The differential contractile responses to capsaicin and anandamide in muscle strips isolated from the rat urinary bladder

The contractile responses to capsaicin and anandamide, exogenous and endogenous agonists for transient receptor potential vanilloid receptor 1 (TRPV1), respectively, were investigated in muscle strips isolated from the rat urinary bladder. Capsaicin and anandamide produced concentration-dependent contractions of the muscle strips. The contractile response induced by capsaicin disappeared within approximately 20 min. In contrast, anandamide produced contractile responses lasting at least for 30 min. Capsaicin produced additive contractile responses in anandamide-treated muscle strips. The contractile response to anandamide was attenuated, but not abolished in strips desensitized by capsaicin. The response to capsaicin was abolished in the presence of a TRPV1 antagonist, N-(4-tertiarybutylphenyl)-4-(3-chlorphyridin-2-yl)tetrahydropyrazine-1(2H)-carbox-amide (BCTC), but not altered in the presence of either tetrodotoxin, atropine or indomethacin. In the presence of SR140333, a tachykinin NK₁ receptor antagonist or SR48968, an NK₂ receptor antagonist, the response to capsaicin was attenuated. The response to anandamide was partially attenuated in the presence of ONO8130, a prostanoid EP₁ receptor antagonist, URB597, a fatty-acid amide hydrolase inhibitor, BCTC, SR140333 or SR48968, and almost completely abolished by indomethacin. Neither tetrodotoxin, atropine, a cannabinoid CB₁ receptor antagonist, AM251, nor a cannabinoid CB₂ receptor antagonist, AM630, had any effect on the response to anandamide. These results indicate that capsaicin produces muscle contractions by stimulating the TRPV1 receptor, followed by release of neuropeptides that can activate tachykinin NK₁ and/or NK₂ receptors in the bladder and that the contractile response to anandamide is mediated at least in part by activation of prostanoid EP₁ receptors due to production of prostaglandins in addition to TRPV1 receptor activation.     

Vesico-inhibitory responses and capsaicin-sensitive afferents in rats

Summary (1) The effect of perineal pinching and distension of a balloon inserted into the colon on motility of the urinary bladder has been investigated in adult urethane-anesthetized rats pretreated with capsaicin (50 mg/kg s.c.) or its vehicle 4 days before the experiments. (2) At bladder volumes which were sufficient to elicit reflex micturition, perineal pinching or colonic distension transiently inhibited the ongoing bladder voiding contraction. The somato-vesical inhibitory response was markedly reduced or even abolished by division of pudendal nerves. Neither the somato-vesical nor the colovesical inhibitory response were modified by desensitization with systemically administered capsaicin. (3) Intraurethral administration of capsaicin produced a transient inhibition of the reflexly-activated bladder contractions. A second administration of the drug was less effective, indicating desensitization. Intravenously administered capsaicin had a similar inhibitory effect on bladder motility. (4) The vesico-inhibitory response produced by intraurethral administration of capsaicin was not affected by phentolamine, propranolol, guanethidine, picrotoxin or naloxone, while it was greatly reduced or even abolished by bilateral section of the pudendal nerves. (5) These findings provide evidence that capsaicin-sensitive chemoreceptors in the rat urethra are involved in generating a vesico-inhibitory response via pudendal nerves. On the other hand, no evidence was found for the participation of capsaicin-sensitive nerves in the generation of the somato- or colo-vesical inhibitory response. Send offprint requests to C. A. Maggi at the above address     

The ‘efferent’ function of capsaicin-sensitive nerves: Ruthenium Red discriminates between different mechanisms of activation

