Muscarinic modulation of nitrergic neurotransmission in guinea-pig gastric fundus.

Muscarinic receptor activation by (4-Hydroxy-2-butynyl)-1-trimethylammonium-m-chlorocarbanilate chloride (McN-A-343) was investigated both on NADPH-d staining and on electrically induced responses in guinea-pig gastric fundus. McN-A-343 (10 micromol L(-1)) significantly increased the optical density of NADPH-d positive neurones, while blockade of nitric oxide synthase with N(omega)-nitro-L-arginine (L-NA) decreased it, suggesting facilitation of nitric oxide (NO) production. Electrical field stimulation (EFS; 2 Hz, 0.2 ms, supramaximal current intensity, 10 s train duration) elicited on-contraction followed by off-relaxation in the circular muscle strips. McN-A-343 (10 micromol L(-1)) transformed the EFS-evoked response from on-contraction into on-relaxation, which was neurogenic, tetrodotoxin-sensitive and hexamethonium-resistant. L-NA partly reduced the EFS-evoked relaxation, revealing two components: a nitrergic and a non-nitrergic one. The effect of McN-A-343 on the amplitude of the EFS-evoked relaxation was not changed by the M(3) receptor antagonist para-fluoro-hexahydro-sila-difenidol hydrochloride, but was significantly enhanced by M(1) receptor blockade with telenzepine. In the presence of telenzepine, the L-NA-dependent nitrergic component of the EFS-induced relaxation predominates. We suggest that cholinergic receptor activation has a dual effect on nitrergic neurotransmission: (i) stimulation of NOS by muscarinic receptor(s) different from M(1) and M(3) subtype, (ii) prejunctional inhibition of NO-mediated relaxation via M(1) receptors. In addition, M(1) receptors may facilitate the non-nitrergic relaxation.

Participation of muscarinic acetylcholine receptors in nitric oxide pathway in cat duodenum.

The effect of activation of muscarinic acetylcholine subtype M1 receptors on the electrical field stimulation (EFS)-induced contractions in cat duodenal muscle strips was investigated. EFS elicited two types of responses: a) on-contraction; and b) off-contraction. The EFS-evoked responses were tetrodotoxin- and atropine-sensitive. In all strips the muscarinic acetylcholine subtype M1 agonist (4-Hydroxy-2-butynyl)-1-trimethylammonium-m-chlorocarbanilate chloride (McN-A-343) increased the tone and the amplitude of the spontaneous contractions. On this background EFS produced an inhibition of the phasic contractions at switching on the stimulation, followed by off-contractions. The EFS-induced inhibition was removed by N omega-nitro-L-arginine and was partly overcome by L-arginine. The results suggest the participation of muscarinic acetylcholine subtipe M1 receptors in the mediation of nitrergic inhibitory responses in cat duodenum.

Role of different bombesin receptor subtypes mediating contractile activity in cat upper gastrointestinal tract.

Mammalian bombesin-like peptides, gastrin-releasing peptide (GRP) and neuromedin B (NMB) are known to increase the motility of different segments in the gut. The present study was carried out to identify the bombesin receptor subtypes mediating the contractions induced by exogenous bombesin-like peptides in muscle strips isolated from cat esophagus, fundus, and duodenum. Both GRP-10 and NMB evoked concentration-dependent contractions in circular strips of esophagus and fundus and in longitudinal strips of the duodenum. These contractions were tetrodotoxin- and atropine-resistant. The potency of NMB in esophageal strips was 33 times higher than that of GRP-10. The NMB-preferring receptor antagonists D-Nal-Cys-Tyr-D-Trp-Lys-Val-Cys-Nal-NH2 (SSocta) and D-Nal-cyclo[Cys-Tyr-D-Trp-Orn-Val-Cys]-Nal-NH2 (BIM-23127) shifted the NMB and GRP concentration-response curves to the right, while the GRP-preferring receptor antagonist [D-Phe6]Bombesin(6-13)-methyl-ester (BME) did not affect the response to the peptides. Isolated muscle strips from the cat fundus and duodenum showed a higher sensitivity to GRP-10 than to NMB. In both segments, BME shifted the GRP-10 and NMB concentration-response curves to the right, while SSocta had no effect. The antagonism of BME was competitive on duodenal but not competitive on fundic muscle. We conclude that the direct myogenic action of GRP-10 and NMB in the esophagus is mediated mainly via NMB-preferring receptors, while GRP-preferring receptors are responsible for the contractile responses to bombesin-like peptides in feline fundus and duodenum. Our data suggest that the GRP receptor population located on fundic muscle might be nonhomogeneous.

