Circular Muscle Cells Research Articles

EFFECT OF SECONDARY METABOLITES FROM THREE ENTOMOPATHOGENIC FUNGI ON THE COTTON LEAF WORM, SPODOPTERA LITTORALIS (BOISD.)

Three entomopathogenic fungi isolates, i. e., Beauveria bassiana, Paecilomyces fumosoroseus and Metarhizium anisopliae were used as biological control agents against the Egyptain cotton leaf worm, Spodoptera littoralis (Boisd.), which represent the most severe destructive cotton pest in Egypt and many other countries. The entomopathogenic fungi were inoculated YES medium, incubation period was carried out for 19 days at pH 5.4 and 28 – 30oC. After incubation period, the mats of the fungi were removed and YES broth was mixed individually with chloroform/methanol (2:1, v/v). The mixture was shaken vigorously in a separating funnel and left to settle down forming a dense lower aqueous layer containing the secondary metabolites. Extracted metabolites were then concentrated by using a speed vacuum device (Maxi Dry Plus) to a volume of 1 ml. Virulence of B. bassiana, M. anisopliae and P. fumosoroseus was investigated against 1st instar larvae of the cotton leaf worm S. littoralis. Results obtained revealed that the three metabolites (toxins) were toxic to the 1st instar larvae of S. littoralis. The most effective toxin was M. anisopliae (84.92 %) followed by P. fumosoroseus (66.32%) and B. bassiana (62.65 %). On the other hand, LT50 values in days for M. anisopliae were 8.5414, 6.4808, 4.7721, 3.7211, while for P. fumosoroseus they were 11.0488, 7.7757, 6.4998, 5.1703 and for B. bassiana they were 10.4380, 8.3415, 6.8299, 5.6922 in relation to the concentrations 25, 50, 75, 100% respectively. Treatment of S. littoralis 4th instars with LC50 of the metabolic fungal toxins caused complete disruption of endocuticle of the resulting late 6th instars. Meanwhile, it caused exfoliation of the mid gut epithelium from the underlying circular muscle fibers, leaving a large vacuole or space, disruption of both the peritrophic membrane and columnar cells, beside high vacuolization and collapsing of the fat body.

Analysis of pacemaker activity in the human stomach

Extracellular electrical recording and studies using animal models have helped establish important concepts of human gastric physiology. Accepted standards include electrical quiescence in the fundus, 3 cycles per minute (cpm) pacemaker activity in corpus and antrum, and a proximal-to-distal slow wave frequency gradient. We investigated slow wave pacemaker activity, contractions and distribution of interstitial cells of Cajal (ICC) in human gastric muscles. Muscles were obtained from patients undergoing gastric resection for cancer, and the anatomical locations of each specimen were mapped by the operating surgeon to 16 standardized regions of the stomach. Electrical slow waves were recorded with intracellular microelectrodes and contractions were recorded by isometric force techniques. Slow waves were routinely recorded from gastric fundus muscles. These events had similar waveforms as slow waves in more distal regions and were coupled to phasic contractions. Gastric slow wave frequency was significantly greater than 3 cpm in all regions of the stomach. Antral slow wave frequency often exceeded the highest frequency of pacemaker activity in the corpus. Chronotropic mechanisms such as muscarinic and prostaglandin receptor binding, stretch, extracelluar Ca(2+) and temperature were unable to explain the observed slow wave frequency that exceeded accepted normal levels. Muscles from all regions through the thickness of the muscularis demonstrated intrinsic pacemaker activity, and this corresponded with the widespread distribution in ICC we mapped throughout the tunica muscularis. Our findings suggest that extracellular electrical recording has underestimated human slow wave frequency and mechanisms of human gastric function may differ from standard laboratory animal models.

