Syntheses and antifungal activities of novel 3-amido bearing pseudomycin analogues.

As a result of our core SAR effort, we discovered a large number of 3-amido pseudomycin B (PSB) analogues (e.g., 4e LY448212 and 5b LY448731) that retain good in vitro and in vivo (IP) activities against Candida and Cryptococcus without inherent tail vein irritation. Several dimethylamino termini bearing 3-amides (e.g., 5b) also exhibited improved potency against Aspergillus in vitro. When evaluated in a two-week rat toxicology study, it was found that all animals receiving 4e (up to 75 mg/kg) were found to be normal. On the basis of these observations, we are convinced that it is possible to broaden the antifungal spectrum and improve the safety profile of pseudomycin analogues at the same time.

Pharmacology of butylthio[2.2.2] (LY297802/NNC11-1053): a novel analgesic with mixed muscarinic receptor agonist and antagonist activity.

Butylthio[2.2.2], ((+)-(S)-3-(4-butylthio-1,2,5-thiadiazol-3-yl)-1-azabicyclo[2.2.2] octane; LY297802/NNC11-1053) is a muscarinic receptor ligand which is equiefficacious to morphine in producing antinociception. In vitro, butylthio[2.2.2] had high affinity for muscarinic receptors in brain homogenates, but had substantially less or no affinity for several other neurotransmiter receptors and uptake sites. In isolated tissues, butylthio[2.2.2] was an agonist with high affinity for M1 receptors in the rabbit vas deferens (IC50 = 0.33 nM), but it was an antagonist at M2 receptors in guinea pig atria (pA2 = 6.9) and at M3 receptors in guinea pig urinary bladder (pA2 = 7.4) and a weak partial agonist in guinea pig ileum, which contains a heterogeneous population of muscarinic receptors. In vivo, butylthio[2.2.2] was without effect on acetylcholine, dopamine and serotonin levels in rat brain. Moreover, butylthio[2.2.2] did not decrease charcoal meal transit in mice, nor did it significantly alter heart rate in rats. Further, butylthio[2.2.2] did not produce parasympathomimetic effects such as salivation or tremor in mice, but it antagonized salivation and tremor produced by the nonselective muscarinic agonist oxotremorine. The present data demonstrate that butylthio[2.2.2] is a novel muscarinic receptor mixed agonist/antagonist and its pharmacological profile suggests that it may have clinical utility in the management of pain as an alternative to opioids.

Antagonism of serotonin3 (5-HT3) receptors within the blood-brain barrier prevents cisplatin-induced emesis in dogs.

Recently discovered serotonin3 (5-HT3) receptor antagonists are potent antiemetics in cytotoxic drug-induced vomiting. The specific site where 5-HT3 receptor antagonists act to abolish emesis is controversial. The major objective of this study was to determine whether the antiemetic effect of 5-HT3 receptor antagonists is exerted in the brain areas that reside inside or outside of the blood-brain barrier. Tropisetron, zatosetron (LY277359 maleate) and its quaternary analog zatosetron-QUAT were used in this study. Zatosetron and zatosetron-QUAT showed high affinity and selectivity for 5-HT3 receptors in radioligand binding studies. Both compounds antagonized 5-HT-induced bradycardia in rats with an approximate ID50 of 0.7 and 0.2 microgram/kg i.v., respectively. Zatosetron and tropisetron significantly inhibited cisplatin-evoked emesis in dogs (estimated ID50 values of 34.4 +/- 2.3 micrograms/kg and 108.3 +/- 4.8 micrograms/kg i.v., respectively). Zatosetron-QUAT (0.01-1.0 mg/kg i.v.) had no effect. [14C]-zatosetron-QUAT (100 micrograms/kg) was not detected in the brain after i.v. administration to rats, consistent with the inability of charged compounds to achieve significant brain concentrations. However, i.c.v. administration (100 ng/kg) of zatosetron-QUAT reduced emetic episodes significantly (11.6 +/- 1.6 vs. 2.8 +/- 1.2). These studies suggest that, in dogs, antagonism of 5-HT3 receptors located within the blood-brain barrier is important to block cisplatin-induced emesis.

Discovery of a potent, peripherally selective trans-3,4-dimethyl-4-(3-hydroxyphenyl)piperidine opioid antagonist for the treatment of gastrointestinal motility disorders.