Participation of M1 receptors in NO pathway in cat ileum.

Electrical field stimulation (EFS) elicited two types of responses from longitudinal muscle strips of the distal cat ileum: contraction at switching on the stimulation (in 62% of the strips) and inhibition of the phasic contractions at switching on the stimulation followed by contraction in 38% of the strips. In all strips the muscarinic acetylcholine subtype M1-receptor agonist (4-Hydroxy-2-butynyl)-1-trimethylammonium-m-chlorocarbanilate chloride (McN-A-343) increased the tone and amplitude of the spontaneous contractions. On this background EFS elicited an inhibition of the phasic contractions at switching on the stimulation followed by contraction. Blockade of nitric oxide (NO)-synthase by Nw-nitro-L-arginine (L-NNA) transformed the inhibitory response into contraction and this transformation was partly overcome by L-arginine. The results suggest that presynaptically situated M1 receptors are involved in the NO pathway.

Non-adrenergic non-cholinergic neuron stimulation in the cat lower esophageal sphincter.

Both electrical field stimulation and nicotine produced non-adrenergic non-cholinergic (NANC) relaxation of the circular muscle strips from the cat lower esophageal sphincter in the presence of 5 microM guanethidine and 5 microM scopolamine. Low-frequency stimulation (2 Hz, 0.2 ms duration, supramaximal current intensity, 20-s train) provoked a transient relaxation, while at high-frequency stimulation (20 Hz) a slow restoration to the resting tone was observed. Blockade of nitric oxide (NO) synthesis by 1 mM N omega -nitro-L-arginine decreased by 20% the amplitude of the 20 Hz-induced relaxation and changed the pattern of relaxation, making it similar to the sustained relaxation evoked by exogenously applied vasoactive intestinal peptide (VIP). After chymotrypsin (4 U/ml), the pattern of the high-frequency-induced relaxation resembled that of the low-frequency-induced relaxation. Similarly, chymotrypsin changed the shape of nicotine-provoked relaxation, increasing the speed of restoration to the resting tone. We suggest that the fast relaxation elicited in cat lower esophageal sphincter by electrical field stimulation or nicotine is initiated by NO. The slow restoration to the resting tone in the case of high-frequency- or nicotine-induced relaxation seems to be due to the release of VIP or VIP-like peptides. The possibility of participation of another transmitter(s) involved in NANC relaxation should not be excluded.

Participation of nitric oxide in the nicotine-induced relaxation of the cat lower esophageal sphincter.

The participation of nitric oxide in the relaxation of the cat lower esophageal sphincter muscle strip in response to electrical field stimulation or administration of nicotine was studied. The nicotine-induced relaxation was mediated via a neuronal pathway, since it was inhibited by administration of hexamethonium or tetrodotoxin. Inhibition of nitric oxide biosynthesis by N-nitro-L-arginine decreased the relaxation induced by nicotine (50 microM) or field stimulation. With the maximal concentration of N-nitro-L-arginine (1 mM) electrical field stimulation-induced relaxation was abolished, while nicotine-induced relaxation decreased by 70%. L-Arginine (1 mM) partly restored this relaxation. Desensitization of P2x receptors by alpha, beta methylene-adenosine 5-triphosphate (alpha, beta-m-ATP) did not change the relaxation induced by either electrical field stimulation or administration of nicotine. It is therefore suggested that the field stimulation-induced relaxation is mediated by the release of nitric oxide, but in the nicotine-produced relaxation is only partly due to nitric oxide, other factor(s) might be also be involved.

