0.01, respectively). CONCLUSIONS: The distribution of cannabinoid receptor 2 on sensory nerves and in the urothelium, and effects by CP55940 on the micturition interval and threshold pressure suggest a role for cannabinoid receptor 2 in bladder afferent signals. Co-expression of VAChT and cannabinoid receptor 2, and effects by CP55940 on nerve contractions suggest a cannabinoid receptor 2 mediated modulatory effect on cholinergic nerve activity. Anandamide may not be a good tool for cannabinoid receptor studies due to its activity at other receptors.

Transient receptor potential A1 (TRPA1) activity in the human urethra--evidence for a functional role for TRPA1 in the outflow region.

BACKGROUND: A role for the transient receptor potential (TRP) A1 ion channel in rat lower urinary tract (LUT) sensation and disease has been proposed, but in the human LUT no information on TRPA1 activity is available. OBJECTIVES: To investigate the distribution of TRPA1 in the human urethra and to study the effect of TRPA1 agonists on isolated urethral strip preparations. DESIGN, SETTINGS, AND PARTICIPANTS: Urethral specimens were obtained preoperatively from 10 patients and were freshly prepared for Western blot, immunohistochemistry, and functional in vitro investigations. MEASUREMENTS: The expression patterns of TRPA1 were studied with Western blot and immunohistochemistry. The effects of allyl isothiocyanate (AI), cinnamaldehyde (CA), and NaHS (donor of H(2)S) on tension of urethral strips were investigated in tissue baths. RESULTS AND LIMITATIONS: TRPA1 immunoreactivity (-IR) was found in nerve fibres in the suburothelial space and was also located to nerve fibres of the muscle layer. Single TRPA1-IR nerves extended into the urothelium. A majority, but not all TRPA1-IR nerves also expressed immunoreactivity for CGRP or TRPV1. In the urothelium, TRPV1 was located to the outer layers whereas TRPA1 was observed in basal urothelial cells. Interspersed between strands of smooth muscle cells of the urethral wall, TRPA1- and vimentin-IR cells containing central nuclei and slender cytoplasmatic extensions were observed. In functional experiments, TRPA1-agonists had no contractile effect in urethral preparations. After precontraction with phenylephrine, AI, CA, and NaHS caused concentration-dependent relaxations of urethral strip preparations. CONCLUSIONS: The localization of TRPA1 to nerves that also express TRPV1 and CGRP, and in urothelial cells and interstitial cells, as well as the findings that TRPA1 agonists can modify tone of urethral preparations, propose a role for TRPA1 in afferent and efferent sensory signaling of the human outflow region.

Human HSD17B1 expression masculinizes transgenic female mice.

When present in excess amounts during fetal life, androgens can impair female development by inducing masculinization. On way to modify fetal steroid concentration is by altering the expression of hydroxysteroid (17beta) dehydrogenases (HSD17Bs). Human HSD17B1 converts weak estrogen estrone to estradiol, and with lower catalytic efficiency, weak androgen androstenedione to testosterone. We have recently shown that over-expression of human HSD17B1 in transgenic mice results in masculinized phenotype in female mice. In the present study, we further show that in addition to the Müllerian ducts, HSD17B1TG females have internal structures resembling Wolffian ducts, and enlarged Skene paraurethral gland, also called the female prostate. HSD17B1 expression has been found in fetal human ovary, thus, it is possible that HSD17B1 contributes to maintain the normal steroid hormone concentration during development. Thereby, abnormal increase in the fetal expression of HSD17B1 could contribute to the development of hormonal imbalances, and so result in female masculinization.

Prostatic inflammation and obstructive voiding in the adult Noble rat: impact of the testosterone to estradiol ratio in serum.