Myoanatomy and serotonergic nervous system of plumatellid and fredericellid phylactolaemata (lophotrochozoa, ectoprocta)

The phylogenetic position of the Ectoprocta within the Lophotrochozoa is discussed controversially. For gaining more insight into ectoproct relationships and comparing it with other potentially related phyla, we analysed the myoanatomy and serotonergic nervous system of adult representatives of the Phylactolaemata (Plumatella emarginata, Plumatellavaihiriae, Plumatella fungosa, Fredericella sultana). The bodywall contains a mesh of circular and longitudinal muscles. On its distal end, the orifice possesses a prominent sphincter and continues into the vestibular wall, which has longitudinal and circular musculature. The tentacle sheath carries mostly longitudinal muscle fibres in Plumatella sp., whereas F. sultana also possesses regular circular muscle fibres. Three groups of muscles are associated with the lophophore: 1) Lophophoral arm muscles (missing in Fredericella), 2) epistome musculature and 3) tentacle musculature. The epistome flap is encompassed by smooth muscle fibres. A few fibres extend medially over the ganglion to its proximal floor. Abfrontal tentacle muscles have diagonally arranged muscle fibres in their proximal region, whereas the distal region is formed by a stack of muscles that resemble an inverted 'V'. Frontal tentacle muscles show more variation and either possess one or two bases. The digestive tract possesses circular musculature which is striated except at the intestine where it is composed of smooth muscle fibres. The serotonergic nervous system is concentrated in the cerebral ganglion. From the latter a serotonergic nerve extends to each tentacle base. In Plumatella the inner row of tentacles at the lophophoral concavity lacks serotonergic nerves. Bodywall musculature is a common feature in many lophotrochozoan phyla, but among other filter feeders like the Ectoprocta is only present in the 'lophophorate' Phoronida. The longitudinal tentacle musculature is reminiscent of the condition found in phoronids and brachiopods, but differs to entoproct tentacles. Although this study shows some support for the 'Lophophorata', more comparative analyses of possibly related phyla are required.

Ventricular Torsion and Untwisting: Further Insights into Mechanics and Timing Interdependence: A Viewpoint

Ventricular torsion and untwisting are essential for normal ventricular function and their mechanisms are related to the temporal responses of the helical and circular muscle fibers that comprise cardiac architecture. Explanation of the presystolic isovolumic contraction (IVC) period is essential for analysis of these interactions. Structural and imaging studies by magnetic resonance, speckle tracking, velocity vector encoding, and sonomicrometer crystals are described to define why and how different muscular components contract asynchronously. Mechanical and functional relationships are described for pre-systolic IVC, torsion, postejection isovolumic interval, and rapid and slow filling. Circular fibers dominate to cause pre- and posttwisting global counterclockwise and clockwise movement, and helical fibers govern torsion whereby the base rotates clockwise and apex counterclockwise; untwisting cannot begin until torsion is completed. Prolonged torsion extends into the postejection isovolumic interval and delays untwisting, and is caused by prolonged contraction of the right-handed helical arm or descending segment of the helical ventricular myocardial band that narrows the ∼80 ms "timing hiatus" between end of shortening of the descending and the ascending segment or left-handed arm of the helical muscle. Longer torsion duration by this mechanism becomes the common theme for unbalanced torsion and untwisting in diastolic dysfunction, physiological, structural, and electrical disease processes, whose management may be guided by changing the interconnected reasons for these adverse mechanical and timing factors.

Identification of GIRK2-4 subunits in human esophageal smooth muscle cells

Acetylcholine (ACh) secreted from the vagus nerve contributes to the physiological and pathological regulation of the contraction and relaxation of human esophageal smooth muscle. Expression of acetylcholine-sensitive G protein‑activated inwardly rectifying potassium channels (GIRKs) occurs widely in the heart, nervous system and gastrointestine, but the role of GIRKs in the esophagus remains unclear. In the present study, expression of the GIRK1-4 subunits in mRNA and total protein was examined in human esophageal smooth muscle cells (SMCs) by reverse transcription polymerase chain reaction (RT-PCR) and Western blotting. mRNA and protein expression of the GIRK2-4 subunits was detected in human esophageal longitudinal muscle (LM) and circular muscle (CM) cells. However, GIRK1 mRNA and protein were not observed in either the esophageal LM or CM. This study is the first to identify the expression of GIRK2-4 subunits in human esophageal SMCs.