Role of nitric oxide in mediating non-adrenergic non-cholinergic relaxation of the cat ileocecal sphincter.

The participation of nitric oxide (NO) in field stimulation- or nicotine-evoked non-adrenergic non-cholinergic (NANC) relaxation of cat ileocecal sphincter was studied in vitro. During a 30 microM noradrenaline-induced contraction, both the application of electrical field stimulation (2-20 Hz, 0.2 ms, supramaximal current intensity, 10 s duration) and (-)-nicotine (10-500 microM) produced a tetrodotoxin-sensitive relaxation. The maximal relaxation was observed at 10 Hz or 100 microM (-)-nicotine. In 12 out of 19 strips the pretreatment with N omega-nitro-L-arginine (100 microM) decreased the amplitude of the field stimulation-evoked relaxation, while in the remaining strips the relaxation was transformed into a contraction. By increasing the concentration of N omega-nitro-L-arginine up to 1 mM all strips responded to field stimulation with a frequency-dependent tetrodotoxin-resistant contraction. N omega-Nitro-L-arginine (100 microM) completely inhibited the nicotine-induced relaxation. L-Arginine (1 mM) restored the amplitude of both field stimulation- and nicotine-evoked relaxations. These data indicate that NO appears to be involved in both field stimulation- and nicotine-evoked NANC relaxations. Evidence has been obtained for the existence of tetrodotoxin-resistant NANC contraction in cat ileocecal sphincter.

Some characteristics of the muscularis mucosae of the cat lower esophageal sphincter.

1. The contractile activity of muscularis mucosae strips isolated from cat lower esophageal sphincter (LES) was studied. 2. LES muscularis mucosae strips were characterized by spontaneous phasic contractions. 3. Acetylcholine (ACh) at a threshold concentration of 10(-8) M dose-dependently increased the basal tone and decreased the amplitude of the phasic contractions. Noradrenaline (NA) also increased the basal tone but at a higher threshold concentration (10(-6) M). 4. Nicotine (N) at concentrations of 5 x 10(-5)-10(-4) M induced a relaxation of the LES muscularis mucosae. The N-induced relaxation was tetrodotoxin (TTX) (10(-6) M) sensitive and was insignificantly reduced after guanethidine (5 x 10(-6) M) plus scopolamine (5 x 10(-6) M) or propranolol (10(-6) M). The N-induced relaxation was strongly decreased by the blocker of nitric oxide (NO) synthesis N omega Nitro-L-Arginine (L-NNA) (10(-4) M). The relaxation was restored after addition of L-arginine (10(-3) M). 5. These results suggest the involvement of NO in the N-induced relaxation of the muscularis mucosae.

GRP-preferring bombesin receptor subtype mediates contractile activity in cat terminal ileum.

Mammalian bombesin-like peptides, such as gastrin-releasing peptide and neuromedin B, are known to increase motility of different segments in the gut. The present study was carried out to identify the receptor subtype mediating these contractions of ileal longitudinal muscles in cats, in vitro. Both gastrin-releasing peptide (GRP) and neuromedin B (NMB) evoked concentration-dependent contractions of the strips. [D-Phe6]Bombesin(6-13)-methyl-ester, a highly selective GRP-preferring receptor antagonist, competitively inhibited contractions induced by either agonist. On the other hand, D-Nal-Cys-Tyr-D-Trp-Lys-Val-Cys-Nal-NH2, a selective NMB receptor antagonist, did not affect the actions of either gastrin-releasing peptide or neuromedin B. Our results suggest that bombesin-like peptides contract cat terminal ileum via activating GRP receptors.

Effect of cromakalim on the smooth muscle of the cat gastric antrum.

