Peptide immunoreactivities in the ganglionated plexuses and nerve fibers innervating the human gallbladder.

The mammalian gallbladder is innervated by a well-developed intrinsic neural network. However, little is known about the neurochemistry and organization of the innervation of this organ in humans. The aim of this study was to analyze the distribution of immunoreactivity (IR) for the neuropeptides, vasoactive intestinal polypeptide (VIP), neuropeptide Y (NPY), tachykinins (TK) and calcitonin gene-related peptide (CGRP) in the human gallbladder by means of immunohistochemistry. Neuropeptide-IRs are found in neurons and processes of the two ganglionated plexuses, i.e., the innermost plexus located in the lamina propria at the base of the mucosal folds, and the outermost plexus situated within the fibro-muscular layer. In these two plexuses, VIP-, NPY- and TK-IRs are present in ganglion cells and varicose fibers, whereas CGRP-IR is confined to nerve processes. VIP-IR is present in most, if not all, neurons. NPY- and TK-IRs are also found in many neurons. The densities of the peptide-IR nerves in the mucosa are NPY and VIP > TK >> CGRP, and in the fibro-muscular layer are NPY > VIP and TK > CGRP. The vasculature is richly innervated by NPY-IR nerves, which are mostly perivascular. CGRP-, VIP- and TK-IR processes are found only occasionally around blood vessels and in a paravascular position. Double-label studies demonstrated that a large number of VIP-containing neurons expresses NPY- or TK-IR. On the other hand, all neurons positive for either NPY- or TK-IR are immunostained for VIP. In agreement with these findings, most of the NPY-IR fibers in the lamina propria and fibro-muscular layer contain VIP-IR, and numerous TK-IR fibers are positive for VIP. However, the perivascular NPY-IR processes do not contain VIP-IR, suggesting an extrinsic origin. In addition, a population of TK-IR processes contains CGRP-IR and presumably originates from extrinsic sources, since CGRP/TK-IR intrinsic neurons could not be detected in the gallbladder. Peptide-IRs have a similar distribution in the neck, body and fundus of the gallbladder. No peptide-containing endocrine/paracrine cells are observed in the epithelium. The presence of peptide-IRs in the ganglionated plexuses and the abundance of peptidergic innervation suggest that peptides exert their effects on gallbladder function by acting directly on tissue targets and influencing intrinsic ganglion cells. Furthermore, the co-localization of more than one peptide in the same neuron raises the possibility that peptides are co-released upon stimulation and might interact at the same target.

Nitric oxide producing neurons in the monkey and human digestive system.

Nitric oxide has been proposed as an inhibitory transmitter molecule that plays a role in muscle relaxation and vasodilation in the gastrointestinal tract. The present study analyzes the distribution of nitric-oxide-producing neurons in the monkey and human digestive system by means of nicotinamide-adenine-dinucleotide-phosphate-diaphorase histochemistry. This histochemical method is reliable and convenient for the visualization of neuronal nitric-oxide synthase, the enzyme responsible for nitric-oxide generation. In the gastrointestinal tract, nitric-oxide-synthase-related diaphorase activity was present in nerve fibers running throughout the muscular layer (circular > longitudinal) and in numerous ganglion cells and processes in the myenteric plexus of monkeys and humans. Labelled ganglion cells and fibers also were observed in the submucous plexus, although they were much less numerous than those seen in the myenteric plexus. In the submucosa, a few positive fibers were seen around blood vessels. In the mucosa, stained fibers were sparse at the base of the villi and crypts, whereas they were quite abundant in the muscularis mucosae, especially in the small intestine and colon. In the gallbladder (human), labelling was found in ganglion cells and processes of the innermost and outermost ganglionated plexuses. Stained fibers also were distributed to the muscular layer and, less abundantly, to the mucosa and vasculature. Labelled fibers were more abundant in the sphincter of Oddi (human) than in the gallbladder. In the monkey and human pancreas, nicotinamide-adenine-dinucleotide-diaphorase staining was seen mainly in ganglion cells and fibers of intrapancreatic ganglia, and in processes running among acini, around ducts and in the stroma. A moderate density of stained fibers also was distributed to the vasculature, whereas the islets showed few positive processes. Finally, double label experiments performed in the pancreas showed that the vast majority of neurons producing nitric oxide are immunoreactive for vasoactive intestinal peptide.(ABSTRACT TRUNCATED AT 250 WORDS)

Preservation and propagation of cyclic myoelectric activity after feeding in rat small intestine.