Role of TLR4 and MAPK in the local effect of LPS on intestinal contractility

Lipopolysaccharide (LPS) has been shown to alter intestinal contractility. Toll-like receptor 4 (TLR4), K(+) channels and mitogen-activated protein kinases (MAPKs) have been proposed to be involved in the mechanism of action of LPS. The aim of this study was to determine the role of TLR4, K(+) channels and MAPKs (p38, JNK and MEK1/2) in the local effect of LPS on the acetylcholine (ACh)-induced contractions in rabbit small intestine in vitro. Segments of rabbit duodenum were suspended in the direction of longitudinal or circular smooth muscle fibres in a thermostatically controlled organ bath. LPS (0.3 µg/ml) reduced the contractions induced by ACh (100 µm) in the longitudinal and circular smooth muscle of the duodenum after 90 min of incubation. Polymyxin (TLR4 inhibitor), SB203580 (p38 MAPK inhibitor), SP600125 (JNK1/2 inhibitor) and U0126 (MEK1/2 inhibitor) antagonized the effects of the LPS on ACh-induced contractions in duodenal smooth muscle. Incubation with the blockers of K(+) channels, TEA, apamin, charybdotoxin, iberiotoxin, glibenclamide or quinine, did not reverse the effect of LPS on ACh-induced contractions. These results suggest that the effect of LPS on ACh-induced contractions in the rabbit duodenum might be mediated by TLR4 and p38, JNK1/2 and MEK1/2 MAPKs.

New idea of intestinal lengthening and tailoring

Both the Bianchi and the serial transverse enteroplasty (STEP) procedure have been reported to be successful in short bowel syndrome, but both have their limitations. The Bianchi technique is surgically challenging and can only tailor the diameter to half and double the length. The STEP procedure is easy, adjustable, but it changes the orientation of the muscle fibres making the circular; longitudinal and the longitudinal; circular. We have created a model to test our idea of Spiral Intestinal Lengthening and Tailoring (SILT). A double layer intestinal simulator was used as the bowel model. The orientation of the circular muscle fibres was marked. The simulator was cut spirally at a 45° and 60° angle, stretched longitudinally and retubularised. The procedure was adapted to porcine small bowel ex vivo and mucosal microcirculation was observed in three separate points by intravital videomicroscopy in vivo. The simulator was lengthened by 60% and the diameter was tailored by 33% using the 45° spiral cut, 73% lengthening and 44% tailoring was achieved at 60°. The circular muscle fibres showed oblique orientation. The porcine bowel was lengthened by 136 ± 21% and the diameter was tailored by 56 ± 8%. The linear regression analysis of variants showed significant linear regression R = 0.9689 (R(2) = 0.9388), p = 0.0014. The angle of the spiral cut (α) showed the relation: α ≥ 90-arc sin R(2)/R(1). (R(1) is the radius of the original segment and R(2) the desired radius of the tailored and lengthened bowel). The bowel remained viable macroscopically 90 min after the procedure. The median red blood cell velocity reduced from 570 (control) to 558 μm s(-1) in point 1, to 382 μm s(-1) in point 2, and to 482 μm s(-1) in point 3. Oscillation of the capillary flow has not been observed. The SILT may be an easy, more physiological and adjustable alternative to the existing techniques.

