Fernando R. Báu

Long-term nitric oxide deficiency causes muscarinic supersensitivity and reduces β3-adrenoceptor-mediated relaxation, causing rat detrusor overactivity

Overactive bladder is a complex and widely prevalent condition, but little is known about its physiopathology. We have carried out morphological, biochemical and functional assays to investigate the effects of long‐term nitric oxide (NO) deficiency on muscarinic receptor and β‐adrenoceptor modulation leading to overactivity of rat detrusor muscle.

Evaluation of the relaxant effect of the nitric oxide-independent soluble guanylyl cyclase stimulator BAY 41-2272 in isolated detrusor smooth muscle

The nitric oxide (NO)-independent soluble guanylyl cyclase stimulator stimulator BAY 41-2272 was reported to produce relaxant response in different types of smooth muscle. However no study was carried out to investigate the effects of BAY 412282 in detrusor smooth muscle. Thus, this study aimed to evaluate the relaxant effects of BAY 41-2272, in isolated mouse, rat and rabbit detrusor smooth muscle. Mouse, rat and rabbit were anesthetized, and urinary bladder removed. Detrusor smooth muscle was transferred to 10-mL organ baths containing oxygenated and warmed Krebs-Henseleit solution. Tissues were connected to force-displacement transducers and changes in isometric force were recorded. BAY 41-2272 (0.001-100 microM) produced concentration-dependent detrusor smooth muscle relaxations in mouse, rat and rabbit with maximal responses of 61.3+/-6.6%, 95.1+/-9.9% and 91.7+/-5.9%, respectively. Sodium nitroprusside and glyceryl trinitrate, as well as 8-bromo-cGMP also produced detrusor relaxations, but to a much lesser extent than BAY 41-2272. The NO synthesis inhibitor L-NAME and the phosphodiesterase-5 inhibitor sildenafil had no effect in BAY 41-2272-induced responses. However, the soluble guanylyl cyclase inhibitor ODQ significantly reduced BAY 41-2272-induced relaxations. BAY 41-2272 increased the bladder cGMP levels by about of 14- and 20-fold for 10 and 100 microM, respectively, which were markedly reduced by ODQ. The cAMP levels were unaffected by BAY 41-2272. Moreover, BAY 41-2272 significantly reduced the contractile responses to extracellular Ca(2+) in an ODQ-insensitive manner. In conclusion, rabbit detrusor smooth muscle relaxations by BAY 41-2272 involve mainly cGMP production, but an additional mechanism involving Ca(2+) influx blockade independently of cGMP production appears to be involved.

Insulin relaxes bladder via PI3K/AKT/eNOS pathway activation in mucosa: unfolded protein response-dependent insulin resistance as a cause of obesity-associated overactive bladder

•  Metabolic syndrome, obesity and insulin resistance are risk factors for overactive bladder, but little is known about the impact of insulin resistance on bladder functioning. •  We aimed to investigate the insulin effects in murine and human bladder, and whether its defective action contributes to the bladder dysfunction associated with obesity. •  Under physiological conditions, insulin relaxes urinary bladder through the release of nitric oxide from the mucosal layer, and this mechanism is impaired in bladders from insulin‐resistant obese mice. •  The defective insulin action in bladder mucosa from obese mice is due to endoplasmic reticulum stress, which remarkably contributes to the bladder overactivity in obesity conditions. •  Our results enable a better understanding of the mechanism of action of insulin in the urinary bladder and how its defective action in mucosa contributes to bladder dysfunction in conditions of obesity‐associated insulin resistance.

Characterization of the urinary bladder dysfunction in renovascular hypertensive rats

Association between arterial hypertension and urinary bladder dysfunction has been reported in humans and spontaneously hypertensive rats. However, no study exists evaluating the bladder dysfunction in conditions of renovascular hypertension. The purpose of this study was to characterize the bladder dysfunction in two kidney‐one clip (2K‐1C) hypertensive rats.