We have investigated the ability of Ruthenium Red, an inorganic dye with calcium entry blocking properties, to interfere with the ‘efferent’ function of capsaicin-sensitive sensory nerves. These nerves were activated in the guinea-pig isolated bronchus (atropine in the bath) or left atria (reserpine-pretreated animals, atropine in the bath) by electrical field stimulation or with capsaicin. Both stimuli produced a contraction of the bronchus and a positive inotropic response in the atria, responses which are mediated by endogenous neuropeptides (tachykinins in the bronchus, calcitonin gene-related peptide in the atria) released from sensory nerves. Ruthenium Red (10 μM for 20 min in the cases) selectively inhibited the responses produced by the administration of capsaicin, while leaving the responses to electrical field stimulation unaffected. Likewise, the bronchonconstrictor response to exogenous neurokinin A and the atrial positive inotropic response to calcitonin gene-related peptide were unaffected by Ruthenium Red. A prejunctional site of action Ruthenium Red was confirmed in release experiments where the dye strongly inhibited the capsaicin-evoked outflow of calcitonin gene-related peptide, which is taken as a marker of activation in sensory nerves. Together with other observations, these findings support the concept that there are two independent mechanisms for activating the ‘efferent’ function of sensory nerves, one of which is activated by capsaicin and is Ruthenium Red-sensitive but ω-conotoxin-resistant, while the other is activated by propagated action potentials (field stimulation) and is ω-conotoxin-sensitive and Ruthenium Red-resistant.     

Cadmium chloride induces contractions of the rat isolated urinary bladder by activation of capsaicin-sensitive sensory nerves

Cadmium chloride (CdCl2)(30 microM-1 mM) produced a concentration-related contraction of the rat isolated urinary bladder which was abolished by tetrodotoxin (1 microM) or extrinsic bladder denervation (72 h before). Complete cross-desensitization was observed between the contractile response to Cd and capsaicin, indicating that, at the peripheral level, this inorganic calcium channel blocker can activate the 'efferent' function of capsaicin-sensitive sensory nerves.     

Capsaicin-sensitive afferents in the rat urinary bladder activate a spinal sympathetic cardiovascular reflex

Summary In urethane-anesthetized rats with an intact spinal cord, application of capsaicin on the outer surface of the urinary bladder produced a transient bradycardia, hypotension and negative cardiac inotropism which were neither prevented by i. v. atropine (0.5 mg/kg) nor by cervical vagotomy. In acute spinal rats (C2-C3) application of capsaicin (0.2 and 2 pg in 25 pl) on the urinary bladder induced a transient hypertension, tachycardia and positive cardiac inotropism. A second application (30 min later) induced minor cardiovascular effects, expecially with the higher dose, indicating desensitization. All cardiovascular responses to topical capsaicin were abolished by systemic capsaicin desensitization (50 mg/kg s. c., 4 days before). The excitatory cardiovascular response to capsaicin in acute spinal rats was markedly reduced by bilateral section of pelvic but not hypogastric nerves. Further, it was abolished by pretreatment with hexamethonium (20 mg/kg i.v.) or reserpine (5 mg/kg i. p., 2 days before) and reduced, at various extent for the different components, by phentolamine (0.5 mg/kg i. v.) or propranolol (1 mg/kg). In rats with pelvic and hypogastric nerves intact, section of the cord at a level (T12-L1), just above the medullary segments which receive primary afferent input from the bladder (L6-S1), abolished the excitatory cardiovascular response to application of capsaicin on the bladder. In spinal rats (C2-C3) rapid distension of the urinary bladder with saline produced transient tachycardia, hypertension and positive cardiac inotropism similar to that evoked by capsaicin. These responses were not observed in rats systemically pretreated with capsaicin. These findings indicate that certain bladder afferents which are susceptible to capsaicin desensitization in adult rats activate a spinal reflex having excitatory influence on cardiovascular function. This response is apparently mediated by spinal centers located above the site of entry of bladder pelvic afferents into the cord and most likely involves excitation of preganglionic sympathetic neurons in the spinal cord.Send offprint requests to S. Giuliani at the above address     

Involvement of capsaicin-sensitive sensory nerves in early and delayed cardioprotection induced by a brief ischaemia of the small intestine