The effect of the K(+)-channel opener cromakalim (BRL 34915) on the electrical and contractile activity of the smooth muscle of the cat gastric antrum has been studied. Cromakalim induced a concentration-dependent inhibition of the contractions and shortening of the sustained partial repolarization phase of the plateau action potential. High concentrations of cromakalim produced hyperpolarization and shortening of the repolarization and depolarization phases of the plateau action potential. The K(+)-channel blockers 4-aminopyridine (10(-2) M) and tetraethylammonium (10(-2) M) decreased the effect of cromakalim on the phasic contractions, while glibenclamide (5 x 10(-5) M) completely abolished it. We suggested that the inhibitory effect of cromakalim on the electrical and contractile activity of the gastric antrum smooth muscle is due to the cromakalim-induced increase of the outward K(+)-current through glibenclamide-dependent K(+)-channels.

[Gastrointestinal sphincters--pharmacologic viewpoints]. / Gastrointestinale Sphinkteren--pharmakologische Gesichtspunkte.

Bearing in mind the specificity of the functions of the gastrointestinal sphincters (GIS), we studied the responses of the lower esophageal sphincter (LES), the pyloric sphincter (PS), the ileocecal sphincter (ICS) and the internal anal sphincter (IAS) to noradrenaline (NA), acetylcholine (ACh), PGE1, PGF2 alpha and the peptide bombesin (B). The electrical and contractile activities as well as the response to field electrical stimulation (FES) were recorded. All sphincters except for PS responded to NA with depolarization and contractions. ACh at concentrations higher than 5 x 10(-6)M elicited a biphasic response from LES: depolarization followed by hyperpolarization and respectively contractions and relaxation. At concentrations higher than 10(-6)M ACh produced hyperpolarization and relaxation in IAS. Data were obtained about the modulating role of presynaptic N-cholinoreceptors in the release of a non-adrenergic, non-cholinergic (NANC) inhibitory neurotransmitter leading to hyperpolarization and relaxation in LES and IAS under the effect of exogenous ACh. PGE1 and PGF2 alpha increased the tone and decreased the FES-induced relaxation in LES. In PS PGE1 evoked relaxation and completely inhibited the response to FES. The pharmacological analysis showed that PGs modulated not only the adrenergic and cholinergic but also the NANC neurotransmission. Evidence is presented concerning the role of bombesin in the adrenergic, cholinergic, and NANC neurotransmission in LES.

[The effect of microinjections of bombesin into the amygdala on the slow-wave frequency of the gastroduodenal smooth muscles and on the migrating myoelectric complex]. / Vliianie mikroin''ektsii bombezina v mindalinu na chastotu medlennykh voln gladkikh myshts gastroduodenal'noi zony i migriruiushchii mioélektricheskii kompleks.

In dogs with electrodes implanted in the smooth muscle wall of the stomach and duodenum, microinjection of bombesin (5 micrograms) during the 2nd phase of the migrating myoelectric complex decreased frequency of slow waves in the stomach, leaving unaltered the frequency of slow waves in the duodenum. Bombesin caused a short-term inhibition of the gastric and intestinal spike activity followed by its increase, the active period of the migrating myoelectric complex being prolonged.

[Hormonal changes and cellular proliferation following partial resection of the pancreas]. / Khormonalni promeni i kletuchna proliferatsiia sled chastichna rezektsiia na pankreasa.

This study is an attempt to elucidate the histomorphologic changes in the different portions of the gastrointestinal tract, occurring after partial resection of the pancreas with occlusion of the canal of the pancreas that remains. Experiments were carried out on dogs. A number of gastrointestinal hormones were determined by radioimmunoassay. Serious hormonal disturbances after operative intervention on the pancreas were found. The hormonal derangements following pancreatic surgery were attributed to histomorphologic changes in the digestive system.

[Effect of bombesin on the frequency of slow potentials of the smooth muscles of the gastrointestinal tract and migrating myoelectric complex]. / Vliianie bombezina na chastotu medlennykh potentialov gladkikh myshts zheudochno-kishechnogo trakta i migriruiushchii mioélektricheskii kompleks.

