Histamine receptors in guinea-pig isolated urinary bladder

Spasmogenic action of histamine, 2-(2-aminoethyl) pyridine (H1 receptor agonist) and 4 methyl-histamine (H2 receptor agonist), have been studied in guinea pig isolated urinary bladder in the presence of mepyramine (H1 antagonist) and metiamide (H2 antagonist) to identify the presence of H1 and H2 receptors. The study suggested the presence of H1 as well as H2 receptors in this preparation.     

Mechanism of action of nicotine in isolated urinary bladder of guinea-pig.

1. Nicotine produced a transient contraction of isolated strips of guinea-pig urinary bladder. The response to nicotine was antagonized by the nicotinic receptor antagonist, hexamethonium but was insensitive to tetrodotoxin. 2. The nicotine-induced contraction was potentiated by the cholinesterase inhibitor, physostigmine, and was reduced to 50% and 70% by the muscarinic cholinoceptor antagonist, atropine and the sympathetic neurone blocking drug, guanethidine, respectively. Chemical denervation with 6-hydroxydopamine abolished the inhibitory effect of guanethidine. Simultaneous treatment with atropine and guanethidine did not abolish the response to nicotine, but the degree of inhibition was comparable to that obtained with atropine alone. 3. The nicotine-induced contraction was insensitive to bunazosin and yohimbine (alpha 1- and alpha 2-adrenoceptor antagonists, respectively), and exogenously applied noradrenaline did not cause a contraction even in the presence of blockade of noradrenaline uptake mechanisms with desipramine and normetanephrine and of beta-adrenoceptors with propranolol, suggesting a non-adrenergic nature of the sympathomimetic effect of nicotine in this tissue. 4. The nicotine-induced contraction in the presence of atropine was abolished after desensitization of P2-purinoceptors with alpha, beta-methylene adenosine 5'-triphosphate, a slowly degradable ATP analogue selective for P2-purinoceptors. By this desensitization, the response to ATP, but not to histamine, was also abolished. 5. A cyclo-oxygenase inhibitor flurbiprofen partially inhibited the nicotine-induced contraction. The degree of the inhibition was more pronounced in the presence of atropine than in its absence. Flurbiprofen antagonized the response to exogenously applied ATP in an unsurmountable manner, but not that to carbachol.(ABSTRACT TRUNCATED AT 250 WORDS)     

Blockade of kinin-induced responses of the guinea-pig isolated urinary bladder by the extract of Mandevilla velutina

1. The crude extract (CE) from M. velutina (0.2-0.8 mg/ml), antagonized in a dose-dependent and reversible manner bradykinin (BK)- and lysyl-BK-induced contractions of the guinea-pig urinary bladder in vitro. 2. Schild plots yielded nominal pA2 values (as g/ml) of 3.5 and 3.7, but the slopes were significantly greater than unity. 3. At concentrations 2-10-fold higher, the CE failed to affect responses to acetylcholine or histamine. 4. In electrically-stimulated tissue, BK, L-BK and M-L-BK potentiated nerve-mediated twitch responses and increased muscular tone. The B1 agonist [des-Arg9]-BK, was inactive up to 10 microM. 5. The CE from M. velutina (0.4 mg/ml) inhibited both actions of the kinins.     

Release of prostaglandin E from the isolated urinary bladder of the guinea-pig.

1 Release of prostaglandin E (PGE) from guinea-pig urinary bladder in vitro has been demonstrated both in the resting state and during electrical stimulation. 2 The electrically evoked release of PGE was significantly higher than the resting release and was frequency-dependent. 3 The released substance was characterized as PGE pharmacologically by (a) blockade of its response by SC-19220 on guinea-pig ileum, (b) reduction of the amount of the released substance by indomethacin and (c) the inhibitory effect of the released substance on adrenergic neurotransmission in guinea-pig vas deferens. 4 The prostaglandin seemed to originate from the muscle since tetrodotoxin treatment did not abolish the release during direct muscle stimulation; however, concomitant release from neuronal tissue could not be excluded in the present experiments. 5 Indomethacin failed to inhibit the mechanical responses of the bladder to transmural stimulation. 6 The present experiments suggest that PGE is not involved in mediating the non-cholinergic non-adrenergic neurotransmission in the guinea-pig urinary bladder.     

Adrenoreceptors of the guinea-pig urinary bladder.