Early cardioprotection can be achieved by a brief ischaemia of noncardiac tissues. Our study examined whether a brief ischaemia of the small intestine induces both early and delayed cardioprotection in the rabbit and assessed the possible mechanism involved in the activation of capsaicin-sensitive sensory nerves. The plasma concentration of creatine kinase (CK) and infarct size (necrotic zone/left ventricular zone) after 30 min coronary artery occlusion and 180 min reperfusion were determined in rabbits. Infarct size was 35.5±6.8% in the control non-preconditioned group. Preconditioning induced by a brief period of 10-min small intestine ischaemia significantly reduced infarct size (6.5±1.9%, P<0.01 vs. the control non-preconditioned group) and decreased CK release (3092±236 and 1094±117 U/l for myocardial ischaemia-reperfusion and preconditioning plus myocardial ischemia-reperfusion, respectively, P<0.01), and the protection was partly abolished by pretreatment with capsaicin (50 mg/kg, s.c.) 4 days before the experiments. A brief period of anterior mesenteric artery occlusion caused an increase in the plasma level of calcitonin gene-related peptide-like immunoreactivity (CGRP-LI), an effect which was abolished by pretreatment with capsaicin. Similar protection was shown in the animals subjected to a brief period of anterior mesenteric artery occlusion 24 h before coronary artery occlusion, and this delayed protection was also abolished partly by pretreatment with capsaicin. Capsaicin treatment (50 mg/kg, s.c.) alone also protected the ischa-emic myocardium. The results suggest that brief ischaemia of the small intestine induces both early and delayed protection against reperfusion-induced myocardial injury, and the effects are, at least partly, related to the activation of capsaicin-sensitive sensory nerves. Received: 13 August 1998 / Accepted: 4 January 1999     

Excitatory and inhibitory urinary bladder reflexes induced by stimulation of cervicovaginal capsaicin-sensitive sensory fibers in rats

We have investigated the effect of intravaginal application of capsaicin on micturition reflex in female rats. Urinary bladder contractility was measured by transurethral pressure recording at isovolumetric and subthreshold conditions in anaesthetized rats. The intravaginal application of capsaicin (15 mug/50 mul rat) induced reproducible bladder phasic contractions, without desensitization upon repeated applications, that were blocked by intravenous atropine (1 mg/kg) or hexamethonium (5 mg/kg) and prevented by removal of paracervical ganglia or systemic capsaicin pretreatment (125 mg/kg, s.c.). The inhibition of sympathetic transmission by guanethidine (30 mg/kg, s.c.) produced significant increase of the bladder reflex contractions activated by intravaginal capsaicin. Intravenous administration of the TRPV1 antagonist, capsazepine (3 mg/kg), significantly reduced the excitatory reflex response to capsaicin. Intravaginal administration of capsaicin (15 mug/50 mul), during distension-induced reflex bladder contractions, produced a transient block of reflexes, unaffected by guanethidine pretreatment. In conclusion, the stimulation of capsaicin-sensitive sensory nerve endings in the rat cervix-vagina induced a dual excitatory or inhibitory bladder response in anaesthetized female rats depending on the degree of bladder distension.     

Effects of carbonyl cyanide p-trichloromethoxyphenylhydrazone (CCCP) and of ruthenium red (RR) on capsaicin-evoked neuropeptide release from peripheral terminals of primary afferent neurones