Structure-activity relationship studies were pursued within N-substituted-trans-3,4-dimethyl-4-(3-hydroxyphenyl)piperidines in an effort to discover a peripherally selective opioid antagonist with high activity following systemic administration. Altering the size and the polarity of the N-substituent led to the discovery of 3 (LY246736). Compound 3 has high affinity for opioid receptors (Ki = 0.77, 40, and 4.4 nM for mu, kappa, and delta receptors, respectively). It is a potent mu receptor antagonist following parenteral and oral administration and distributes selectively (> 200-fold selectivity) to peripheral receptors. Thus, 3 has properties suitable for the clinical investigation of mu opioid receptor involvement in GI motility disorders.

Effects of LY267108, an erythromycin analogue derivative, on lower esophageal sphincter function in the cat.

BACKGROUND/AIMS: Erythromycin (EM-A) and some of its analogues stimulate gastrointestinal smooth muscle contractions. Because gastroesophageal reflux disease (GERD) in humans is in part caused by a reduction in lower esophageal sphincter (LES) pressure, the aim of this study was to investigate the effect of LY267108 (an EM-A analogue with no significant antimicrobial activity) on LES function. METHODS: In ketamine-anesthetized cats, LES pressure was recorded using a Dent sleeve. RESULTS: In cats, LY267108 increased LES pressure, as did motilin and EM-A. Neither LY267108, EM-A, nor motilin altered LES relaxation in response to a swallow. LY267108 increased LES pressure in cats in which the basal LES pressure was lowered experimentally by perfusing the distal esophagus with HCl (0.1 N for 3 days) or following isoproterenol (3.0 micrograms/kg intravenously). In summary, LY267108 increases LES pressure in normal cats, did not affect the relaxation of the LES in response to a swallow, and increases LES pressure in animals with an experimentally induced decrease in LES pressure. CONCLUSIONS: The results suggest that LY267108 may be useful in treating GERD because of its ability to increase LES pressure and thus present a barrier for gastroesophageal reflux.

Effect of zatosetron on ipecac-induced emesis in dogs and healthy men.

Serotonin receptor (5-HT3) antagonists provide effective antiemetic therapy in cancer patients receiving emetogenic chemotherapy, such as cisplatin. Animal studies have shown that 5-HT3 receptor antagonists also have antiemetic activity in ipecac-induced emesis. The authors investigated the antiemetic activity of zatosetron maleate, a 5-HT3 receptor antagonist, on ipecac-induced emesis in dogs and healthy men. They also evaluated the effect of ipecac administration on serotonin release and metabolism by measuring urinary 5-hydroxyindoleacetic acid (5-HIAA) excretion in healthy men. In separate randomized, placebo-controlled trials, 20 dogs received zatosetron intravenously and eight healthy men received zatosetron (50 mg) orally, followed by ipecac syrup. In both trials, emetic response to ipecac was recorded, including the number and time of vomits and retches. Zatosetron treatment inhibited and delayed ipecac-induced emesis in both groups. In dogs, zatosetron inhibited ipecac-induced emesis in a dose-dependent manner with a 100-micrograms/kg dose producing complete inhibition. In men, zatosetron administration resulted in fewer emetic episodes after ipecac than had occurred with placebo administration (P = .03); vomiting was completely inhibited by zatosetron. In men, ipecac administration did not affect the urinary 5-HIAA/creatinine ratio (mg/g) or 5-HIAA excretion rate (microgram/hour). Our study demonstrates that zatosetron has similar efficacy on ipecac-induced emesis in healthy men, as has been shown previously with other 5-HT3 receptor antagonists in chemotherapy-induced emesis in cancer patients. We did not observe the increase of urinary 5-HIAA in our study with ipecac-induced emesis, however, as has been described previously in cisplatin-induced emesis.(ABSTRACT TRUNCATED AT 250 WORDS)

LY277359 maleate: a potent and selective 5-HT3 receptor antagonist without gastroprokinetic activity.