The cyclic nature and distal propagation of the fasting migrating motor complex (MMC) of the small intestine have been well described. The fate of the MMC after feeding, however, has not been elucidated. We used time series analysis with fast Fourier transforms (FFT) to study myoelectric cycling before and after feeding. Ten rats were chronically prepared with bipolar electrodes secured to the duodenum and proximal jejunum. Spikeburst frequency was recorded before and after feeding simple nutrients. During fasting, cyclic activity occurred at the MMC frequency. Although after feeding this periodicity appeared disrupted, FFT analysis showed persistent cycling at the fasted (MMC) rate. Digital filtering of data at the MMC frequency isolated cycling from background noise during both fasted and fed states and showed a depression in the amplitude of the waveform after feeding. Root-mean-square analysis of the waveform confirmed statistically significant depression of amplitude by 47-57%. The waveform propagated from the duodenum to the jejunum during both the fasted and fed state at an unchanged rate. We conclude that factors controlling myoelectric cycling during the fasted state persist after feeding, allowing continued net abroad propulsion of food.

Late functional adaptation after colectomy, mucosal proctectomy, and ileal pouch-anal anastomosis.

Ileal pouch-anal anastomosis (IPAA) is currently an alternative to proctocolectomy and ileostomy for patients with ulcerative colitis or familial polyposis. Some studies have suggested significant anal sphincter damage after mucosal proctectomy. Our aim was to assess prospectively late sphincter function after IPAA. In 250 patients, anorectal pressures were assessed with a pneumohydraulic perfused catheter manometry system. Each patient underwent colectomy, mucosal proctectomy, ileoanal anastomosis of a 15 cm ileal J-pouch, and loop ileostomy. Eight weeks after IPAA, anal manometry was repeated, and the ileostomy was closed. Manometry was repeated at yearly intervals. A decline in resting tone of the anal sphincter occurred early after IPAA with a gradual recovery toward control. External sphincter squeeze after pressures were not affected by IPAA and steadily increased to 8 years after operation. During this time, a progressive increase in J-pouch capacity was noted, and 24-hour stool frequency declined from 7.9 +/- 0.3 stools to 6.5 +/- 0.3 stools (p less than 0.05). We conclude that mucosal proctectomy results in internal anal sphincter trauma but is associated with long-term sphincter recovery, coupled with a significant improvement in external sphincter capacity, ileal pouch volume, and stool frequency.

Substance P stimulates the opossum sphincter of Oddi in vitro.

We have previously shown that substance P (SP) regulates sphincter of Oddi (SO) motility in vivo. However, its mechanism of action remains unclear. Our aim was to develop an in vitro model to measure spikeburst (SB) an contractile frequency (CMC) of the SO and to characterize further SP effects. In 16 opossums, SO rings were excised, mounted within a Kreb's tissue bath with bipolar electrodes and force transducers, allowed to equilibrate, and exposed to increasing SP concentrations with washout between each test solution. Spikeburst and CMC frequencies were recorded on a polygraph, quantitated, expressed as differences before and during SP, and statistically analyzed with Student's test. Although SP induced a significant concentration-dependent increase in phasic SB frequency and CMC, the amplitude of concentrations was not affected by SP. A close correlation was found between basal and SP-stimulated SB and CMC, suggesting myoelectric and mechanical coupling. Previous exposure of SO to SP antagonist [D-Arg1, D-Pro2, D-Trp7,9, Leu11]-SP significantly decreased the response to SP. Tetrodotoxin (TTX), did not affect the delta CMC response to SP. In conclusion an in vitro preparation was developed to study the effect of SP on the SO. Substance P increased SB and CMC of the SO in a concentration-dependent fashion, thus acting as a stimulatory peptide. Perfusion of SO rings with SP antagonist had no effect on basal CMC but significantly inhibited the action of SP in a competitive manner. The effect of SP was not altered by TTX. These data suggest that the action of SP on the SO is primarily myogenic.

Characterization of substance P effects on sphincter of Oddi myoelectric activity.