1 Adrenaline, noradrenaline and isoprenaline (5 mug/ml) did not affect the resting tone of the isolated urinary bladder of the guinea-pig. 2 The catecholamines (1-2 mug/ml) inhibited neuronally evoked contractions at various stimulation frequencies; the inhibition was maximum at 2 Hz and minimum at 50 Hz. Isoprenaline produced maximum inhibition. 3 Propranolol (0.5 mug/ml) completely blocked the catecholamine-induced inhibition at all the frequencies employed. The concentration-response curves of isoprenaline at 2, 10 and 50 Hz were characteristically shifted by propranolol (50 ng/ml). Phenoxybenzamine (0.2 mug/ml) was totally ineffective. 4 In some experiments adrenaline significantly raised the tone of the bladder exposed to propranolol; this effect could be blocked by phenoxybenzamine. 5 Acetylcholine-induced bladder contractions were inhibited by adrenaline (2 mug/ml); the inhibition was completely blocked by propranolol (0.5 mug/ml). 6 The results indicate the presence of an inhibitory beta-adrenoceptor and suggest the possibility of an excitatory alpha-adrenoceptor in guinea-pig urinary bladder.     

Pharmacological analysis of the local and reflex responses to bradykinin on rat urinary bladder motility in vivo.

1. The topical application of bradykinin (BK) (0.05-5000 pmol/rat) onto the serosal surface of the urinary bladder in urethane-anaesthetized rats, evoked low amplitude tonic contractions (not exceeding 25 mmHg) or high amplitude (about 50 mmHg), phasic reflex contractions (chemoceptive micturition reflex) which were abolished by bilateral ablation of the pelvic ganglia. In ganglionectomized rats, BK induced only a local, tonic-type contraction. 2. Systemic capsaicin pretreatment (164 mumol kg-1, 4 days before) reduced the incidence of chemoceptive reflex induced by BK (500 pmol/rat) but had no effect on the magnitude of the tonic-type contraction elicited by BK in ganglionectomized rats. Indomethacin (11 mumol kg-1, 20 min before) reduced the incidence but not the amplitude of the reflex contractions induced by topical application of BK (500 pmol/rat). In ganglionectomized rats, indomethacin (11 mumol kg-1, 20 min before) decreased the amplitude of the tonic contraction evoked by BK. Indomethacin did not affect the chemoceptive reflex induced by topical application of capsaicin (15 nmol/rat) onto the bladder. 3. Intrathecal administration of the tachykinin NK1 receptor antagonists, RP 67,580 (10 nmol/rat) or SR 140,333 (10 nmol/rat), abolished the chemoceptive reflex induced by BK without modifying the magnitude of the tonic contraction. SR 140,333 (10 nmol/rat) also abolished the occurrence of the chemoceptive reflex induced by capsaicin. 4. Intravenous administration of the B2 receptor antagonist, Hoe 140 (35 nmol kg-1, 10 min before) abolished the reflex and local effects induced by BK on bladder motility but failed to modify the chemoceptive reflex induced by topical application of capsaicin (15 nmol/rat).(ABSTRACT TRUNCATED AT 250 WORDS)     

Contribution of cyclooxygenase-dependent mechanisms to contractile responses to donepezil in the rat urinary bladder

Donepezil, an inhibitor of acetylcholinesterase, is used to improve cholinergic neurotransmission and cognitive function in Alzheimer's disease. In the present study, contractile effects of donepezil on the rat urinary bladder were examined and compared with those of the nonselective cholinesterase inhibitor neostigmine. Both donepezil and neostigmine produced concentration-dependent contractile responses of the isolated rat urinary bladder strips. The neostigmine-induced contractions were abolished by atropine. However, donepezil produced contractions of urinary bladders partly through atropine-insensitive mechanisms. The atropine-resistant component of donepezil-induced contraction was significantly reduced by the cyclooxygenase inhibitor indomethacin. These results suggest that cyclooxygenase-derived prostanoids contribute, at least in part, to contractile effects of donepezil in the rat urinary bladder.     

