Effect of cholecystokinin-octapeptide on opossum lower esophageal sphincter

We evaluated the action of cholecystokini-octapeptide (CCK-OP) on lower esophageal sphincter (LES) pressure in the opossum. LES pressure was recorded by an infused sleeve device that straddled the sphincter, whereas intraluminal esophageal pressure and gastric pressure were recorded via conventional manometric catheters. Progressive intravenous pulse doses of CCK-OP caused 1) graded increases in LES pressure, 2) circular and longitudinal smooth muscle contraction in esophageal body, and 3) mild increases in intragastric pressure. Pressor effect of CCK-OP on the LES was weakly antagonized by tetrodotoxin (TTX), but not by atropine, phentolamine, or pyrilamine. TTX antagonism of CCK-OP appeared to be nonspecific because TTX also partially antagonized LES contractions induced by pentagastrin, substance P, and bethanechol. We conclude that CCK-OP at doses that cause LES relaxation in other species induces LES contraction in the opossum. This pressor effect appears to be elicited by a direct action of the hormone on LES smooth muscle.     

Modulation of lower esophageal sphincter relaxation in the opossum.

The relationship of lower esophageal sphincter (LES) relaxation and sphincter pressure was studied in the basal state and after neurohumoral stimulation in the anesthetized opossum. LES relaxation was induced by electrical vagal stimulation, balloon distension, and swallowing, whereas LES pressure was increased by infusions of gastrin I, histamine, phenylephrine, or bethanechol. Each agent was selected to give an approximately twofold increase in LES pressure, at the highest dosage. The lower dosage gave a submaximal increase in LES pressure. Neither gastrin nor histamine modified sphincter relaxation as induced by vagal stimulation, esophageal distension, or swallowing. Phenylephrine or bethanechol made the LES less sensitive to vagal stimulation or esophageal distension without reducing maximal LES relaxation. Only bethanechol significantly decreased sphincter relaxation in response to swallowing (P less than 0.05). The results of this study suggest that LES relaxation is not a fixed response simply dependent on the inhibitory stimulus or the resting tonic level of LES pressure, but may be modulated by both alpha-adrenergic and cholinergic drugs.     

Interaction of gastrin I, secretin, and cholecystokinin on gallbladder smooth muscle.

The effect of cholecystokinin (CCK), the octapeptide of cholecystokinin (CCK-OP), gastrin I, and secretin was studied on guinea pig gallbladder smooth muscle in vitro. Both CCK and CCK-OP stimulated gallbladder contraction, with CCK-OP being more potent. Gastrin I, over a wide dose range, had no effect on gallbladder contractility. Secretin alone also showed no effect on gallbladder smooth muscle but in combination with CCK-OP it produced a noncompetitive type of inhibition. Michaelis-Menten kinetics showed the calculated maximum response of the secretin plus CCK-OP interaction to be less than with CCK-OP alone. There was no change in the dose required to achieve half-maximal response, D50. These studies indicate that: a) CCK-OP has a greater effect on gallbladder contractility than CCK, b) gastrin I has no effect on gallbladder muscle tone, and c) secretin acts as a noncompetitive antagonist of CCK-OP. These findings suggest that gallbladder motor function may be determined in part by the interaction of secretin and CCK rather than solely in response to CCK.     

Inhibition of lower esophageal sphincter circular muscle by female sex hormones

To determine the effect of estrogenic and progesteronic activity on lower esophageal sphincter (LES) circular muscle, studies were performed on 20 adult opossums. Does-response curves on circular smooth muscle strips from the LES were constructed for gastrin and acetylcholine alone, and with 17beta-estradiol and/or progesterone added. Each female hormone significantly decreased the maximal LES muscle responses to gastrin and acetylcholine. A combination of 17beta-estradiol and progesterone abolished the response to gastrin. In contrast, the male sex hormone, dihydrotestosterone, had no effect. In conclusion, administration of estrogen and progesteron, but not dihydrotestosterone, in vitro reduced LES muscle responses to gastrin and acetylcholine. These studies suggest that the female sex hormones can alter LES function and potentially may be of importance in the pathogenesis of heartburn of pregnancy.     