Sp707 How to Successfully Manage Novel Technical Challenges in Per Oral Endoscopic Myotomy for Achalasia

Achalasia is a primary esophageal motility disorder and surgical disruption of the lower esophageal sphincter is usually performed, when conservative approaches failed to be sufficient. Since the first report of laparoscopic Heller myotomy in 1991, minimally invasive techniques have been increasingly used for definitive treatment of achalasia. Efforts in natural orifice translumenal surgery (NOTES) led to the development of novel methods such as submucosal endoscopic myotomy in an animal model. Recently, the clinical implementation of per oral endoscopic myotomy (POEM) was demonstrated. The clinical implementation of such novel procedures might conceal a novel variety of challenges, which have not been dealt with before. We describe the successful management of unexpected difficulties as well as lessons learned in an early clinical series of per oral endoscopic myotomies for the treatment of achalasia. We studied the initial five patients, enrolled in an institutional review board approved trial, who had per oral endoscopic myotomy for symptomatic achalasia. A conventional flexible endoscope with a transparent cap mounted onto the distal end was used. After mucosal incision and entry into the submucosal space, a tunnel within the esophageal wall was created using a blunt dissection or hook needle-knife cautery. The inner circular muscle fibers at the lower esophageal sphincter were then divided at a length of approximately 7 cm. The mucosal incision was then closed using hemostatic clips. All interventions were videotaped and analyzed on unexpected technical difficulties as well as their management were analyzed. POEM could be successfully performed in all patients (5/5) and resulted in immediate smooth passage of the endoscope through the gastro-esophageal sphincter. Although no clinical complications were observed, novel intra-operative technical challenges were observed. The transparent cap used on the endoscope loosened three times during retrograde movements and was subsequently lost within the submucosal tunnel. Several attempts to remove the submucosal foreign body with graspers or an endoscopic net failed as the cap twisted and was caught underneath the mucosa. With the aid of a balloon catheter the lost cap was finally lined-up with the endoscope and could be safely removed without injuring the esophageal mucosa. POEM appears to be an appealing therapeutic approach for achalasia and the knowledge of basic rescue maneuvers in case of technical challenges might be essential to shorten the learning curve of endoscopists and ensure patient's safety.

Histomorphology of the Stomach, Proventriculus and Ventriculus of the Red Jungle Fowl

The cranial chamber (proventriculus) and caudal chamber (ventriculus) of the stomach of the Red jungle fowl (Gallus gallus spadiceus) were examined by means of light microscopy. Both chambers presented folds of the tunica mucosa lined by a simple prismatic epithelium that was positive for neutral mucin. Simple tubular glands occupied the lamina propria of both chambers; in the ventriculus of older birds, they showed a coiled base. These ventricular glands were lined by simple cuboidal cells represented by the chief cells and a few large basal cells. The luminal and tubular koilin rodlets and folds of the ventriculus were positive to periodic acid Schiff (PAS) stain. The proventricular glands were situated between the inner and outer layers of the lamina muscularis mucosae. Cells lining the tubulo-alveolar units of the proventricular glands showed a dentate appearance. Vacuoles were not observed, and the cells were negative for Alcian-PAS stain. The tunica submucosa was very thin in the proventricular wall. In the ventriculus, it was not separated from the lamina propria owing to the absence of any lamina muscularis mucosae. The tunica muscularis of the proventriculus was formed by a thick inner layer of circular smooth muscle fibres and a thin outer layer of longitudinal fibres. In addition to these layers, oblique muscle fibres formed the most internal layer of the tunica muscularis in the ventriculus.

Fine structure of Longicollum pagrosomi (Acanthocephala: Pomphorhynchidae) and intestinal histopathology of the red sea bream, Pagrus major, infected with acanthocephalans