Beta-adrenoceptor subtypes in the detrusor of guinea-pig urinary bladder.

beta-Adrenoceptors have been demonstrated in the urinary bladders of many animals including the guinea pig. However, there is little information on the subtypes involved in the antispasmodic activity of beta-adrenoceptor activation in the guinea-pig detrusor. The present study uses the non-selective beta-agonist isoproterenol, the antagonist nadolol, the beta 2-selective agonists salbutamol and terbutaline, the antagonist ICI 118551, and the beta 1-selective antagonist metoprolol, to demonstrate functionally the subtypes existing in the guinea-pig detrusor. Isoproterenol dose-dependently reduces the myogenic activity in the guinea-pig detrusor induced by mild depolarization with 20 mM potassium in the tissue bath. At the supramaximal concentration of 30 microM, isoproterenol achieves 73 +/- 2% of the reference maximal response. This activity of isoproterenol is reduced to 9 +/- 5, 24 +/- 6 and 54 +/- 1% in the total blockade of beta, beta 1 and beta 2 with nadolol, metoprolol and ICI 118551, respectively. Consistently, salbutamol and terbutaline at the same concentration produce only 35 +/- 1 and 38 +/- 4% of the response, respectively. Thus, both beta 1- and beta 2-adrenoceptors are present in the detrusor of the guinea-pig urinary bladder. Although activation of either subtype results in antispasmodic action, the larger portion of the antispasmodic activity appears to be associated with the activation of the beta 1-subtype.     

Multiple tachykinin binding sites in hamster, rat and guinea-pig urinary bladder

Binding of the 125I-Bolton-Hunter labelled tachykinins substance P, substance K, eledoisin and neuromedin K (BHSP, BHSK, BHE, BHNK) was examined in urinary bladders of hamster, rat and guinea-pig using crude membrane suspensions and by autoradiography. High-affinity binding of BHSK was observed in hamster and rat bladder and high-affinity binding of BHSP was seen in rat and guinea-pig bladder. Characterization of this binding showed that the hamster bladder contains very large numbers of K-type binding sites, where BHSK is displaced by substance K greater than kassinin greater than eledoisin greater than neuromedin K greater than substance P greater than physalaemin, and has very few P-type binding sites, where BHSP is displaced by substance P greater than substance K much greater than neuromedin K. In contrast, the rat bladder contains moderate and approximately equal numbers of both K (KD, 0.74 nM; Bmax 2.9 fmol/mg wet weight tissue) and P (KD, 0.12 +/- 0.01 nM; Bmax 2.6 +/- 0.2 fmol/mg wet weight tissue) sites. The guinea-pig bladder possesses predominantly P sites. Most tachykinin binding sites are localized over smooth muscle and probably represent functional receptors mediating the direct contractile effects of tachykinins in these tissues. Few E-type binding sites, as previously described in rat brain, were found, although some BHNK binding sites were seen in the mucosa of guinea-pig bladder.     

Effects of propiverine hydrochloride on the spontaneous contractions of isolated guinea-pig urinary bladder strip and rhythmic urinary bladder contractions of anesthetized dog.

The effects of propiverine hydrochloride (P-4, CAS 60569-19-9), a new drug to treat pollakiuria, was investigated on the spontaneous contractions of isolated guinea-pig urinary bladder strip and rhythmic urinary bladder contractions of anesthetized dog. At 10(-6)-10(-5) mol/l P-4 raised the base line of an isolated guinea-pig urinary bladder strip and accelerated its spontaneous contraction. At 10(-4) mol/l P-4 raised, and then lowered the baseline, and accelerated then suppressed its spontaneous contractions. Papaverine at 10(-6)-10(-4) mol/l also showed a similar action as P-4 in the isolated guinea-pig urinary bladder strip. Flavoxate at 10(-6)-10(-4) mol/l raised its base line and accelerated its spontaneous contractions. Those of P-4 at 10(-5) mol/l were not inhibited by tetrodotoxin 10(-6) mol/l). At doses of 50 mg/kg or more, intraduodenal administration of P-4 suppressed the frequency of rhythmic urinary bladder contractions of anesthetized dog in a dose-dependent manner. These results indicate that P-4 shows mainly an accelerating action on the endogenous spontaneous contractions of urinary bladder, but on exogenous contractions induced by the Balloon's method it shows an suppressing action and regulates the functions of the urinary bladder, so P-4 might become a useful drug for the clinical treatment of micturitional dysfunction, for example, pollakiuria.     

Nitric oxide pathway-mediated relaxant effect of bradykinin in the guinea-pig isolated trachea.