Several 5-hydroxytryptamine (5-HT3) receptor antagonists have been described. In addition to 5-HT3 receptor antagonist activity, many of these agents also possess gastroprokinetic activity. In the present report, we identify compound LY277359 maleate as a potent, p.o. active, highly selective 5-HT3 receptor antagonist lacking gastroprokinetic effects. LY277359 maleate was a potent and selective antagonist of 2-methyl 5-HT-induced contraction in the guinea pig ileum (KB = 1.6 nM), a response mediated by activation of 5-HT3 receptors. Given both i.v. (0.0003, 0.001 and 0.003 mg/kg) and p.o. (0.01 and 0.03 mg/kg), LY277359 maleate inhibited the bradycardic response to i.v. administered 5-HT in urethane-anesthetized rats. The duration of antagonism of 5-HT-induced bradycardia persisted beyond 6 hr after p.o. administration of LY277359 maleate (0.03 mg/kg p.o.). In contrast to its potent 5-HT3 receptor antagonist activity, LY277359 maleate, in doses up to 1 mg/kg p.o., did not affect gastric emptying in rats, suggesting minimal, if any, gastroprokinetic activity of LY277359 maleate. This is in contrast to another 5-HT3 receptor antagonist, zacopride, which did produce a marked increase in gastric emptying in rats at doses of 0.1 mg/kg p.o. and higher. LY277359 maleate (0.03 and 0.1 mg/kg i.v. and 0.07, 0.1, 0.3 and 1.0 mg/kg p.o.) did have effects consistent with other 5-HT3 antagonists, to inhibit cisplatin-evoked emesis in dogs.(ABSTRACT TRUNCATED AT 250 WORDS)

Preclinical studies on quinelorane, a potent and highly selective D2-dopaminergic agonist.

Quinelorane (LY163502) has the endocrine, neurochemical and behavioral profile of a potent and highly selective D2-dopaminergic agonist. The administration of quinelorane produced dose-related decreases in serum prolactin concentration of reserpinized, male rats and increases in serum corticosterone concentration of male rats. The minimum effective doses (MED) for these effects were 10 and 30 micrograms/kg i.p., respectively. Quinelorane induced increases in 3-methoxy-4-hydroxyphenylglycol-sulfate levels in the brain stem (MED, 30 micrograms/kg i.p.) and decreases in hypothalamic epinephrine levels (MED, 100 micrograms/kg i.p.) in male rats as determined by high-pressure liquid chromatography with electrochemical detection methods. Quinelorane induced increases in extracellular ascorbic acid as determined by in vivo voltammetry in the nucleus accumbens and striatum of male rats. Quinelorane produced concentration-dependent suppression of K+-evoked release of acetylcholine from superfused caudate slices, with an IC50 of approximately 10(-8)M. Quinelorane administration produced dose-related increases in compulsive, contralateral turning in male rats with unilateral nigrostriatal lesions and increases in locomotor activity and stereotypic behavior in male rats. In dogs, quinelorane administration produced dose-related increases in emetic response with an ED50 of 7 micrograms/kg i.v. Quinelorane administration also produced dose-related decreases in the striatal concentrations of the dopamine metabolites, 3,4-dihydroxyphenylacetic acid and homovanillic (MED, 1 microgram/kg i.p. for both metabolites) as determined by high-pressure liquid chromatography with electrochemical detection methods and decreases in extracellular concentrations of homovanillic acid in the nucleus accumbens and striatum as determined by in vivo voltammetry., Quinelorane produced concentration-dependent decreases in K+-evoked dopamine release from superfused striatal slices (IC50 = 3 X 10(-9) M).(ABSTRACT TRUNCATED AT 250 WORDS)

General pharmacology of nizatidine in animals.

Nizatidine (N-[2-[[[2-[(dimethylamino)methyl-4-thiazolyl]- methyl]thio]ethyl]-N'-methyl-2-nitro-1,1-ethenediamine, LY139037, Axid) is a novel, potent, and selective H2-antagonist. The potential secondary pharmacologic effects of this agent on the cardiovascular, respiratory, gastrointestinal, renal, hepatic, autonomic, and central nervous systems as well as effects on circulating blood glucose and the acute inflammatory response were examined. Nizatidine was generally inactive in the tests conducted in mice, rats, guinea pigs, rabbits, and dogs. Nizatidine and the reference H2-antagonist, cimetidine, both produced effects upon the cardiovascular and respiratory systems by intravenous administration in anesthetized dogs at doses in excess of the intended clinical exposure. In summary, these studies confirm the selective pharmacologic activity of nizatidine and indicate a low potential for secondary pharmacologic side effects to be encountered clinically.

Comparison of the 5-HT3 receptor antagonist properties of ICS 205-930, GR38032F and zacopride.