The results described the structure of Longicollum pagrosomi and histopathological characters of the intestine of the red sea bream, Pagrus major, infected with acanthocephalans, using the light and electron microscopes. Among the six samples of P. major, L. pagrosomi was identified in the posterior intestine of five fish samples. Adult L. pagrosomi (total length, 8-27 mm) is divided into the presoma (proboscis, anterior neck, and posterior neck) and metasoma (trunk). The proboscis had vertically arranged hooks (40 μm in length), with ten hooks per row, and the septum was observed between the posterior neck and trunk. The tegument thickness of the proboscis was approximately 15 μm, and it was composed of thin, circular muscle fibers. The outer fibrous membrane was approximately 1 μm, and the connective tissue layer was approximately 35 μm in thickness in the anterior neck. The tegument of the posterior neck enclosed the cephalic ganglion and had longitudinal and vertical muscle fibers, and the tegument thickness was approximately 45 μm. The tegument of the body, which was approximately 1 mm in thickness, was composed primarily of muscle and collagen fibers, and the structure of the tegument was different, depending on the body region. The acanthocephalans had ovaries and oval-shaped eggs with an eggshell (77.5 × 17.1 μm), floating within the body cavity of the trunk. In the infected posterior intestine of P. major, the presoma and the anterior part of the metasoma of L. pagrosomi passed through the intestinal wall and infected the intestinal tissue, perforating the loose connective tissue. In the inflammatory connective tissue, collagen and muscle fibers were fragmented and revealed partial necrosis. Lipid drops and eosinophilic granular cells aggregated in the connective tissue of the tissue capsule. In the vicinity of the acanthocephalan, the mucosal epithelia contained hypertrophied nuclei, and the epithelial layer was collapsed. In an extreme case, the mucosal fold was degenerated because of pressure from the acanthocephalan.

Embryonic development of the internal anal sphincter in rats with anorectal malformations

Background/Purpose The embryogenesis of the internal anal sphincter (IAS) in anorectal malformations (ARMs) remains unclear. This study aimed to investigate the development of the smooth muscle in the terminus of the digestive tract in normal and abnormal rats. Methods Rat embryos with ARMs were generated by administration of ethylenethiourea to pregnant rats. The normal rat embryos and embryos with ARMs from E13.5 to E21 were serially sectioned in the sagittal plane and stained immunohistochemically using specific antibody to α–smooth muscle actin (SMA). Temporospatial study was carried out on circular muscle of the distal portion of the hindgut. Results α-Smooth muscle actin immunolabeling cells could not be observed in the hindgut on E13.5, E14, and E14.5. On E15, there were α-SMA immunolabeling circular muscle cells in the hindgut; and the distal portion of the circular muscle was not thickened in the normal and ARMs rats. From E16 onward, the smooth muscle with slight dilated terminus, which was characterized by the features of IAS, could be noted in the primitive anorectum. In the normal group, the circular muscle in the distal portion of the hindgut thickened slightly and became the musculature with shutter-like bundles. In the ARMs group, the α-SMA immunolabeling myogenic precursors of the smooth muscle could be observed in the primitive anorectum as well. The musculature was similar to that in the normal group. On E15 and E16, there was no significant difference in the development of the circular muscle in the 2 groups. Moreover, the terminus of the circular muscle in the hindgut did not reach the orificium fistulae in ARMs rats. From E17 onward, in ARMs rats, the funnel-shaped distal hindgut communicated the genitourinary tract with a narrow fistula; the dilated musculature at this portion thinned gradually and formed an acute angled extremity in the ARMs group rather than formed blunt extremity in the normal group; the terminus circular muscle in the dorsal hindgut reached the orificium fistulae. During the following gestational days, the circular muscle of the hindgut in both normal and ARMs rats continued its own tendency. Conclusion The IAS primordium started to appear at the terminus of the hindgut on E15 in the 2 groups. The IAS in the ARMs group failed to develop as well as that in the normal group. The IAS dysplasia occurred in the late embryonic development (E17-E21).

A new family of the Hoplonemertea: Cinclidonemertes mooreae fam., gen. et sp. nov. (Nemertea: Hoplonemertea)

Recent morphological, cladistic and molecular phylogenetic studies suggest a strong relationship between Cratenemertidae and Reptantia. In contrast to other hoplonemerteans, these groups have complex cerebral organs located behind the pre-cerebral septum possessing both sensory and “sac-type” epithelial components in the canal structure and a robust single-layered rhynchocoel wall composed of interwoven circular and longitudinal muscle fibres. Four specimens collected by the US Antarctic Program, listed in an earlier cladistic study as “Antarctic C”, distinct from other Hoplonemertea and related to Cratenemertidae and Reptantia, are described herein as Cinclidonemertes mooreae fam., gen. et sp. nov. They have complex cerebral organs, with the same two types of canal epithelium, lying against the sides of the brain and extending below and behind it ventrally, a lattice-like rhynchocoel wall with partial interweaving, and four large shallow cup-shaped eyes with little pigment as opposed to the numerous heavily pigmented deep-cup ocelli of Cratenemertidae and Reptantia.

