Dog and human bladders have different neurogenic and nicotinic responses in inner versus outer detrusor muscle layers.

The aim of this study was to investigate layer and species variations in detrusor muscle strip responses to myogenic, neurogenic, and nicotinic, and muscarinic receptor stimulations. Strips from bladders of 9 dogs and 6 human organ transplant donors were dissected from inner and outer longitudinal muscle layers, at least 1 cm above urethral orifices. Strips were mounted in muscle baths and maximal responses to neurogenic stimulation using electrical field stimulation (EFS) and myogenic stimulation using potassium chloride (KCl, 120 mM) determined. After washing and re-equilibration was completed, responses to nicotinic receptor agonist epibatidine (10 µM) were determined followed by responses to EFS and muscarinic receptor agonist bethanechol (30 µM) in continued presence of epibatidine. Thereafter, strips and full-thickness bladder sections from four additional dogs and three human donors were examined for axonal density and intramural ganglia. In dog bladders, contractions to KCl, epibatidine, and bethanechol were 1.5- to 2-fold higher in the inner longitudinal muscle layer, whereas contractions to EFS were 1.5-fold higher in the outer (both pre- and post-epibatidine). Human bladders showed 1.2-fold greater contractions to epibatidine in the inner layer and to EFS in the outer, yet no layer differences to KCl or bethanechol were noted. In both species, axonal density was 2- to 2.5-fold greater in the outer layer. Dogs had more intramural ganglia in the adventitia/serosa layer, compared with more internal layers and to humans. These findings indicate several layer-dependent differences in receptor expression or distribution, and neurogenic responses in dog and human detrusor muscles, and myogenic/muscarinic differences between dog versus humans.

Anti-ulcerogenic mechanisms of the sesquiterpene lactone onopordopicrin-enriched fraction from Arctium lappa L. (Asteraceae): role of somatostatin, gastrin, and endogenous sulfhydryls and nitric oxide.

Arctium lappa L. has been used in folk medicine as a diuretic, depurative, and digestive stimulant and in dermatological conditions. The mechanisms involved in the anti-ulcerogenic activity of the sesquiterpene onopordopicrin (ONP)-enriched fraction (termed the ONP fraction), obtained from A. lappa leaves, were studied. The gastroprotective mechanism of the ONP fraction was evaluated in experimental in vivo models in rodents, mimicking this disease in humans. ONP fraction (50 mg/kg, p.o.) significantly inhibited the mucosal injury induced by ethanol/HCl solution (75%), indomethacin/bethanecol (68.9%), and stress (58.3%). When the ONP fraction was investigated in pylorus ligature, it did not induce alteration in the gastric volume but did modify the pH and total acid concentration of gastric juice. ONP fraction significantly increased serum somatostatin levels (82.1±4.1 vs. control group 12.7±4 pmol/L) and decreased serum gastrin levels (62.6±6.04 vs. control group 361.5±8.2 µU/mL). Mucus production was not significantly altered by the ONP fraction. Gastroprotection by the ONP fraction was completely inhibited by N-ethylmaleimide treatment and did not modify the effect in the animals pretreated with l-N(G)-nitroarginine methyl ester. These results suggest an antisecretory mechanism involved with the antiulcerogenic effect of the ONP fraction. However, only endogenous sulfhydryls play an important role in gastroprotection of the ONP fraction.

Beneficial effects of estrogen in a mouse model of cerebrovascular insufficiency.

BACKGROUND: The M(5) muscarinic acetylcholine receptor is known to play a crucial role in mediating acetylcholine dependent dilation of cerebral blood vessels. Previously, we reported that male M(5) muscarinic acetylcholine knockout mice (M5R(-/-) mice) suffer from a constitutive constriction of cerebral arteries, reduced cerebral blood flow, dendritic atrophy, and short-term memory loss, without necrosis and/or inflammation in the brain. METHODOLOGY/PRINCIPAL FINDINGS: We employed the Magnetic Resonance Angiography to study the area of the basilar artery in male and female M5R(-/-) mice. Here we show that female M5R(-/-) mice did not show the reduction in vascular area observed in male M5R(-/-) mice. However, ovariectomized female M5R(-/-) mice displayed phenotypic changes similar to male M5R(-/-) mice, strongly suggesting that estrogen plays a key role in the observed gender differences. We found that 17beta-estradiol (E2) induced nitric oxide release and ERK activation in a conditional immortalized mouse brain cerebrovascular endothelial cell line. Agonists of ERalpha, ERbeta, and GPR30 promoted ERK activation in this cell line. Moreover, in vivo magnetic resonance imaging studies showed that the cross section of the basilar artery was restored to normal in male M5R(-/-) mice treated with E2. Treatment with E2 also improved the performance of male M5R(-/-) mice in a cognitive test and reduced the atrophy of neural dendrites in the cerebral cortex and hippocampus. M5R(-/-) mice also showed astrocyte swelling in cortex and hippocampus using the three-dimensional reconstruction of electron microscope images. This phenotype was reversed by E2 treatment, similar to the observed deficits in dendrite morphology and the number of synapses. CONCLUSIONS/SIGNIFICANCE: Our findings indicate that M5R(-/-) mice represent an excellent novel model system to study the beneficial effects of estrogen on cerebrovascular function and cognition. E2 may offer new therapeutic perspectives for the treatment of cerebrovascular insufficiency related memory dysfunction.

Necessity of intracellular cyclic AMP in inducing gastric acid secretion via muscarinic M3 and cholecystokinin2 receptors on parietal cells in isolated mouse stomach.

The existence of a direct action of acetylcholine and gastrin on muscarinic M3 and cholecystokinin2 (CCK2) receptors on gastric parietal cells has not yet been convincingly established because these stimulated acid secretions are remarkably inhibited by histamine H2 receptor antagonists. In the present study, we investigated the necessity of intracellular cyclic AMP in inducing gastric acid secretion via muscarinic M3 and CCK2 receptors on parietal cells using an isolated mouse stomach preparation. Bethanechol (10-300 microM) produced a marked increase in acid output and this increase was completely blocked by famotidine (10 microM). In the presence of famotidine, bethanechol (1-30 microM) augmented the acid secretory response to dibutyryl AMP (200 microM) in a concentration-dependent manner. The augmentation was blocked by atropine (1 microM), 4-DAMP (0.1 microM), a muscarinic M3-selective antagonist, and by Ca2+ exclusion from the serosal nutrient solution. Pentagastrin (0.3-3 microM) also concentration-dependently stimulated gastric acid secretion, but the effect was completely inhibited by famotidine. In the presence of famotidine, pentagastrin (0.1-0.3 microM) elicited a definite potentiation of the acid secretory response to dibutyryl cyclic AMP (200 microM). This potentiation was inhibited by YM022 (1 microM), a CCK2 receptor antagonist, and by exclusion of Ca2+ from the serosal nutrient solution. The present results suggest that gastric acid secretion via the activation of muscarinic M3 and CCK2 receptors on the parietal cells is induced by activation of the cyclic AMP-dependent secretory pathway.