The well-documented 5-HT3 receptor antagonists, ICS 205-930 and GR38032F, have been compared with regard to their inhibitory activity at 5-HT3 receptors to another gastrokinetic agent, zacopride. Zacopride and ICS 205-930 showed similar affinity (-log kB approximately 8.0), whereas GR38032F showed lower affinity (-log ka approximately 7.0) at 5-HT3 receptors in the guinea pig ileum. After i.v. administration to anesthetized rats, zacopride was approximately 10-fold more potent than either ICS 205-930 or GR38032F, which were equipotent as inhibitors of serotonin-induced bradycardia (5-HT3-mediated activation of the von Bezold Jarisch reflex). After oral administration to anesthetized rats, zacopride remained approximately 10-fold more potent than ICS 205-903, which was approximately 2-fold more potent than GR38032F as an inhibitor of serotonin-induced bradycardia. Furthermore, the inhibitory effectiveness of GR38032F persisted for less than 3 hr after oral administration and for less than 15 min after intravenous administration. ICS 205-930 produced maximal inhibition of serotonin-induced bradycardia for over 3 hr with heart rate returning to control values 6 hr after oral administration. Zacopride possessed the longest duration of inhibitory effectiveness in urethane-anesthetized rats with maximal inhibition still apparent 6 hr after oral administration. All three agents inhibited cisplatin-induced emesis after i.v. administration in dogs with zacopride being 10-fold more potent than ICS 205-930 or GR38032F, which were equipotent. These comparative data with three 5-HT3 receptor antagonists indicate that in animals, zacopride was more potent and longer acting than either ICS 205-930 or GR38032F. Furthermore, after oral administration to rats, GR38032F was slightly less potent than ICS 205-930 and possessed the shortest duration of action.

Swallow-evoked peristalsis in opossum esophagus: role of cholinergic mechanisms.

This study examined the role of cholinergic mechanisms in esophageal smooth muscle of anesthetized opossums. Swallow-induced motility was recorded manometrically before and after treatment with physostigmine or metoclopramide. At the 5-cm site above lower esophageal sphincter (LES) the latency, duration, and amplitude of swallow-evoked contractions were 2.1 +/- 0.12 (SE) s, 1.8 +/- 0.31 s, and 74.5 +/- 6.1 mmHg, respectively. At the 1-cm site the respective values were 4.6 +/- 0.38 s, 3.8 +/- 0.24 s, and 42.0 +/- 4.6 mmHg. The calculated speed of peristalsis was 1.72 +/- 0.21 cm/s. After intravenous treatment with the physostigmine (100 micrograms X kg-1 X h-1), each swallow produced contractions that were simultaneous, repetitive, and of long duration. The values of duration and amplitude of contraction at the 5- and 1-cm site after physostigmine treatment were significantly higher than controls (P less than 0.01). There was a significant decrease in the latency of contraction at all sites. This resulted in a significant faster speed of peristalsis. Atropine (30 micrograms/kg iv) reversed the influence of physostigmine, resulting in a significantly slower speed of peristalsis. Metoclopramide (10 mg/kg iv) resulted in several changes in swallow-evoked contractions similar to those observed after physostigmine treatment. These data suggest that alteration in cholinergic mechanisms results in disordered esophageal peristalsis in the opossum.

Role of substance P nerves in longitudinal smooth muscle contractions of the esophagus.

Longitudinal muscle strips from different sites along the opossum esophagus were stimulated transmurally so as to produce neurally mediated contractions. Low-frequency transmural stimulation produced contractions after termination of the stimulus ("off" contractions), whereas high-frequency stimuli produced contractions beginning during the stimulus and extending beyond termination of the stimulus (extended-duration contractions). The intrastimulus portion of the extended-duration contraction was partially antagonized by atropine or substance P desensitization, whereas the poststimulus portion of the contraction was selectively and fully antagonized by desensitization with substance P. A combination of atropine and substance P desensitization abolished the extended-duration contraction. The amplitude of contraction was greater in the proximal than in the distal strips, irrespective of the mode of stimulation. The poststimulus portion of the extended-duration contraction was significantly longer in muscle strips taken from more distal than proximal portions of the esophagus. This gradient in duration of contractions was abolished by substance P desensitization but was not affected by atropine. Exogenously applied substance P (10 microM) produced equally sustained long-duration contractions at all sites along the esophagus. These observations suggest that a) both acetylcholine- and substance P-containing nerves are responsible for the extended-duration contraction of longitudinal muscle, and b) transmural stimulation causes an aborally directed increase in the duration of contractions; this gradient of increasing duration of contraction appears to be due to a more prolonged neural release of substance P at more distal sites.