Generation of nitric oxide in the opossum lower esophageal sphincter during physiological experimentation

Lipopolysaccharide (LPS), given in vivo, modulates opossum esophageal motor functions by inducing the inducible nitric oxide synthase (iNOS), which increases nitric oxide (NO) production. Superoxide, a NO scavenger, is generated during this endotoxemia. Superoxide is cleared by superoxide dismutase (SOD) and catalase (CAT) to protect the physiological function of NO. This study examined whether lower esophageal sphincter (LES) motility, NO release, and iNOS and nitrotyrosine accumulation in the LES are affected by LPS in vitro. Muscle strips from the opossum LES were placed in tissue baths containing oxygenated Krebs buffer. NO release was measured with a chemiluminescence NOx analyzer, and Western blots were performed to analyze iNOS and nitrotyrosine production. The percent change in resting LES tone after a 6-hour exposure to LPS was significantly increased compared to pretreatment values. The percent LES relaxation upon electrical stimulation was significantly decreased in the control group at 6 hours, indicating that the LPS treatment had an effect. The NO concentration in the tissue bath of LPS- treated muscle without nerve stimulation was significantly less than that of LPS treatment combined with SOD/CAT or SOD/CAT alone. iNOS and nitrotyrosine were detectable and increased over time in the LES muscle of both the control and LPS-treated groups. Antioxidant enzymes may play a role in regulating NO-mediated neuromuscular functions in the LES.     

LES pressure response to pentagastrin: effect of cholinergic augmentation and inhibition

Studies were performed on the anesthetized adult opossum to examine a) the pressure response of the lower esophageal sphincter (LES) to both bolus injections and continuous infusions of pentagastrin (PG), and b) how this response is affected by either cholinergic antagonism with atropine or cholinergic enhancement with edrophonium, an anticholinesterase. Both the bolus injection and the continuous infusion of PG produced dose-dependent rises in LES pressure. The peak pressure responses occurred at 1 microgram/kg and 32 microgram/kg-h, respectively. Atropine (100 microgram/kg), in a dose that significantly diminished the LES response to exogenous acetylcholine, had no significant effect on PG-induced increases in LES pressure. Similarly, edrophonium (100 microgram/kg) had no effect on the response of the LES to PG stimulation. The Sphincter's response to exogenous acetylcholine was significantly enhanced after edrophonium pretreatment. From these studied, we conclude that PG-induced increases in the LES pressure are due primarily to the direct stimulation of sphincter smooth muscle rather than to activation of excitatory cholinergic neurons.     

Effect of atropine on esophageal motor function in humans

In this study, we used a high-fidelity manometric recording system to quantitate the effects of atropine on lower esophageal sphincter (LES) pressure and primary peristalsis (1 degree P). A sleeve sensor recorded LES pressure, and seven recording orifices spaced at 3-cm intervals registered motor activity in the esophageal body. Five randomized manometric studies were done in each of five normal subjects. LES pressure and 1 degree P with wet swallows were recorded for 30 min before and 70 min after intravenous injection of saline or atropine, 3, 6, 12, and 24 micrograms/kg. We also studied the effect of atropine on LES pressure in five additional subjects, four dogs, four opossums, and six monkeys. In humans, saline and 3 micrograms/kg atropine caused no significant change in pulse rate, LES pressure, or the incidence of complete peristaltic sequences. The 6, 12, and 24 micrograms/kg atropine doses caused significant inhibition of LES pressure and the incidence of intact 1 degree P. Only the 12 and 24 micrograms/kg doses increased pulse rate. When 1 degree P occurred in the smooth muscle portion of the esophagus its appearance in the proximal portion of the smooth muscle segment was delayed for several seconds. The amplitude of 1 degree P was decreased 30-60% in the smooth muscle segment, but 1 degree P was not affected in the proximal striated muscle esophageal segment. Atropine reduced canine LES pressure substantially but caused no change in opossums or monkeys. We conclude that 1) basal LES tone in humans and dogs, unlike that of the opossum and monkey, is partially generated by cholinergic neural input, 2) cholinergic nerves elicit 1 degree P in human esophageal smooth muscle, and 3) species variation exists in esophageal responses to atropine.     