Summary In the superfused isolated rat urinary bladder, capsaicin as well as electrical field stimulation evoked the release of calcitonin gene-related peptide-like immunoreactivity (CGRP-IR). Carbonyl cyanide p-trichloromethoxyphenylhydrazone (CCCP, threshold 2 M) reduced both, the capsaicin- and the electrical field stimulation-evoked release of CGRP-IR while a low concentration of Ruthenium Red (RR, 0.6 M and 2 M) selectively attenuated the capsaicin-evoked release of CGRP-IR but did not influence the effect of electrical field stimulation. 20 M RR nearly abolished the capsaicin-evoked release, but also attenuated the effect of electrical field stimulation.In the isolated guinea-pig bronchus, electrical field stimulation and capsaicin induced non-cholinergic contractions which are known to be caused by tachykinin release from afferent nerve terminals. CCCP (0.6 M) only reduced the response to field stimulation; a ten-fold higher concentration of CCCP attenuated field stimulation as well as capsaicin-induced contractions. This is in contrast to the reported selective inhibition of capsaic-ininduced contractions by RR.The present data demonstrate that CCCP generally inhibits evoked neuropeptide release, regardless of the kind of stimulation used while low concentrations of RR preferentially inhibit capsaicin-evoked neuropeptide release.Send offprint requests to: R. Amann at the above address     

Pharmacologic responses of the mouse urinary bladder

The aim of the study was to determine pathways involved in contraction and relaxation of the mouse urinary bladder. Mouse bladder strips were set up in gassed Krebs-bicarbonate solution and responses to various drugs and electrical field stimulation were obtained. Isoprenaline (b-receptor agonist) caused a 63% inhibition of carbachol precontracted detrusor (EC50=2nM). Carbachol caused contraction (EC50=0.3μM), responses were antagonised more potently by 4-DAMP (M3-antagonist) than methoctramine (M2-antagonist). Electrical field stimulation caused contraction, which was inhibited by atropine (60%) and less by guanethidine and α,β-methylene-ATP. The neurogenic responses were not potentiated by inhibition of nitric oxide synthase. Presence of an intact urothelium significantly depressed responses to carbachol (p=0.02) and addition of indomethacin and L-NNA to remove prostaglandin and nitric oxide production respectively did not prevent the inhibitory effect of the urothelium. In conclusion, b-receptor agonists cause relaxation and muscarinic agonists cause contraction via the M3-receptor. Acetylcholine is the main neurotransmitter causing contraction while nitric oxide has a minor role. The mouse and human urothelium are similar in releasing a factor that inhibits contraction of the detrusor muscle which is unidentified but is not nitric oxide or a prostaglandin. Therefore, the mouse may be used as a model to study the lower urinary tract.     

Reflex activation of the adrenal medulla during hypoglycemia and circulatory dysregulations is regulated by capsaicin-sensitive afferents

Summary 1. Catecholamines were measured in the adrenal venous outflow during hypoglycemia, hypotension, hypovolaemia and efferent splanchnic nerve stimulation in anaesthetized capsaicin-pretreated rats and their vehicle controls. In control rats, efferent splanchnic nerve stimulation caused a marked rise in adrenaline and noradrenaline levels. In contrast, the other stimuli mainly elicited adrenaline release. 2. The release of adrenaline and noradrenaline evoked by splanchnic nerve stimulation was of the same magnitude in capsaicin-pretreated rats (whose afferent C-fibres were destroyed by this pretreatment) as in their untreated controls. 3. Capsaicin-pretreatment of rats resulted, however, in a reduced adrenaline release during insulin-induced hypoglycemia for up to 30 min and, as a consequence, generated a greater fall in blood glucose. The adrenal response to hypoglycemia in the first 30 min was also reduced by bilateral vagotomy indicating the existence of glucoreceptors on peripheral vagal terminals. 4. Adrenaline secretion following central glucopenia induced by 2-deoxy-d-glucose remained unaffected by capsaicin-pretreatment, indicating an intact function of the central regulation of adrenaline release. 5. Hypotension evoked either by sodium nitroprusside or by haemorrhage resulted in a pronounced increase in adrenaline release which was almost absent in the capsaicin-pretreated rats. 6. It is concluded that the stimulation of the adrenal medulla during hypoglycemia, hypotension, and hypovolaemia is based on a reflex mechanism initiated by capsaicin sensitive afferents.This work was supported by the Austrian Scientific Research Funds, grant No. P6646, the Austrian National Bank (grant No. 2811) and the Pain Research Commission of the Austrian Academy of Sciences Send offprint requests to J. Donnerer at the above address     