Regional gradient of initial inhibition and refractoriness in esophageal smooth muscle.

The influence of two successive vagal stimuli on esophageal contractions was studied by recording intraluminal pressures in the smooth muscle portion of the opossum esophagus. The esophageal contraction in response to the first or second stimulus in a pair of vagal stimuli was inhibited depending on the interstimulus interval, the frequency of the stimulus, and the esophageal site. The esophageal contraction in response to the first vagal stimulus was inhibited by a second vagal stimulus if the latter stimulus was applied before the peak of the first contraction. This phenomenon is termed initial inhibition. Initial inhibition is a graded phenomenon. It was greater at higher frequencies than at lower frequencies (p less than 0.001), and was significantly greater in the distal esophagus than in the proximal esophagus (p less than 0.01). The term "refractoriness" has been used to denote inhibition of the second esophageal contraction by the first. Refractoriness was observed during and beyond the duration of the first esophageal response. Refractoriness was also observed at all esophageal levels; however, the interstimulus intervals that demonstrated refractoriness were significantly greater in the distal than in the proximal esophagus (p less than 0.01). Refractoriness was complete (effective refractory period) during the ascending phase of the first contraction. Refractoriness was incomplete after the peak of the contraction (relative refractory period). These studies show gradients of durations and degrees of initial inhibition and refractoriness along the esophagus. The gradient is responsible for the peristaltic nature of esophageal contraction. The gradients of initial inhibition and refractoriness determine esophageal response to multiple successive swallows.

Swallow-evoked action potentials in vagal preganglionic efferents.

Swallow-evoked potentials in the preganglionic vagal fibers were studied using the single-fiber recording technique in anesthetized opossums. Swallows were evoked by tactile pharyngeal stimulation or electrical stimulation of the cut central end of the superior laryngeal nerve (SLN). Swallowing activity was recorded by the mylohyoid electromyogram and esophageal motility. Sixty-six fibers were studied in which swallowing evoked action potentials. The latencies (from the onset of mylohyoid activity) of evoked responses in different fibers varied from 100 ms to 5 s. The discharge rate of the evoked response was 3-8 action potentials per burst. Each burst lasted 1.1 +/- 0.02 (SE)s. The latencies of evoked spike bursts showed a bimodal distribution. In 34 fibers the latencies were less than 1 s, and in 32 fibers the latencies ranged between 1 and 5 s; these are the short- and long-latency fibers, respectively. Short-latency fibers could easily be distinguished from long-latency fibers based on the influence of SLN-stimulus frequency. Short-latency discharges had low thresholds of activation and were sensitive to changes in the frequency of SLN stimulation, since their latencies decreased and their discharge rate increased with increasing SLN-stimulus frequency. On the other hand, the latencies and discharge rates of long-latency discharges were not modified with changing SLN stimulus frequencies. The conduction velocities of 6 short- and 9 long-latency fibers were 5.64 +/- 0.12 and 5.78 +/- 0.12 (SE) m/s, respectively (P greater than 0.05). The relationship between the latencies of swallow-evoked discharges in the short- and long-latency fibers and the esophageal smooth muscle responses suggested that the short-latency discharges may correlate with the latency of initial inhibition, and the long-latency fibers may correlate with latencies of peristaltic contractions. Based on these temporal relationships, we speculate that vagal efferent fibers showing swallow-evoked, short-latency discharges make contact with intramural inhibitory neurons. They may mediate deglutitive inhibition in the body of the esophagus, relaxation of the lower esophageal sphincter, and receptive relaxation of the fundus of the stomach. The fibers showing late discharges make contact with intramural excitatory neurons and participate in their sequential activation. This dual pathway of activation may be responsible for physiological esophageal peristalsis.

Intramural mechanism of esophageal peristalsis: roles of cholinergic and noncholinergic nerves.