We examined the effects of substance P (SP) on the myoelectric activity of the opossum sphincter of Oddi (SO). Myoelectric data from the SO in five adult opossums were recorded using thin stainless steel electrodes and computer-assisted analog-to-digital conversion. In fully awake and conscious animals, baseline spikeburst activity during phase I of the MMC occurred at a frequency of 28.6 +/- 3.1 spikebursts (SB) per 20-min period. Intravenous infusion of graded doses of substance P (from 0.5 to 8.0 micrograms/kg) stimulated SO myoelectric activity in a dose-related manner (from 80 +/- 8 to 235 +/- 11 SB/20 min, respectively, P less than 0.05 when compared to baseline). The effect of substance P on SO myoelectric activity was antagonized by administration of the H2-blocker, cimetidine (92.0 +/- 6.1 vs 48.2 +/- 7.0, n = 5, P less than 0.05). Administration of the antimuscarinic drug atropine only slightly affected the SO spikeburst frequency when infused prior to SP (73.0 +/- 10.4 vs 70.8 +/- 8.2, P greater than 0.05). We conclude that SP stimulated the SO spikeburst frequency in a dose-dependent fashion. Cimetidine markedly inhibited the response of the SO to SP but atropine did not. The excitatory effect of substance P on the opossum SO is mediated at least in part by a histaminergic, noncholinergic pathway.

Graded effect of protein on regional myoelectric activity of the opossum sphincter of Oddi.

Although it is known that the sphincter of Oddi exhibits a myoelectric response to intraluminal nutrients, the effect of specific dietary components has not been well characterized. The aim of this study was to determine the effect of intragastric instillation of a protein meal on regional myoelectric activity of the opossum sphincter of Oddi. Seven adult opossums were chronically prepared with self-retaining bipolar electrodes secured to the proximal and distal parts of the sphincter. After a 2-week recovery period, and with the animals fasted and conscious, myoelectric activity was recorded for two cycles of the migrating myoelectric complex. The animals then received graded calories of protein through an orogastric tube and recordings were continued until return of the fasted cyclic activity. Doses of 60, 120, and 240 kcal of protein elicited increases in myoelectric spike activity of the proximal sphincter of 18.7%, 41.9%, and 66.6%, respectively. Similarly, the length of the fed state was prolonged by increasing caloric loads, with mean (plus or minus standard error of the mean) times of 266 +/- 25, 292 +/- 15, and 375 +/- 42 minutes for each dose. A similar relationship was noted in the distal part of the sphincter. Regression analysis between the proximal and distal parts of the sphincter during the fasted and fed states showed a strong and persistent correlation of gradient of spikeburst frequency. We conclude that myoelectric spike frequency of the opossum sphincter of Oddi exhibits a dose-related response to intragastric protein calories. Furthermore, the proximal and distal parts of the sphincter remain electrically coupled, with the proximal region pacing the distal region during the fasted state and after protein administration.

Postprandial sphincter of Oddi myoelectric activity and gallbladder emptying.

Feeding initiates gallbladder emptying and bile delivery into the duodenum. It is not yet defined how the sphincter of Oddi regulates flow of bile into the duodenum during gallbladder emptying. The aim of this study was to assess postprandial spike burst activity in the sphincter of Oddi, while quantitating gallbladder emptying with noninvasive radioisotope imaging. Six adult opossums were prepared with bipolar electrodes in the sphincter of Oddi. After 2 weeks of recovery the animals were fasted overnight and positioned under a gamma camera, and myoelectric recordings were begun. After two cycles of the migrating motor complex (MMC), 2 mCi 99Tc-HIDA was infused intravenously and permitted to concentrate in the gallbladder for a period of 30 min. Subsequently, a 30-ml liquid meal, containing 0.9 g protein, 3.5 g carbohydrate, and 3.3 g fat, was instilled into the stomach. Sphincter of Oddi myoelectric activity (spike bursts/min) and gallbladder emptying (expressed as percentage of original 99Tc counts in the gallbladder) were measured at intervals for 120 min following feeding. Feeding resulted in prompt gallbladder emptying. Sphincter of Oddi spike burst activity was not altered significantly in the first 30 min after feeding, suggesting that motor activity in the sphincter of Oddi does not initially influence bile flow. Subsequently, spike burst activity increased progressively, suggesting that sphincter of Oddi motor activity may accelerate bile delivery into the duodenum during later phases of gallbladder emptying.