A novel pathway underlying the inhibitory effects of melatonin on isolated rat urinary bladder contraction

The aim of the present study was to elucidate the direct effects of melatonin on bladder activity and to determine the mechanisms responsible for the detrusor activity of melatonin in the isolated rat bladder. We evaluated the effects of melatonin on the contractions induced by phenylephrine (PE), acetylcholine (ACh), bethanechol (BCh), KCl, and electrical field stimulation (EFS) in 20 detrusor smooth muscle samples from Sprague-Dawley rats. To determine the mechanisms underlying the inhibitory responses to melatonin, melatonin-pretreated muscle strips were exposed to a calcium channel antagonist (verapamil), three potassium channel blockers [tetraethyl ammonium (TEA), 4-aminopyridine (4-AP), and glibenclamide], a direct voltage-dependent calcium channel opener (Bay K 8644), and a specific calcium/calmodulin-dependent kinase II (CaMKII) inhibitor (KN-93). Melatonin pretreatment (10(-8)~10(-6) M) decreased the contractile responses induced by PE (10(-9)~10(-4) M) and Ach (10(-9)~10(-4) M) in a dose-dependent manner. Melatonin (10(-7) M) also blocked contraction induced by high KCl ([KCl](ECF); 35 mM, 70 mM, 105 mM, and 140 mM) and EFS. Melatonin (10(-7) M) potentiated the relaxation response of the strips by verapamil, but other potassium channel blockers did not change melatonin activity. Melatonin pretreatment significantly decreased contractile responses induced by Bay K 8644 (10(-11)~10(-7) M). KN-93 enhanced melatonin-induced relaxation. The present results suggest that melatonin can inhibit bladder smooth muscle contraction through a voltage-dependent, calcium-antagonistic mechanism and through the inhibition of the calmodulin/CaMKII system.     

The contractile effect of bombesin on the rat isolated urinary bladder

We have investigated and compared the myotropic effects of bombesin (BB) and carbachol (C) in the rat isolated urinary bladder. BB (0.5 x 10(-9) to 0.5 x 10(-5) M) and C (2.7 x 10(-8) to 5.4 x 10(-5) M) were found to produce dose-dependent increases of the basal tone of the rat detrusor muscle. The maximal contraction produced by C was about 4 times greater than that elicited by BB or substance P (SP). However, the threshold concentrations of BB and SP required to stimulate the detrusor muscle were much lower than those of C. The pD2 (-log ED50) values of BB, SP and C were respectively 7.63, 7.05 and 5.8. The tissues exposed to BB relaxed more slowly after washout than those challenged with C or SP. The contractile effects of medium range concentrations of BB were not affected by pretreating the tissues with tetrodotoxin, atropine, antihistaminics, indomethacin, alpha- and beta-adrenergic blockers, methysergide or 8-leucine-angiotensin II. Tissues desensitized with high concentrations of bradykinin maintained their sensitivity to BB. The result suggest that the contractile effect of BB on the rat isolated urinary bladder is likely to be the result of a direct effect on the smooth muscle cells.     

Effects of testosterone on neuromuscular transmission in rat isolated urinary bladder