Histaminergic pharmacology of primate lower esophageal sphincter.

We have studied the lower esophageal sphincter (LES) response to exogenous histamine and to H1- and H2-blocking agents in the awake baboon. Increasing intravenous bolus doses of histamine produce an increase in LES pressure with a maximum response at a dose of 12 microgram/kg. H1-receptor blockade with chlorpheniramine over a wide dose range did not alter basal LES pressure but did abolish the response of the LES to exogenous histamine. H2-receptor blockade with cimetidine at doses markedly inhibiting gastric acid secretion (2 mg/kg.h) did not alter basal LES pressure or the response of the LES to exogenous histamine. In addition, cimetidine did not alter the response of the LES to pentagastrin and bethanechol. Although histamine and histamine receptors are important in gastric secretion, they appear to have no identifiable role in the maintenance of basal LES smooth muscle tone in the baboon. These results demonstrate the presence of a stimulatory H1 receptor on baboon LES smooth muscle, but provide no evidence for the presence of an H2-inhibitory receptor. As opposed to the parietal cell, the LES response to pentagastrin and bethanechol does not require a H2 receptor.     

Incretin physiology beyond glucagon-like peptide 1 and glucose-dependent insulinotropic polypeptide: cholecystokinin and gastrin peptides

Gastrin and cholecystokinin (CCK) are homologous hormone systems known to regulate gastric acid secretion, gallbladder emptying, and cell growth in the pancreas and stomach. They are, however, also involved in the development and secretory functions of pancreatic islet cells. For instance, foetal and neonatal islets express significant amounts of gastrin, and human as well as porcine islet cells express the gastrin/CCK-B receptor abundantly. Therefore, exogenous gastrin and CCK peptides stimulate insulin and glucagon secretion in man. Accordingly, endogenous hypergastrinaemia is accompanied by islet cell hyperplasia and increased insulin secretion. Conventionally, the effect of gastrointestinal hormones on insulin secretion (the incretin effect) has been defined and quantified in relation to oral versus intravenous glucose loadings. Under these unphysiological conditions, the release of gastrin and CCK and, hence, their effect on insulin secretion are modest in comparison with the effects of glucose-dependent insulinotropic polypeptide and glucagon-like peptide 1 (GLP-1). Consequently, the interest of CCK and gastrin in incretin research has for decades been limited. A few years ago, however, it was suggested that gastrin together with epidermal growth factor or later GLP-1 might stimulate beta cell growth and secretion. Recent studies have shown that the combination of gastrin and GLP-1 actually restores normoglycaemia in diabetic mice. Therefore, a short review of the incretin system in a broader functional context that includes gastrin and CCK peptides may be timely.     

Interaction between gastrin, CCK, and secretin on canine antral smooth muscle in vitro.

This study compared the actions and interactions of human synthetic gastrin, octapeptide-cholecystokinin (OP-CCK), cholecystokinin (CCK), and secretin on the amplitude of isometric tension developed in strips of dog antral smooth muscle in vitro. Cholecystokinin, OP-CCK, and gastrin produced maximal stimulatory effects at 7.5 x 10(-9), 4.5 x 10(-9), and 3.5 x 10(-9) M, respectively. Secretin alone was ineffective up to 2.5 x 10(-8) M. Observed maximal responses to gastrin, OP-CCK, and CCK tested alone were not significantly different. A submaximal gastrin dose added with OP-CCK, shifted the OP-CCK dose-response curve to the left and significantly reduced the D50, but the calculated maximal response (CMR) did not change. Also, submaximal OP-CCK plus gastrin shifted the gastrin dose-response curve to the left and significantly lowered the D50 with no change in CMR. Secretin decreased CMR but did not change the D50 for gastrin. Responses obtained to gastrin and OP-CCK tested alone were not affected by tetrodotoxin (1 x 10(-5) M), hexamethonium bromide (4 x 10(-5) M), or atropine (1 x 10(-7) M). Larger doses of atropine (5 x 10(-6) M) reduced peptide responses an average 30%. The results indicate that OP-CCK, CCK, and gastrin share a common noncholinergic receptor site. Secretin acts at a different receptor site.     