1. The effects of two nitric oxide (NO) biosynthesis-inhibitors NG-nitro-L-arginine (L-NOARG) and NG-monomethyl-L-arginine (L-NMMA) on the relaxation induced by bradykinin (BK, 100 nM), isoprenaline (Iso, 1 microM) and sodium nitroprusside (SNP, 1 microM) were investigated in epithelium-intact strips of guinea-pig isolated trachea. 2. Relaxations induced by BK (100 nM) in guinea-pig tracheal strips under spontaneous tone were inhibited in a concentration-related manner by L-NOARG and L-NMMA (1 to 100 microM), with IC50s (and 95% confidence limits) of 9.1 (6.9-11.6) microM and 7.0 (4.2-12.3) microM, respectively. However, at the maximal concentration (100 microM) used, neither of these drugs inhibited completely BK-induced relaxation (maximal inhibition of 74 +/- 7 and 67 +/- 7%, respectively). On the other hand, D-NMMA, the D-enantiomer of L-NMMA, up to 100 microM failed to inhibit BK-induced relaxation. The relaxation induced by Iso (1 microM) and SNP (1 microM) were not affected by either L-NOARG or L-NMMA (30 microM). 3. The inhibition of BK-induced relaxation caused by L-NOARG and L-NMMA was partially reversed by addition of excess of L-arginine but not D-arginine (1 mM). 4. Like L-NOARG and L-NMMA, methylene blue (10 microM), an agent that inhibits the activation of soluble guanylate cyclase by NO, also significantly inhibited BK-induced relaxation, leaving responses to Iso unaffected. 5. Indomethacin (0.3 nM to 10 nM), a cyclo-oxygenase inhibitor, concentration-dependently inhibited BK-mediated relaxation, with an IC50 of 2.6 (1.7-3.8) nM, without affecting Iso and SNP-mediated relaxant responses. 6. A combination of a very low concentration of indomethacin (1 nM) and either L-NOARG or L-NMMA (100 microM) changed the response of tracheal preparations to BK (100 nM) from a relaxation to a sustained contraction. 7. These findings indicate that BK-induced relaxation in guinea-pig trachea is mediated jointly by the release of NO or a NO-related substance and a prostanoid, probably prostaglandin E2.     

Purinergic innervation of the guinea-pig urinary bladder

1 A number of criteria for considering adenosine 5′-triphosphate (ATP) as a neurotransmitter in the guinea-pig urinary bladder have been examined. In addition, the effect of tachyphylaxis to ATP on the response to non-adrenergic, non-cholinergic nerve stimulation has been re-examined.

2 Quinacrine fluorescence histochemistry revealed a population of nerve fibres, ganglion cells, and nerve bundles in the bladder which were not seen in either the iris or vas deferens, where adrenergic and cholinergic nerves predominate. The distribution and morphology of the quinacrine-positive nerves in the bladder were different from those observed with catecholamine fluorescence and cholinesterase histochemistry, and were unaffected by chemical sympathectomy.

3 Release of ATP from the bladder during stimulation of intramural excitatory nerves, in the presence of atropine and guanethidine increased to 3-12 times prestimulation levels. Tetrodotoxin abolished both the contractile response and the increase in ATP release resulting from intramural nerve stimulation. There was no increase in ATP release during contraction resulting from direct muscle stimulation following nerve paralysis with tetrodotoxin.

4 Sympathectomy with 6-hydroxydopamine did not affect release of ATP in response to intramural nerve stimulation.

5 Release of ATP was dependent on the concentration of calcium ion in the medium.

6 Contractions in response to non-adrenergic, non-cholinergic intramural nerve stimulation were closely mimicked by ATP, but not by acetylcholine or histamine.

7 Adenosine and dipyridamole reduced the contractions to both ATP and non-cholinergic nerve stimulation.

8 2-2′-Pyridylisatogen was not a specific blocker of either ATP or intramural nerve stimulation in the guinea-pig bladder. 2-Substituted imidazolines initiated spontaneous activity making it impossible to assess any blocking action that they may have had.

9 Prostaglandins (E₁, E₂ and F_2α) gave weak, slow contractions and an increase in spontaneous activity. Both the response to ATP and non-adrenergic, non-cholinergic nerve stimulation were greatly potentiated in the presence of prostaglandins.

10 In the presence of indomethacin the response to non-adrenergic, non-cholinergic nerve stimulation was virtually abolished following desensitization to ATP.

     

The properties of the ATP-induced depolarization and current in single cells isolated from the guinea-pig urinary bladder.