Vas deferens smooth muscle responses to the nitric oxide-independent soluble guanylate cyclase stimulator BAY 41‐2272

The nitric oxide-cGMP signaling pathway modulates the ejaculatory functions. The nitric oxide (NO)-independent soluble guanylate cyclase haem-dependent stimulator BAY 41-2272 potently relaxes different types of smooth muscles. However, no study investigated its effects in vas deferens smooth muscle. Therefore, we designed experiments to evaluate the in vitro relaxing responses of vas deferens to BAY 41-2272. The effects of prolonged oral intake with BAY 41-2272 in vas deferens contractions of rats treated chronically with the NO synthase inhibitor N(ω)-nitro-L-arginine methyl ester (L-NAME) were also investigated. BAY 41-2272 (0.001-100 μM) produced concentration-dependent relaxations in the prostatic and epididymal portions of vas deferens, an effect markedly reduced by the soluble guanylate cyclase inhibitor ODQ (100 μM). BAY 41-2272 significantly increased cGMP levels that were fully prevented by ODQ. In separate protocols, rats received L-NAME (20mg/rat/day) concomitantly with BAY 41-2272 (10mg/kg/day, 4 weeks), after which vas deferens contractions to electrical-field stimulation and noradrenaline were achieved. Electrical-field stimulation (1-32 Hz) evoked frequency-dependent contractions that were significantly enhanced in L-NAME-treated rats. Co-treatment with BAY 41-2272 fully reversed the increased contractile responses in L-NAME group. Noradrenaline (0.01-100 μM)-induced contractions were also greater in L-NAME-treated rats, and that was normalized by BAY 41-2272. In conclusion, BAY 41-2272 potently relaxes in vitro rat vas deferens smooth muscle and elevates the cGMP levels in an ODQ-sensitive manner. Moreover, prolonged oral intake with BAY 41-2272 restores the enhanced contractile vas deferens activity in rats treated with L-NAME. NO-independent soluble guanylate cyclase stimulators may be an alternative treatment for premature ejaculation.

Increased Rho-kinase-mediated prostate contractions associated with impairment of β-adrenergic-cAMP-signaling pathway by chronic nitric oxide deficiency.

Impairment of nitric oxide (NO) - cyclic GMP signaling pathway is likely to contribute to human begnin prostate hyperplasia (BPH). In the present study we have used a model of chronic NO synthesis inhibition to evaluate the functional alterations of prostate smooth muscle (PSM) machinery, and involvement of Rho-kinase pathway. Wistar rats were treated with the NO inhibitor N(ω)-nitro-l-arginine methyl ester (L-NAME, 20mg/kg/day; 4 weeks), after which contractile responses to phenylephrine (α1-adrenoceptor agonist; 1nM to 100µM), carbachol (muscarinic agonist; 1nM to 1mM) and α,β-methylene ATP (P2X receptor agonist; 1-10µM), as well as to electrical-field stimulation (EFS; 1-32Hz) were evaluated. PSM relaxations to isoproterenol (non-selective β-adrenoceptor agonist, 0.1nM to 10µM) and sodium nitroprusside (NO donor, 1nM to 10mM) were also evaluated. The ratio prostate weight/body weight was 22% greater (P<0.05) in L-NAME compared with control group. The PSM contractions to phenylephrine, carbachol and α,β-methylene ATP were higher in L-NAME (Emax: 3.85±0.25, 3.52±0.35 and 2.03±0.2mN, respectively) compared with control group (Emax: 3.08±0.17, 2.37±0.18 and 1.57±0.18mN, respectively). The PSM contractions induced by EFS were also significantly greater in L-NAME group. Prior incubation with the Rho-kinase inhibitor Y27632 (1µM) fully reversed the enhanced contractions to phenylephrine and carbachol. Isoproterenol-induced PSM relaxations were 34% lower in L-NAME group, which was associated with reduced levels of cAMP in prostate tissue. The relaxations to sodium nitroprusside remained unaltered in L-NAME group. In summary, chronic NO deficiency leads to increased Rho-kinase-mediated PSM contractile responses accompanied by impairment of β-adrenergic-cAMP-signaling pathway.