We examined the role of peripheral cholinergic and noncholinergic mechanisms in esophageal peristalsis. Intramural nerve elements in rings of circular muscle from six different levels of the opossum esophagus were stimulated transmurally so as to cause neurally mediated muscle contractions. Stimulus frequency was varied from 2 to 40 Hz. An increase in stimulus frequency caused an increase in latencies of contractions in rings from distal esophageal sites and a decrease in latencies in rings from proximal sites. This resulted in a marked slowing of the calculated peristaltic speed. Increasing stimulus frequency also caused an increase in duration and amplitude of contractions. These effects were reversed by atropine (0.1 microM), suggesting that higher stimulus frequencies recruited more cholinergic nerves. In the presence of atropine, increasing the stimulus frequency caused an increase in latencies of contraction at all sites, suggesting that increasing stimulation frequency applied to noncholinergic nerves causes an increase in latencies of contraction at all sites. The results of this study indicate that both noncholinergic and cholinergic nerves play a role in the peripheral mechanism of esophageal peristalsis. Cholinergic nerve stimulation reduces the latency and enhances the amplitude and duration of contractions seen with noncholinergic nerve stimulation alone. The influence of cholinergic innervation is most prominent proximally and decreases distally along the smooth muscle portion of the esophagus. This peripherally located gradient of cholinergic innervation plays an important role in determining the speed and amplitude of esophageal peristalsis.

Acetylcholinesterase and choline acetyltransferase staining of neurons in the opossum esophagus.

The purpose of the present investigation was to identify and compare cholinergic intramural neurons in the lower esophageal sphincter and esophageal body by histochemical staining for acetylcholinesterase and the enzyme that synthesizes acetylcholine, choline acetyltransferase. Opossums were anesthetized and their abdominal cavity was opened by a midline incision to expose the esophagogastric junction. The lower esophageal sphincter was identified manometerically and localized in situ with markers. Tissues were removed, rapidly frozen in freon cooled with liquid nitrogen and serial cryostat sections were obtained from the lower esophageal sphincter and esophageal body. Sections were stained with one of the above histochemical procedures and adjacent sections were stained with Solachrome cyanin , which differentially stains nerve elements from muscle fibers. The muscle of the lower esophageal sphincter and esophageal body was stained with nonspecific cholinesterase with some selectivity of intensity of reaction in the various smooth muscle layers. All identifiable plexus neurons in the esophagus stained for nonspecific cholinesterase and acetylcholinesterase. Nerve fiber tracts were also stained for acetylcholinesterase within the longitudinal and circular layers of the tunica muscularis. Reaction for choline acetyltransferase showed no staining in the muscle layers or nerve fiber tracts of either part of the esophagus studied; however, selected neurons within the myenteric plexus of both regions (approximately 38%) were reactive. There was no significant difference in the number of positive choline acetyltransferase neurons in the lower esophageal sphincter or esophageal body.

Characteristics of "on" and "off" contractions in esophageal circular muscle in vitro.

The prevalence, amplitude, and latency periods of "on," "off," and "intermediate" contractions in response to transmural stimulation were recorded in transverse rings of circular muscle from different levels of the opossum esophagus. Ten-second train stimuli consistently produced off contractions. On contractions were not seen at lower stimulus frequencies (2 and 5 Hz); however, their incidence approached 90% at higher frequencies (40 Hz). Intermediate contractions occurred only at stimulus frequencies of 10 Hz or greater and were less frequent than on contractions. In general, the on and intermediate contractions had significantly lower amplitudes than the off contraction. The on contraction occurred with a latency period from initiation of the stimulus. This latency was greater in the more distal sites and decreased with increasing stimulus frequency. The off contractions occurred with a latency period from termination of the stimulus. This latency was not dependent on either stimulus frequency or site along the esophagus. Atropine antagonized the on and intermediate contractions but had no such effect on the off contraction. Tetrodotoxin abolished the on, off, and intermediate contractions. This study suggests that an intramural mechanism exists that upon stimulation produces atropine-sensitive on contractions. These contractions may play a role in esophageal peristalsis.

Membrane potential and mechanical responses of the opossum esophagus to vagal stimulation and swallowing.