Testosterone was examined for its effects on neuromuscular transmission in rat and shrew urinary bladder. In isolated preparations of detrusor muscle from sexually immature male rats (8-10 weeks old) at concentrations of 100-300 microM, it inhibited neuromuscular transmission in a concentration-dependent manner and it also inhibited responses to applied carbachol and diadenosine pentaphosphate (Ap(5)A, a P2X receptor agonist). Ethanol (at or above 38 mM), the solvent for testosterone, also caused significant inhibition of neurogenic contractions as well as carbachol- and Ap(5)A-induced contractions. In older, sexually mature male rats (over 16 weeks old), testosterone and ethanol had similar effects to those observed in the young male rat, although both were slightly less potent. In young virgin female rats (8-12 weeks old), testosterone and ethanol inhibited neuromuscular transmission; testosterone was approximately 1000 times more potent than in male rats, with a threshold concentration of 30 nM. In the insectivore, Suncus murinus, testosterone (0.1 microM-1 x mM) caused inhibition of neurogenic and chemogenic responses, but ethanol had no significant effect. Flutamide (50 microM), a genomic testosterone-receptor antagonist, did not inhibit any of the responses to testosterone. It is concluded that testosterone acts predominantly on a postjunctional nongenomic receptor to inhibit urinary bladder detrusor muscle contraction.     

Capsaicin sensitive nerves in the jejunum of Nippostrongylus brasiliensis-sensitized rats participate in a cardiovascular depressor reflex

Summary Superfusion of capsaicin onto the serosal surface of jejunum of Nippostrongylus brasiliensis-sensitized rats induces a short-lasting (1–3 min), dose-dependent (2 to 20 g) decrease in blood pressure which ranges from –5.3±1.40% to –22.6±2.20%. The hypotension evoked by capsaicin was more marked in sensitized rats than in unsensitized animals, which responded only to the highest dose (20 mg) of capsaicin tested. The hypotensive effects of capsaicin were not affected by intravenous injections of mepyramine (10 mg/kg), a histamine receptor antagonist, or by the cycloxygenase inhibitor indomethacin (10 mg/kg). However, an intravenous injection of a platelet-activating factor (PAT) antagonist, BN 52021 (20 mg/kg), or an intraperitoneal injection of guanethidine (8 mg/kg) 18 h prior to experimentation, to functionally impair the sympathetic nerves, abolished the capsaicin-induced drop in blood pressure. Treatment of neonatal rats with capsaicin reduced by 75% the hypotensive effects of capsaicin, whereas the capsaicin antagonist, ruthenium red, reduced non-significantly the hypotensive action of capsaicin. It is concluded that the activation of jejunal sensory nerves in N. brasiliensis-sensitized rats by capsaicin induced a reflex hypotension that is dependent upon PAF release from mast cells and functional sympathetic nerves. In addition, the afferent function of the sensory nerves are not totally blocked by ruthenium red as capsaicin elicits the reflex hypotension in the presence of this blocker of sensory nerve efferent function.Correspondence to: R. Mathison     

Effects of ageing on muscarinic receptor subtypes and function in rat urinary bladder

We compared the density and function of M₂ and M₃ muscarinic acetylcholine receptor subtypes in the urinary bladder of young adult (3 months) and old (23 months) male Wistar rats. Old rats had a reduced density of muscarinic receptors (96±10 vs. 156±21 fmol/mg protein), but competition experiments with the M₃-selective darifenacin did not indicate alterations in the relative roles of M₂ and M₃ receptors, with the former being more abundant. The amount of immunodetectable -subunits of various G-proteins potentially linked to muscarinic receptor function was unchanged. The potency of carbachol to contract bladder strips was also unaltered; its maximum effects as well as those of a single KCl concentration were unchanged if raw data or those corrected for strip length were analysed, but somewhat reduced when those corrected for strip weight were analysed. Antagonistic effects of atropine, the M₂-selective Ro 320–6206 and the M₃-selective darifenacin were unchanged. Agonistic effects of the M₃-sparing agonist 4-(2-oxo-2,3-dihydro-benzoimidazol-1-yl)-[1,4]bipiperidinyl-1-carboxylic acid ethyl ester were similarly poor in young and old rats. Additional experiments were concomitantly performed in submandibular glands from the same animals. While total muscarinic receptor density in submandibular glands was not significantly affected by age (56±5 vs. 61±4 fmol/mg protein), the relative contribution of M₃ receptors significantly declined from 68±3% to 57±2% based upon darifenacin competition curves. We conclude that aged Wistar rats express fewer muscarinic receptors in their urinary bladder, but there is no change in the relative abundance of M₂ and M₃ receptors; this is accompanied by only minor if any alterations in receptor responsiveness. In contrast, submandibular gland expresses similar receptor numbers in young and old rats, but slightly fewer M₃ receptors in old animals.     