In dogs with electrodes implanted in the smooth muscle wall of the stomach, small intestine and colon, i.v. administration of bombesin (50, 100, 200 and 300 ng/kg) during the 2nd phase of the migrating myoelectric complex (MMC) increased the dose-dependent frequency of slow potential in the small intestine and stomach leaving unchanged the frequency of slow potentials in the colon. Single doses of 50 or 200 ng/kg bombesin during the 2nd phase of the MMC caused a short-term inhibition of the gastric and intestinal spike activity followed by its increase, the active period of the MMC being prolonged.

Participation of cholinergic mechanisms in the realization of the effect of bombesin on the migrating myoelectrical complex.

The experiments were carried out on dogs with chronically implanted bipolar ball-shaped silver electrodes in the antral part of the stomach and duodenum. The effect of bombesin (injected intravenously in a dose of 200 ng/kg) on the migrating myoelectrical complex (MMC) was examined after pretreatment with hexamethonium (10 mg/kg) or atropin (100 micrograms/kg), or alpha-antagonists (phentolamine, 2 mg/kg) and beta-antagonists (propranolol, 2 mg/kg) of the adrenergic receptors. Introduced during the second MMC phase, bombesin caused inhibition of the spike activity, accompanied by an increase of the slow potential frequency. Prolonged intensified spike activity occurred 5-10 min later. Early brief appearance of spike potentials was observed after bombesin in the presence of hexamethonium, which was not manifested on the background of atropin. After hexamethonium or atropin, however, the phase of the late activation of the spike activity did not occur and there was no change in the effect of bombesin on the frequency of the slow potential. The alpha- and beta-antagonists of the adrenergic receptors do not change the effect of bombesin on the electrical activity of the stomach and the intestines.

Changes in the reactivity of the intrinsic nervous system of the stomach to hexamethonium after transthoracic vagotomy.

Experiments were carried out on seven breedless dogs with chronically implanted electrodes in the stomach. The effect of hexamethonium (in doses of 0.2, 0.3, 0.5, 1, 2, 3, 5, 7 and 10 mg/kg, i. v.) on the migrating myoelectrical complex (MMC) was studied before and after transthoracic vagotomy. Injected on the background of the first MMC phase, hexamethonium in doses of up to 2 mg/kg induces the appearance of spike potentials (SP). Applied on the background of spike activity, hexamethonium induces the appearance of spike activity of the type of the 3rd MMC phase. In a dose exceeding 2 mg/kg, introduced during SP activity, hexamethonium interrupts for different periods of time the SP generation and shortens the duration of the active MMC phases. On the background of increased spike activity characteristic of the stomach after vagotomy, 3 mg/kg hexamethonium have almost no effect on the electrical activity, whereas 10 mg/kg only reduce the percentage of slow waves accompanied by SP. It is probably that after vagotomy the excitability of the intrinsic nerve cells is increased, therefore even the high doses of the ganglionic blocker hexamethonium cannot inhibit entirely the active phases of MMC.

Changes in the migrating myoelectrical complex of canine gastro-intestinal tract depending on the dietary regime.

Experiments were made to study the effect of vegetable protein (soya) and animal protein (meat) on the migrating myoelectrical complex (MMC) in the stomach and small intestine of dogs with chronically implanted electrodes. MMC was tested under different dietary conditions: (1) Diet No 1 comprising animal protein; (2) Diet No 2 poor in protein and diet No 3 with vegetable protein (soya). In the case of protein-deficient dietary regime (diet No 2) disturbance in MMC is observed--spike activity is constantly recorded. First phase of MMC is not observed. The transition from diet No 2 to diet No 3 at the end of the first week is accompanied by a tendency toward the formation of brief rest periods, with a considerable increase in the duration of 100% spike activity. At the end of the 30th day the different MMC phases were not well differentiated. Upon transition from protein-poor regime to diet No 1 during the second week, the different MMC of the stomach and small intestine are very well expressed. The problem of the generation and propagation of MMC is discussed. MMC is considered as a manifestation of the "biological clock" which responds to every change taking place in the organism during its interaction with the environment.