Intestinal effects of lipopolysaccharide in rabbit are mediated by cyclooxygenase-2 through p38 mitogen activated protein kinase

The mediators of the pathophysiological symptoms of septic shock are not completely understood. The intracellular signalling mechanisms of lipopolysaccharide (LPS)-induced effects need further investigation. This study investigates (1) the role of COX-2 in the effect of LPS on (a) the KCl, acetylcholine and prostaglandin E 2-induced contractions of rabbit duodenum and (b) the oxidative stress status in plasma and intestine and (2) the relationship between p38 MAPK and COX-2 expression in rabbit duodenum. Rabbits were injected i.v. with either (1) saline, (2) LPS, (3) SB203580, a p38 MAPK inhibitor, (4) SB203580 + LPS, (5) NS-398, a COX-2 inhibitor or (6) NS-398 + LPS. Contractility studies were performed by suspending pieces of duodenum in an organ bath in the direction of longitudinal and circular smooth muscle fibres. The formation of products of oxidative damage to proteins (carbonyls) and lipids [malondialdehyde (MDA) and 4-hydroxyalkenals (4-HDA)] was quantified in intestinal tissue and plasma. The protein expression of COX-2 was measured by western blot. The KCl, acetylcholine and prostaglandin E 2-induced contractions decreased with LPS. In addition, LPS increased the levels of carbonyls and MDA + 4-HDA in plasma and duodenum as well as COX-2 expression in duodenal tissue. All these effects were blocked by NS-398. The p38 MAPK inhibitor SB203580 blocked the effect of LPS on COX-2 expression. These results suggest that the effect of LPS on KCl, acetylcholine and prostaglandin E 2-induced contractions in the rabbit duodenum and oxidative stress might be mediated by an increase in COX-2 expression through the p38 MAPK pathway.

Morphological Maturation Level of the Esophagus Is Associated With the Number of Circumesophageal Muscle Fibers During Archenteron Formation in the Starfish Patiria (Asterina) pectinifera

In echinoderms, the circumesophageal muscle is mesodermal in origin. Several studies of sea urchins have reported that the molecular events of myogenesis occur during the differentiation of the circumesophageal muscle in early embryogenesis. In contrast, few detailed reports have examined the differentiation of the circumesophageal muscle in larval starfish. Here, we examined the temporal-numeric distribution and differentiation of esophagus circular muscle fibers in the starfish Patiria pectinifera by using rhodamine-phalloidin staining. Muscle fibers were not detected in mouth-forming larvae, but a mean of about 10 muscle fibers was observed in 48-h larvae, and about 26 bundles were observed after 60 h. During the next 12 h, the number of muscle fiber bundles increased slightly to about 31 bundles and was stable until 96 h.

Gastrointestinal Tract Disorder in Natriuretic Peptide Receptor B Gene Mutant Mice