Motor and inflammatory effect of hyperosmolar solutions on the rat urinary bladder in relation to capsaicin-sensitive sensory nerves

1. Intravesical administration of hyperosmolar NaCl or urea solutions produced a concentration-dependent stimulatory action on the micturition reflex in urethane-anesthetized rats. This effect was not modified in rats pretreated with capsaicin as adults (50 mg/kg s.c. 4 days before). 2. Hyperosmolar NaCl also produced Evans blue leakage (plasma extravasation) in the rat bladder. This effect was greatly reduced by extrinsic bladder denervation and in rats desensitized to capsaicin as newborns but not as adults. 3. Cumulative addition of NaCl produced a concentration-dependent increase in tone and biphasic effects on neurogenic contractions of the rat isolated bladder. These effects were not modified by in vitro capsaicin desensitization. 4. These findings do not support the idea that true osmoreceptors are present in the rat urinary bladder. The neurogenic component of the inflammatory response to hyperosmolar NaCl could involve activation of a subpopulation of bladder sensory fibers susceptible to the neurotoxic action of capsaicin in the early postnatal period only.     

Effects of ochratoxin A on DNA repair in cultures of rat hepatocytes and porcine urinary bladder epithelial cells

In cultured rat hepatocytes the mycotoxin ochratoxin A (OTA) induced unscheduled DNA synthesis (UDS) only in a narrow concentration range. Using a culture medium supplemented with 1% fetal calf serum, at 750 nM OTA a weak induction and at 1 μM OTA a marked induction of DNA repair was observed (15 ± 11 and 38 ± 24% cells in repair, respectively). Concentrations >1 μM OTA were cytotoxic, and <750 nM no induction occurred. In cultures of cells from the urinary bladder (porcine urinary bladder epithelial cells; PUBEC), a target organ of the mycotoxin, OTA induced UDS in a concentration-dependent manner. To inhibit the proliferation of the cultured epithelial cells, which would counteract the detection of DNA repair, epidermal growth factor was omitted and an arginine-deficient medium (ADM) was used. Under these serum-free culture conditions the amount of cells undergoing DNA repair in PUBEC control cultures was ∼7±4%, a value also comparable to those of control cultures of rat hepatocytes. At concentrations between 250 nM and 1 μM OTA a concentration-dependent increase of cells in repair was observed. Above 1 μM OTA was cytotoxic. At this concentration a maximum of ∼61±9% of the cells undergo DNA repair. This amount is comparable to control cultures incubated with 5 or 10 mM ethylmethane-sulphonate (EMS) (49±9 and 69±10% cells in repair, respectively), used as a positive control. These results show that in cultured rat hepatocytes induction of UDS is relatively weak whereas in urothelial cells this effect was significant. Whether this effect is due to OTA metabolites formed locally in the urothelium cannot be excluded since PUBEC have been shown to be able to metabolize xenobiotics independently from the liver. Received: 13 May 1997 / Accepted: 11 June 1997     