1. The actions of exogenously applied ATP were investigated with the whole-cell patch clamp method in single cells isolated from guinea-pig urinary bladder with a modified concentration jump technique. 2. Rapid application of ATP (threshold ca. 100 nM) depolarized the cell membrane with superimposition of action potentials which was followed by transient hyperpolarization. In the presence of D600, the amplitude of the ATP-induced depolarization was a function of the ATP concentration (EC50: 0.5-1 microM). 3. ATP activated a dose-dependent inward current with a short latency (18 ms with 10 microM ATP; measured as the time between the start of application and 10% of the peak). The relationship of the peak current versus ATP concentration was well fitted by a Michaelis-Menten equation with a Hill coefficient (n) of 1.7 and a dissociation constant (Kd) of 2.3 microM. The current desensitized rapidly and the time course of desensitization was a function of the ATP concentration and could be fitted by two exponentials. 4. The reversal potential of the ATP-activated current was near 0 mV. Replacement of extracellular Na by other monovalent or divalent cations indicated that the current flows through nonselective cation channels. 5. alpha,beta-Methylene ATP also produced a dose-dependent inward current but was less potent than ATP (n: 1.6, Kd: 10.4 microM). alpha,beta-Methylene ATP blocked the response to ATP by desensitization of the receptor. 6. alpha,beta-Methylene ATP was 50-100 times more potent than ATP at eliciting a contractile response of strips of detrusor smooth muscle. 7. The relevance of the above results to the possible role of ATP as the fast excitatory transmitter is discussed.     

The biphasic response of the isolated guinea-pig ileum by bradykinin

The isolated guinea-pig ileum, challenged by agonist, was used to study the effect of bradykinin. In the presence of acetylcholine producing approximately 60% of maximum contraction, bradykinin caused relaxation followed by contraction. The biphasic response to bradykinin was also found in the presence of histamine, eledoisin, angiotensin, prostaglandin F_2α and transmural electrical stimulation. The conditions for bradykinin-induced relaxation were not found after treatment by bradykinin, and potassium or barium chloride. Under conditions where bradykinin produced a biphasic response, acetylcholine, histamine, eledoisin, angiotensin, prostaglandin F_2α and lysine-vasopressin only contracted the ileum, while adrenaline, noradrenaline, oxytocin, calcium and magnesium chloride only relaxed. On increasing the percentage of maximum contraction with acetylcholine, an inverse relationship with relaxation by bradykinin was found. Tachyphylaxis was not present with the bradykinin-induced relaxation. The relaxing effect of bradykinin is more likely to be due to a direct action on the muscle cell membrane than to a release of a mediator or to blockade of a receptor mediating contraction.     

The role of sensory neuropeptides in motor innervation of the hamster isolated urinary bladder

In this study we have characterized the role of sensory fibers and of the sensory peptides, neurokinin A (NKA) and calcitonin gene-related peptide (CGRP), on the contractile responses evoked by single pulse electrical field stimulation (EFS) in the hamster urinary bladder. EFS of the hamster isolated urinary bladder produced twitch contractions which were unaffected by atropine but abolished by tetrodotoxin. The P2 purinoreceptor antagonist PPADS (30 microM) inhibited twitches by 66+/-4% on its own and by 78+/-3% in the presence of atropine. The selective tachykinin NK2 receptor antagonist nepadutant produced a slight but consistent reduction of twitch amplitude (-21+/-3%) at 1 microM. Addition of nepadutant to atropine and PPADS did not further increase their inhibitory effect. The application of hCGRP (10-300 nM) produced a concentration-dependent inhibition of twitches (Emax -38+/-3%, EC50=12 nM) and a small reduction of tone (0.5+/-0.09 mN). Similar effects were obtained with capsaicin (0.1-10 microM) which inhibited EFS-evoked contractions with an EC50 of 100.0 nM and a maximal effect of 34+/-4% inhibition at 1 microM. Under submaximal parameters of stimulation NKA (10 nM) increased the amplitude of twitches by 45+/-6% and produced a concentration-dependent tonic contraction (EC50=55.9 nM). The CGRP1 receptor subtype antagonist, hCGRP(8-37), increased by 29+/-8% the EFS-evoked contractions and significantly reduced the response to 0.1 microM CGRP. Capsaicin (10 microM) increased both CGRP-LI and NKA-LI release from superfused slices of hamster urinary bladder by about sixfold and by about 70%, over baseline, respectively. A second application of capsaicin was ineffective, indicating a complete desensitization of sensory nerve efferent function. In the hamster urinary bladder the sensory neuropeptides NKA and CGRP are co-released by sensory fibers after stimulation either by EFS or capsaicin. However, the role of CGRP appears functionally predominant.