Natriuretic peptide receptor B (NPR-B), which has high affinity for C-type natriuretic peptide (CNP) and synthesizes intracellular cGMP, may be involved in gastrointestinal tract (GIT) regulation. A mutant allele of the NPR-B-encoding gene (Npr2) is responsible for the phenotype of the short-limb dwarfism (SLW) mouse. Homozygosity for this autosomal-recessive gene (slw/slw) leads to dwarfism and death before weaning because of milk retention in the stomach and intestinal distention. To elucidate the relationship between CNP/NPR-B signaling and GIT function, we investigated the association between Npr2 mutation and the GIT phenotype in slw/slw mice. The pylorus and large intestine of the mutants did not respond to CNP stimulation; further, they showed pyloric lumen narrowing with randomly aligned circular muscle cells. Comparison of the cGMP and neuronal marker distribution in GIT tissues confirmed cGMP expression in neuronal tissues. An Auerbach's plexus and submucosal tissues of the mutants didn't express cGMP and expressed Ca(2+). In contrast, those of normal mice (controls) expressed both cGMP and Ca(2+). Sequencing revealed that the causative Npr2 mutation was a 7-base deletion in exon 8, resulting in a frameshift and premature termination codon appearance. Therefore, the GIT phenotype of slw/slw mice is because of a CNP/NPR-B-signaling defect caused by an Npr2 mutation. These results facilitate better understanding of the role of CNP/NPR-B signaling in GIT motility.

Left–right asymmetric morphogenesis of the anterior midgut depends on the activation of a non-muscle myosin II in Drosophila

Many animals exhibit stereotypical left–right (LR) asymmetry in their internal organs. The mechanisms of LR axis formation required for the subsequent LR asymmetric development are well understood, especially in some vertebrates. However, the molecular mechanisms underlying LR asymmetric morphogenesis, particularly how mechanical force is integrated into the LR asymmetric morphogenesis of organs, are poorly understood. Here, we identified zipper ( zip), encoding a Drosophila non-muscle myosin II (myosin II) heavy chain, as a gene required for LR asymmetric development of the embryonic anterior midgut (AMG). Myosin II is known to directly generate mechanical force in various types of cells during morphogenesis and cell migration. We found that myosin II was involved in two events in the LR asymmetric development of the AMG. First, it introduced an LR bias to the directional position of circular visceral muscle (CVMU) cells, which externally cover the midgut epithelium. Second, it was required for the LR-biased rotation of the AMG. Our results suggest that myosin II in CVMU cells plays a crucial role in generating the force leading to LR asymmetric morphogenesis. Taken together with previous studies in vertebrates, the involvement of myosin II in LR asymmetric morphogenesis might be conserved evolutionarily.

T1686 Gastric Clasp and Lower Esophageal Circular Muscle Fibers From GERD Patients With Barrett's Esophagitis Have a Decreased Contractile Response to Cholinergic Stimulation

Background: We recently showed that the gastric sling/clasp muscle complex does not contribute to the high pressure zone in GERD patients indicating a role in pathophysiology. Aim: We determined the cholinergic contractile response in clasp, sling and lower esophageal circular (LEC) fibers. Methods: Stomach and esophagi were obtained from 17 human transplant donors: 11 with no GERD history, 4 with probable GERD (proton pump inhibitor use) and 2 with definite GERD (Barrett's esophagitis). The contractile response to increasing carbachol concentrations was determined. Muscarinic receptor density was measured by subtype selective immunoprecipitation of [3]H-QNB binding. Results: Clasp and LEC fibers from definite GERD have decreased maximal contractile response. Contractility is increased in sling fibers of both definite and probable GERD. Concentrations of carbachol higher than 100 μM induce relaxations that are decreased in clasp fibers of probable and definite GERD. Relaxations are greater in sling fibers from definite GERD and higher in LEC from probable and definite GERD. Total and M2 receptor density is statistically lower in the GERD than non-GERD specimens in both clasp and sling fibers and M3 density is statistically lower in GERD than non-GERD clasp fibers. Conclusion: This suggest that a myogenic defect in clasp fibers may be involved in GERD pathophysiology in the organ donors with Barrett's esophagitis. The greater contractile response in the sling fibers from GERD donors suggest a possible compensatory mechanism for the lack of contractility of the clasp muscle fibers.