通尿灵对兔膀胱出口梗阻后逼尿肌功能失调的保护作用

目的 探讨通尿灵对兔膀胱出口部分梗阻(BOO)后逼尿肌在形态学和酶学上的保护作用.方法 28只雄性白兔随机分为四组每组7只,A组为对照组(假手术组),B、C、D组为实验组,A组行手术但不造成梗阻,B、C、D组行手术造成膀胱出口部分梗阻.术后2周起,B组给予通尿灵100 mg · kg-1·d-1口服,C组给予花生油(通尿灵赋形剂),A、D组始终不给任何药物.再饲养3周后,解剖膀胱,取膀胱逼尿肌束标本用透射电镜观察逼尿肌超微结构以及用荧光分光度计分别对MDA含量和SOD、NOS、Ca2+ -Mg2+ -ATP酶的活力进行测定.结果 D组、C组与A组相比,膀胱逼尿肌细胞中内质网明显扩张、脱颗粒,线粒体水肿明显,空泡变性和线粒体内嵴减少、消失,有的细胞内可见大量溶酶体.B组与A组比较,仅见轻度内质网扩张和线粒体水肿.各组逼尿肌中检测到的SOD[(A组:(86.568±4.657)、B组:(89.218±4.430)、C组:(3.696±3.010)、D组:(32.258±2.001) U/mgprotein)];NOS[(A组:(12.871 ±1.240)、B组:(9.274±1.137)、C组:(2.365±0.358)、D组:(3.614±0.147)U/mgprotein)]以及Ca2+-Mg2+-ATP酶[(A组:(5.231±0.329)、B组:(5.362±0.266)、C组:(2.285±0.354)、D组:(2.654±0.307) μmolPi/mgprotein)],各组逼尿肌中检测到的MDA含量[(A组:(8.369±0.170)、B组:(10.358±0.383)、C组:(36.598±3.120)、D组:(42.574±2.009)nmol/mgp)].结论 通尿灵对梗阻后逼尿肌细胞的超微结构改变具有明显保护作用.     

The role of the capsaicin-sensitive innervation of the rat urinary bladder in the activation of micturition reflex

Capsaicin applied on the serosal surface of the urinary bladder in urethane-anaesthetized rats produces two distinct types of motor effects: a tetrodotoxin-, hexamethonium- and lidocaine-insensitive 'tonic' contraction and a series of tetrodotoxin-, hexamethonium- and lidocaine-sensitive rhythmic contractions. Both 'tonic' and rhythmic contractions are abolished by bladder denervation indicating their neurogenic origin. The rhythmic but not the 'tonic' component of the contractile effect of capsaicin is abolished by spinal cord transection indicating activation of a supraspinal micturition reflex. The motor effects of topical capsaicin are unaffected by pretreatment with indomethacin or diphenhydramine plus cimetidine. Pretreatment with a large dose of subcutaneous (SC) capsaicin increases both volume and pressure threshold for micturition while amplitude of micturition contraction is unaffected. Moreover the spinal somatovesical reflex elicited by pinching of the perineal skin is unaffected by capsaicin-desensitization. The intracerebroventricular (ICV) administration of capsaicin reproduces the effects of SC capsaicin on the bladder response to saline filling. Rats pretreated with ICV capsaicin are as sensitive as controls in reacting to noxious heat (hot plate test) while the wiping response to instillation of capsaicin into one eye was abolished. These findings provide functional evidence for the presence in the rat urinary bladder of a capsaicin-sensitive innervation which subserves a sensory function in relaying volume/pressure information from detrusor muscle to central nervous system. Information carried through these capsaicin-sensitive fibers appears to be relevant for initiation of a supraspinal vesico-vesical micturition reflex. Functional evidence indicates that these fibers may terminate at supraspinal level.     

A study of the atropine-resistant component of the neurogenic response of the rabbit urinary bladder

Rabbit bladder body was stimulated to contract by a number of agonists, of which bradykinin was the most potent, and ATP one of the least potent substances tested. The atropine-resistant component of the neurogenic response was unaffected by 2 X 10(-5) M chlorpheniramine or 10(-6) M methysergide, doses which suppressed responses to histamine or 5HT. Indomethacin 10(-5) M, or 10(-5) M capsaicin both reduced the atropine-resistant component. Following treatment with 10(-6) M atropine and 10(-5) M prazosin, 10(-4) M ANAPP3 produced a further suppression of the response, but did not antagonize the response to ATP. In the bladder body, the transmitter(s) responsible for the neurogenic response may be acetylcholine and prostaglandins and possibly ATP and substance P.