Prostaglandins and tetrodotoxin-insensitive relaxation of opossum lower esophageal sphincter.

Field stimulation with pulses of 0.5 or 5 ms relaxed isolated strips of lower esophageal sphincter (LES) of the opossum; only responses to 0.5-ms pulses were inhibited by tetrodotoxin. Black widow spider venom prevented relaxation to both stimuli; thus both stimuli may release nonadrenergic inhibitory mediator. Isoproterenol, but not PGEs or ATP, was a consistent relaxant of LES. PGF2alpha (approximately 1 microgram/ml) and stable endoperoxides (approximately 10 ng/ml) stimulated LES muscle. Doses of indomethacin (IDM) or 5,8,11,14-eicosatetraynoic acid (ETA), which inhibited contractions to arachidonic acid increased then abolished LES tone, inhibited relaxations to 5-ms pulses and less effectively to 0.5-ms pulses. Inhibition of relaxation preceded loss of tone. Tone could be restored by carbachol, PGEs, or PGF2alpha and relaxation after IDM but not ETA was also restored. Prostaglandins may participate in functioning of nonadrenergic inhibitory nerves and in maintaining sphincter tone. Cells that did not appear to be smooth muscle were in gap junction contact with smooth muscles and closely apposed to nerves with small agranular vesicles. A role for these structures, which are postulated to be interstitial cells, in tetrodotoxin-insensitive prostaglandin-related release of nonadrenergic inhibitory mediator was proposed.     

The half-life of exogenous gastrin in the circulation

1. A method of gastrin bio-assay is described which can be used on as little as 30 ng synthetic human gastrin I at a minimum concentration of 2.5 ng/ml.2. Pentagastrin or synthetic human gastrin I added to cat plasma can be stored on ice or at 4 degrees C, for periods up to 27 hr without apparent loss of gastrin activity.3. Between 1(1/2) and 13 min after the rapid I.V. injection of pentagastrin in the anaesthetized cat and between 1(1/2) and 15 min after the injection of synthetic human gastrin I, there is a rapid reduction of the gastrin concentration in the arterial plasma. The data relating log(10) gastrin concentration in arterial plasma with time can be fitted by a single term.4. Studies in vitro show that over the periods of time involved in the in vivo studies, both pentagastrin and synthetic human gastrin I are stable in cat plasma at 37 degrees C in concentrations which occurred in the circulating plasma.5. The half-life of pentagastrin in the circulating arterial plasma of the anaesthetized cat is 1.50 min (S.E. +/- 0.08) and the half-life of synthetic human gastrin I is 2.65 min (S.E. +/- 0.09).     

Age-related changes in gallbladder contractility and gallbladder cholecystokinin receptor population in the guinea pig

We have examined the effects of aging on guinea pig biliary motility both in vitro and in vivo. The first experiment compared contractile tension of gallbladder strips from young adult (6-12 months old) and 3-year-old guinea pigs in vitro. Contraction of gallbladder strips from the young guinea pigs was twice as forceful and was more sensitive to octapeptide of cholecystokinin (CCK-8) stimulation than the gallbladder strips from the older guinea pigs. The two groups were also studied in vivo by measuring changes in the intraluminal pressure of the gallbladder in response to exogenously administered doses of CCK-8. Young adult guinea pigs were more sensitive to CCK-8 at the lower doses tested and demonstrated gallbladder contractions that were more forceful than that of the old guinea pigs. CCK receptors were measured on gallbladder muscularis membranes from young adult and old guinea pigs. The number of receptors on gallbladder membranes decreased with age: 65.0 +/- 17.7 fmoles/mg protein on membranes from 1 year old; 7.9 +/- 2.0 fmoles/mg protein on membranes from 3 years old. The binding affinity of CCK receptors on gallbladder muscularis membranes for binding to CCK-8 was not significantly different in the two age groups studied. We conclude that age-related decreases in gallbladder responses to CCK-8 may be due to decreased concentrations of CCK receptors on gallbladder muscle cells.     