Relaxant effect of BAY 41–2272 in the rabbit isolated detrusor smooth muscle (DSM): involvement of cGMP-independent mechanisms

Background Overactive bladder (OAB) is described as urgency with or without incontinence, usually accompanied by frequency and nocturia. Overactive bladder reflects involuntary DSM as a consequence of enhanced cholinergic-mediated contractions and decreased b-adrenoceptor-mediated relaxations. The nitric oxide (NO)-cyclic GMP signaling pathway has been described to modulate the muscular tone, neurotransmission and blood flow in DSM, but the exact role of NO in the bladder is not completely understood. It is accepted that NO deficiency contributes to triggering OAB. The compound BAY 41–2272 has been described as a drug that directly activates the soluble guanylate cyclase leading to NO-independent cGMP accumulation. BAY 41–2272 produces potent relaxations in vascular and non-vascular smooth muscle tissues by cGMP-dependent and independent mechanisms, but no studies have been carried out with DSM.

Comparative bioavailability of two escitalopram formulations in healthy human volunteers

OBJECTIVE To assess the bioequivalence of two escitalopram formulations (Test formulation: escitalopram (10 mg tablet) manufactured by Apsen Farmacêutica S.A.) Reference formulation: escitalopram (Lexapro; 10 mg tablet) from Lundbeck Brasil Ltda) in healthy volunteers of both sexes. METHODS The study was conducted using an open, randomized, two-period crossover design with at least a 21-day washout interval. Plasma samples were obtained over a 168 h period. Plasma escitalopram concentrations were analyzed by liquid chromatography coupled to tandem mass spectrometry (LC-MS-MS) with positive ion electrospray ionization using multiple reaction monitoring (MRM). The following pharmacokinetic parameters were obtained from the escitalopram plasma concentration vs. time curves: AUC(last), AUC(inf) and C(max). RESULTS The limit of quantification for escitalopram was 0.2 ng x ml(-1). The geometric mean with corresponding 90% confidence interval (CI) for Test/Reference percent ratios were 97.35% (90% CI = 90.28-104.96%) for C(max), 99.60% (90% CI = 92.93-106.74%) for AUC(last) and 99.92% (90% CI = 93.34-106.97%) for AUC(inf). CONCLUSION Since the 90% CI for AUClast, AUCinf and Cmax ratios were within the 80-125% interval proposed by the US FDA, it was concluded that escitalopram formulation manufactured by Apsen Farmacêutica S.A. is bioequivalent to the Lexapro formulation in regard to both the rate and the extent of absorption.

Insulin relaxes bladder via PI3K/AKT/eNOS pathway activation in mucosa: unfolded protein response-dependent insulin resistance as a cause of obesity-associated overactive bladder: Insulin relaxes bladder via PI3K/AKT/eNOS pathway

•  Metabolic syndrome, obesity and insulin resistance are risk factors for overactive bladder, but little is known about the impact of insulin resistance on bladder functioning. •  We aimed to investigate the insulin effects in murine and human bladder, and whether its defective action contributes to the bladder dysfunction associated with obesity. •  Under physiological conditions, insulin relaxes urinary bladder through the release of nitric oxide from the mucosal layer, and this mechanism is impaired in bladders from insulin‐resistant obese mice. •  The defective insulin action in bladder mucosa from obese mice is due to endoplasmic reticulum stress, which remarkably contributes to the bladder overactivity in obesity conditions. •  Our results enable a better understanding of the mechanism of action of insulin in the urinary bladder and how its defective action in mucosa contributes to bladder dysfunction in conditions of obesity‐associated insulin resistance.

Insulin relaxes bladder via PI3K/AKT/eNOS pathway activation in mucosa

•  Metabolic syndrome, obesity and insulin resistance are risk factors for overactive bladder, but little is known about the impact of insulin resistance on bladder functioning. •  We aimed to investigate the insulin effects in murine and human bladder, and whether its defective action contributes to the bladder dysfunction associated with obesity. •  Under physiological conditions, insulin relaxes urinary bladder through the release of nitric oxide from the mucosal layer, and this mechanism is impaired in bladders from insulin‐resistant obese mice. •  The defective insulin action in bladder mucosa from obese mice is due to endoplasmic reticulum stress, which remarkably contributes to the bladder overactivity in obesity conditions. •  Our results enable a better understanding of the mechanism of action of insulin in the urinary bladder and how its defective action in mucosa contributes to bladder dysfunction in conditions of obesity‐associated insulin resistance.