[Relations between the electric activity of the pyloric sphincter and activities of the stomach and duodenum]. / Zavisimost' élektricheskoi aktivnosti piloricheskogo sfinktera ot aktivnosti zheludka i dvenadtsatiperstnoi kishki.

Indwelled electrodes led electrical activity from the stomach, pyloric sphincter and duodenum in hunger, after feeding or administration of carbachol and morphine as well as after transthoracic vagotomy. In intact resting animals the pyloric sphincter develops slow potentials in the rhythm of stomach antral portion and sometimes in the rhythm of duodenum. Acceleration of the slow potentials rhythm occurring after transthoracic vagotomy in the stomach antral portion, develops in the pyloric sphincter as well. Spike activity of the antral portion spreads over to the sphincter depending on amplitude, frequency and duration of the burst of spikes. Migrating myoelectrical complex develops successively in the stomach antral portion and in the pyloric sphincter. During increased activity of the duodenum (after carbachol or morphine) the bursts of spikes in the rhythm of the duodenum slow waves can occur simultaneously in the pyloric sphincter, too.

On the character of the electrical activity of the stomach, pyloric sphincter and duodenum of cats.

In cats with chronically implanted electrodes in the muscle wall of the gastro-intestinal tract, the electrical activity of the stomach (S) pyloric sphincter (PS) and duodenum (D) is characterized by cycles consisting of two phases: active--whereby 2 to 3 slow potential changes (SPC) are accompanied by spike potentials (SP) and a pause during which for 60-90 s only SPC are recorded. After bilateral transthoracic vagotomy there is a disruption of the cyclic character of the electrical activity in the investigated regions of the gastro-intestinal tract--SP from S and PS are recorded with each SPC. The active phase of the cycle from D is strongly prolonged at the expense of the shorter pause. This lasts for 1-2 days, after which there is a tendency towards shaping the cycles characteristic of the electrical activity of S, PS and D of cats. The pauses, however remain relatively short and the number of SP in the group is much greater. Gradually during the second month the character of the electrical activity of S, PS, and D approaches that prior to the vagotomy, but even until the 6th month after vagotomy the number of SP in the group is greater and the active phase of the cycle is longer. On the background of antagonists of the adrenoreceptors (propranolol--2 mg/kg and dihydroergotamine--300 micrograms/kg), the cyclic character of the electrical activity of S, PS and D is disturbed. The relatively rapid restoration of the cyclic character of cat S, PS and D after vagotomy is assumed to be the result of the organizing role of the intrinsic nervous system. The disturbance of the cyclic character after adrenoblockers demonstrates the significance of the balance between the cholinergic and adrenergic parts of the intrinsic nervous system for the realization of the cyclic character of the electrical activity of S, PS and D of cats.

Changes in the myoelectric complex of the stomach and the small intestine under the influence of the intrinsic nervous system.

Chronic experiments have been carried out on dogs with implanted silver bipolar ball-shaped electrodes on the muscle wall of the stomach and of the duodenum after transthoracic truncus vagotomy. The appearance of incoordination in the rhythm of the slow potentials of the corpus and antrum is followed by an increase in the spike activity of the stomach and duodenum. The percentage of the spike activity increases, the active periods of the myoelectric complex become longer, with a decrease and even obliteration of the quiescent periods. These changes are most pronounced during the second post-vagotomy month, after which the spike activity gradually decreases and quiescent periods appear again. Nevertheless, at the end of the 6th month after the vagotomy, the spike activity has higher values than before the vagotomy. Chemical sympathectomy with 6-OHDA results in 100 per cent spike activity of the stomach and duodenum and complete absence of quiescent periods. It is concluded that the intrinsic nervous system is capable of organizing the phases of the myoelectric complex in the cases of absence of influence of the extrinsic nervous system.