Multiple cholecystokinin (CCK) receptors and CCK-monoamine interactions are instrumental in the control of feeding

Almost two decades ago, exogenous cholecystokinin (CCK) was shown to suppress food consumption in rats. Since then, CCK has been detected not only in peripheral tissue but extensively throughout the central nervous system. Furthermore, specific CCK receptors have been described, and a distinction drawn between CCK-A and CCK-B receptors. Recently, potent, orally active CCK antagonists, which show a high degree of selectivity for either CCK-A or CCK-B receptors, have been introduced. The present report reviews recent evidence obtained in studies using devazepide (a selective CCK-A receptor antagonist) and L-365,260 (a selective CCK-B/gastrin receptor antagonist). Both compounds increased food consumption and postponed the onset of satiety in well-satiated rats. L-365,260 was more potent, suggesting that central CCK-B type receptors may mediate the satiety effects of endogenously released CCK. Only devazepide was effective in blocking the feeding-suppressant effect of exogenous CCK, indicating that CCK-A type receptors mediate this effect. In a second series of studies, devazepide but not L-365,260 antagonized the anorectic effect of either d-fenfluramine or systemically administered 5-HT. Hence, CCK-A type receptors appear to be involved in the anorectic effects of these serotonergic drugs. We propose that CCK and 5-HT mechanisms involved in mediating satiety are mutually interdependent. Possible interactions between CCK and catecholaminergic mechanisms are also briefly considered.     

Effect of bombesin and related peptides on the release and action of intestinal hormones on pancreatic secretion.

1. Pancreatic volume flow as well as bicarbonate and protein secretion from pancreatic fistulas have been measured in response to i.v. infusion of graded doses of bombesin and related peptides containing the COOH-terminal fragment of the bombesin molecule in conscious dogs with intact antrum and in anaesthetized animals with antrectomy, or antrectomy and enterectomy. 2. Bombesin and related peptides given to conscious dogs produced a potent and dose-dependent increase in pancreatic protein output reaching a maximum equal to that induced by the octapeptide of cholecystokinin (OP-CCK) as well as a small rise in bicarbonate output attaining a peak amounting to about 10% of that evoked by secretin. The serum gastrin level rose progressively during the infusion of bombesin to reach a peak with the highest dose of peptide. 3. Bombesin infused i.v. in anaesthetized animals with resected antrum also evoked a marked increase in pancreatic protein secretion without significant changes in the serum gastrin level. Following the removal of the antrum and small intestine, bombesin failed to show any stimulation of the pancreatic secretion or any change in the serum gastrin level. It is concluded that the strong stimulatory action of bombesin and related peptides on pancreatic secretion cannot be entirely ascribed to the release of gastrin but might be attributed at least in part to the release of intestinal hormones, particularly CCK. 4. Atropine and the growth hormone-release inhibiting hormone (GH-RIH), which were shown to inhibit the release of CCK induced by duodenal perfusion of an amino acid mixture, also caused the inhibition of pancreatic protein secretion by bombesin but failed to affect the pancreatic response to OP-CCK. The results indicate that bombesin releases, in addition to gastrin, CCK from the gut by a mechanism largely dependent upon cholingeric innervation.     

Vagally mediated release of gastrin and cholecystokinin following sensory stimulation.