BACKGROUND: The age-related decline of the testosterone to estradiol (T-to-E(2)) ratio in serum is associated with the increased prevalence of prostatic inflammation and lower urinary tract symptoms suggesting obstructive voiding. The impact of the T-to-E(2) ratio on the development and reversal of non-bacterial prostatic inflammation and obstructive voiding was tested in adult Noble rats. METHODS: Adult male Noble rats (n = 16) were treated with estradiol (83 microg/day) and two different doses (280 and 830 microg/day) of testosterone to cause hypoandrogenic and hyperandrogenic states with elevated estrogen. After the 13-week hormonal treatment, urodynamical measurements and electrical activity recording of the rhabdosphincter muscle were performed under anesthesia. Testosterone, estradiol, and prolactin concentrations in serum were measured and inflammatory changes in the dorsolateral prostate were classified and counted. RESULTS: Histopathological and urodynamical analyses indicated that the hypoandrogenic animals with a decreased T-to-E(2) ratio (10 versus > 300 in control) developed prostatic inflammation and non-obstructive voiding. The hyperandrogenic state with decreased T-to-E(2) ratio of 50 decreased the aggressiveness of the inflammation and the number of inflamed acini in the prostate and caused urethral obstruction associated with rhabdosphincter dysfunction. CONCLUSIONS: Different responses of the prostatic inflammation and voiding function to the change in T-to-E(2) ratio imply that non-bacterial prostatic inflammation is not a sufficient condition for the development of obstructive voiding. The present study finds no support for the idea that age- and/or obesity-related hypoandrogenic state with a decreased ratio of T-to-E(2) would cause urethral obstruction.

Distribution and function of the hydrogen sulfide-sensitive TRPA1 ion channel in rat urinary bladder.

OBJECTIVES: To investigate the distribution of the transient receptor potential (TRP) A1 ion channel in the rat urinary bladder, and to study the effects of hydrogen sulfide (H(2)S) and known TRPA1 activators on micturition in conscious rats and on heterologously expressed ion channels. METHODS: The expression of TRPA1 in urinary bladder was studied with fluorescence immunohistochemistry and real-time PCR in female Sprague-Dawley rats. Cystometric investigations were performed in conscious animals subjected to intravesical administration of sodium hydrogen sulfide (NaHS, donor of H(2)S), allyl isothiocyanate (AI), and cinnamaldehyde (CA). Fluorometric calcium imaging was used to study the effect of NaHS on human and mouse TRPA1 expressed in CHO cells. RESULTS: TRPA1 immunoreactivity was found on unmyelinated nerve fibres within the urothelium, suburothelial space, and muscle layer as well as around blood vessels throughout the bladder. All TRPA1 immunoreactive nerves fibres also expressed TRPV1 immunoreactivity and vice versa. TRPA1 was also detected in urothelial cells at both transcriptional and protein levels. AI increased micturition frequency and reduced voiding volume. CA and NaHS produced similar changes in urodynamic parameters after disruption of the urothelial barrier with protamine sulfate. NaHS also induced calcium responses in TRPA1-expressing CHO cells, but not in untransfected cells. CONCLUSIONS: The expression of TRPA1 on C-fibre bladder afferents and urothelial cells together with the finding that intravesical TRPA1 activators initiate detrusor overactivity indicate that TRPA1 may have a role in sensory transduction in this organ. The study also highlights H(2)S as a TRPA1 activator potentially involved in inflammatory bladder disease.

Simultaneous registration of intraabdominal and intravesical pressures during cystometry in conscious rats--effects of bladder outlet obstruction and intravesical PGE2.

AIMS: A method was developed and evaluated to simultaneously register intraabdominal pressure (IAP) and intravesical pressure (IVP) during cystometry in conscious rats. In addition, IAP and IVP were recorded in rats with experimental detrusor overactivity (DO). METHODS: Sprague Dawley rats (n = 24) were used. Six female rats were subjected to partial bladder outlet obstruction for 2 weeks. A catheter was implanted into the bladder to record the IVP, and a balloon-fitted catheter was positioned in the abdominal cavity to record the IAP. PGE(2) was given intravesically to induce DO. Detrusor pressure (DP) was defined as the IVP corrected for IAP. RESULTS: Recorded as increases in IAP, all rats of both sexes exhibited abdominal straining during every void. In controls, a maximal IAP of 6.0 +/- 1.4 cmH(2)O (range 3-15 cmH(2)O) was registered (n = 12) at the time of the flow pressure (FP). Intravesical administration of PGE(2) or BOO did not affect the IAP at basal pressure, FP or micturition pressure. Changes in IAP due to movement or non-voiding-related straining were subtracted from IVP to generate DP and to visualize DO after BOO or intravesical PGE(2). CONCLUSIONS: The conscious rat uses abdominal straining during voiding, and maximal IAP is recorded at the onset of urinary flow. Simultaneous registration of IAP and IVP during the micturition cycle in conscious rats is a convenient method for accurate quantification of pressures inside the bladder and for studying "true" DO without interference from movement artifacts.