Studies were performed in anesthetized opossums. The electrical changes, recorded using a suction electrode applied to the outside of the esophagus, and mechanical activity, recorded by an intraluminal catheter, were monitored from 5 cm above the lower esophageal sphincter. Swallowing was associated with membrane hyperpolarization followed by depolarization and spike burst. Electrical stimulation of the decentralized vagus also caused a prompt hyperpolarization followed by an overshoot depolarization. Single pulses of stimulation caused primarily hyperpolarization. The amplitude and duration of hyperpolarization increased with increasing frequencies of vagal stimulation. Spike burst occurred as the membrane potential was recovering from the peak hyperpolarization and moving toward peak depolarization. The latency of onset of spike burst decreased with increasing frequency of vagal stimulation. The muscle contraction occurred after a latency. The latency of contractions, like the latency of spike burst, decreased with increased frequency of vagal stimulation. These studies show that (a) membrane hyperpolarization is present during the latent period of contraction associated with swallowing, suggesting that swallow-induced esophageal response may be mediated by vagal inhibitory pathway to the esophagus and (b) spike bursts can be temporally dissociated from depolarization by changing the vagal stimulation frequency, suggesting that spike burst and depolarization may be mediated by different excitatory mechanisms.

Influence of successive vagal stimulations on contractions in esophageal smooth muscle of opossum.

Studies were performed in anesthetized opossums to investigate the influence of successive vagal stimuli on esophageal contractions. Mechanical activity was recorded manometrically 5 cm above the lower esophageal sphincter. Contractions in the esophagus were evoked by electrical stimuli of 2.5 mA, and 1-ms pulse duration applied to the cervical vagi, at various train durations and frequencies. Paired or multiple stimuli of 1-s train length were also tested at different interstimulus intervals (ISI). Paired stimuli at an ISI of less than or equal to 3 s and at a frequency of less than or equal to 10 Hz showed refractoriness, i.e., the contractions to the first stimulus inhibited the contraction to the second stimulus. A frequency of 50 Hz showed initial inhibition, i.e., the second stimulus inhibited the contraction to the first stimulus. Repetitive stimuli applied at a rate of 8/min (ISI 6.5 s) evoked contractions to each stimulus. At 15/min, every second or third contraction was inhibited. With stimuli applied at 30/min, contractions occurred only in response to the first and/or the last stimulus; depending upon the frequency of vagal stimulation. The intervening stimuli did not evoke any contractions. A long train stimulus produced an initial, a terminal, or both contractions depending on the stimulation value. These studies show that (a) vagal efferent stimulation causes initial inhibition and refractoriness in the esophageal smooth muscle; (b) the degree of initial inhibition increases with increasing frequency of stimulation; (c) the occurrence of contractions only at the onset and the end of a long train stimulus may be due to the influence of initial inhibition and refractoriness.

Modulation of esophageal peristalsis by vagal efferent stimulation in opossum.

Experiments were performed on anesthetized opossums to study the influence of vagal efferent stimulation on peristalsis in the esophageal smooth muscle using various stimulus parameters. Current intensity, pulse duration, frequency, and train duration were varied systematically. Electrical and mechanical activities were recorded simultaneously at 5, 3, and 1 cm above the lower esophageal sphincter (LES). Vagal efferent stimulation produced a spike burst and contraction with a latency after the termination of the stimulus. This latency varied at different sites with the same stimulus parameters. For example, a stimulus of 5 mA, 0.5 ms, 10 Hz, and 1-s train produced latencies for the electrical response of 1.48 +/- 0.04, 2.2 +/- 0.12, and 3.5 +/- 0.09 s (+/- SEM) at 5, 3, and 1 cm above LES, respectively. The differences in latency were statistically significant (P less than 0.01). The latency of response at any one site also changed with different stimulus parameters; e.g. at 1 cm above LES, the latency of electrical response at 10 Hz was 3.5 +/- 0.09 s, but at 20 Hz the latency was 2.01 +/- 0.06 s when current intensity, pulse, and train duration remained at 5 mA, 0.5 ms, and 1 s. This decrease in latency with increasing frequency was statistically significant (P less than 0.01). By changing stimulus parameters, antiperistalsis or peristalsis with different speeds of propagation could be induced. Antiperistalsis or simultaneous responses occurred near threshold stimulus parameters. Suprathreshold stimuli produced peristaltic responses. Speed of peristalsis in the distal esophagus was 1.82 +/- 0.08 cm/s with swallowing, which was not different from 1.98 +/- 0.14 cm/s (P greater than 0.05) with vagal stimulation of 5 mA, 0.5 ms, 10 Hz, and 1-s train. These studies suggest that: (a) peristalsis in the smooth muscle part of the esophagus can be explained entirely on the basis of peripheral mechanisms, and (b) the central nervous system may modulate the occurrence, polarity, and speed of propagation by modifying the intensity and frequency of vagal activation.