The aim of the present study was to investigate whether gastrin, cholecystokinin (CCK) and somatostatin secretion can be influenced by sensory stimulation and if so, whether such effects are mediated via the vagal nerves. Male rats anaesthetized with chloral hydrate were exposed to three different stimuli, i.e. to low frequency (2 Hz) electrical stimulation of muscles via needles (electro-acupuncture), to thermal stimulation at 40 degrees C or to vibration at 100 Hz. The two former stimuli activate mainly small and medium sized myelinated fibres from muscles and skin respectively, whereas vibration activates large myelinated fibres from skin, subcutaneous tissue and muscles. Experiments were also performed on animals that were vagotomized or exposed to prior treatment with atropine (0.5 mg kg-1). Blood was collected at various time intervals and plasma levels of gastrin, CCK and somatostatin were measured with radioimmunoassay (RIA). All three stimuli, i.e. electro-acupuncture, vibration and thermal stimulation caused significant elevations of gastrin (103 +/- 11-151 +/- 16 pM, 105 +/- 8-140 +/- 12 pM and 105 +/- 14-162 +/- 4 pM) and cholecystokinin (9 +/- 0.8-15 +/- 2.8 pM, 8 +/- 0.5-10 +/- 1.5 pM and 8.0 +/- 0.5-10.5 +/- 1.5). Somatostatin was raised in response to electro-acupuncture (10 +/- 1-14 +/- 3 pM). Vagotomy and atropinization abolished the release of gastrin and CCK in response to all three stimuli. CCK levels were significantly reduced following electro-acupuncture in atropinized rats. In conclusion, gastrin and cholecystokinin release is stimulated by activation of sensory afferent, originating in skin, subcutaneous tissue as well as in muscle.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effect of cadmium and bombesin on exocrine pancreatic secretions and plasma levels of gastrin and cholecystokinin (CCK) in rats

The effects of cadmium and bombesin on exocrine pancreatic secretions and plasma levels of gastrin and cholecystokinin (CCK) were studied in anesthetized rats with pancreatic and gastric fistulas. Rats treated only with saline were used as controls. Both control and cadmium (0.1 mg per kg) treated rats were infused with saline, secretin, and bombesin (BBS). Blood and pancreatic juice samples were collected at regular time intervals. Plasma levels of gastrin and CCK were measured in blood samples by specific radioimmunoassay. Pancreatic juice samples were measured for volume, protein, and trypsin outputs. Compared to saline treated rats, outputs of volume, protein, and trypsin were significantly greater in cadmium treated rats. Plasma levels of gastrin were suppressed with secretin but significantly elevated with BBS. Plasma CCK levels were not different from basal after secretin or BBS in rats treated with either cadmium or saline. The results suggest that the administration of cadmium stimulated exocrine pancreatic secretion by a mechanism that does not involve gastrin or CCK. Bombesin may have a direct influence on the stimulation of exocrine pancreatic secretion in rats.     

Inhibition of gastric acid secretion inthe Atlantic cod, Gadus morhua, by sulphated and desulphated gastrin, caerulein, and CCK-octapeptide

Gastric acid secretory effects of gastrin/CCK-like peptides have been assayed in cods rendered "spontaneously" secreting by a continuous intestinal perfusion with diluted (33%) seawater. A high dose of pentagastrin induced a weak stimulation (31%) of acid output, while gastrin 17-II, caerulein and CCK8 were inhibitory. Caerulein was the most potent peptide, with an estimated D50 for inhibition of 0.013 nmol/kg.h. Although displaying lower potencies, also the desulphated forms of gastrin-17, caerulein and CCK8 were inhibitory. The results may be explained by release of an endogenous inhibitor, or by interaction with endogenous "codfish gastrin". In the latter case two alternatives are considered: Either gastrin 17, CCK8, and caerulein possess lower efficacies than "codfish gastrin" and therefore act as partial agonists. Alternatively, "codfish gastrin" is itself an inhibitory principle (gastron), the effect of which is mimicked by gastrin 17, caerulein and CCK8. The actions of gastrin and the gastrin-like peptides in the cod indicate a structure-activity relationship different from previously described systems, both mammalian and submammalian.     

Interaction of cholecystokinin with vasoactive intestinal peptide in body shaking response to ice-water immersion in rats

Interaction of cholecystokinin (CCK) with vasoactive intestinal peptide (VIP) in body shaking response to ice-water immersion was observed in pentobarbital-anesthetized rats. Although CCK itself had no influence on the response, VIP suppressed it and this effect of VIP was antagonized by simultaneous administration of sulfated octapeptide of CCK, but not by non-sulfated CCK.