The ontogeny of muscle structure and locomotory function in the long-finned squidDoryteuthis pealeii

Understanding the extent to which changes in muscle form and function underlie ontogenetic changes in locomotory behaviors and performance is important in understanding the evolution of musculoskeletal systems and also the ecology of different life stages. We explored ontogenetic changes in the structure, myosin heavy chain (MHC) expression and contractile properties of the circular muscles that provide power for jet locomotion in the long-finned squid Doryteuthis pealeii. The circular muscle fibers of newly hatched paralarvae had different sizes, shapes, thick filament lengths, thin:thick filament ratio, myofilament organization and sarcoplasmic reticulum (SR) distribution than those of adults. Viewed in cross section, most circular muscle cells were roughly triangular or ovoid in shape with a core of mitochondria; however, numerous muscle cells with crescent or other unusual cross-sectional shapes and muscle cells with unequal distributions of mitochondria were present in the paralarvae. The frequency of these muscle cells relative to 'normal' circular muscle cells ranged from 1:6 to 1:10 among the 19 paralarvae we surveyed. The thick filaments of the two types of circular fibers, superficial mitochondria-rich (SMR) and central mitochondria-poor (CMP), differed slightly in length among paralarvae with thick filament lengths of 0.83+/-0.15 microm and 0.71+/-0.1 microm for the SMR and CMP fibers, respectively (P 0.05; ANOVA). During ontogeny the thick filament lengths of both the CMP and SMR fibers increased significantly to 1.78+/-0.27 microm and 3.12+/-0.56 microm, respectively, in adults (P<0.0001 for both comparisons; ANOVA with Tukey's highly significant difference post hoc tests). When sectioned parallel to their long axes, the SMR and CMP fibers of both paralarvae and adults exhibited the myofilament arrangements typical of obliquely striated muscle cells but the angle of obliquity of the dense bodies was 22.8+/-2.4 deg. and 4.6+/-0.87 deg. for paralarvae and adults, respectively. There were also differences in the distribution of the anastomosing network of SR. In paralarvae, the outer and central zones of SR were well developed but the intramyoplasmic zone was greatly reduced in some cells or was scattered non-uniformly across the myoplasm. Whereas in adults the intramyoplasmic SR region was composed primarily of flattened tubules, it was composed primarily of rounded vesicles or tubules when present in the paralarvae. The ontogenetic differences in circular muscle structure were correlated with significant differences in their contractile properties. In brief tetanus at 20 degrees C, the mean unloaded shortening velocity of the paralarval circular muscle preparations was 9.1 L(0) s(-1) (where L(0) was the preparation length that generated the peak isometric stress), nearly twice that measured in other studies for the CMP fibers of adults. The mean peak isometric stress was 119+/-15 mN mm(-2) physiological cross section, nearly half that measured for the CMP fibers of adults. Reverse transcriptase-polymerase chain reaction analysis of paralarval and adult mantle samples revealed very similar expression patterns of the two known isoforms of squid MHC. The ontogenetic differences in the structure and physiology of the circular muscles may result in more rapid mantle movements during locomotion. This prediction is consistent with jet pulse durations observed in other studies, with shorter jet pulses providing hydrodynamic advantages for paralarvae.

Purinergic neuromuscular transmission is selectively attenuated in ulcerated regions of inflamed guinea pig distal colon

This study was undertaken to investigate neuromuscular transmission in regions of the inflamed colon in which motility is disrupted. Propulsive motility was evaluated in segments of control guinea pigs and those treated 6 days previously with trinitrobenzene sulfonic acid. Intracellular recordings were then obtained from circular muscle cells to examine excitatory and inhibitory junction potentials (EJPs and IJPs). In inflamed preparations, propulsion of fecal pellets was temporarily halted or obstructed at sites of mucosal damage, whereas the propulsive motility was linear in control colons. The amplitudes of evoked and spontaneous IJPs were significantly reduced in ulcerated regions of inflamed preparations, but EJPs were comparable to controls. Pharmacological dissection of the IJP revealed that the purinergic component was reduced, while the nitrergic IJP was slightly increased. Furthermore, the reduction in the purinergic IJP in inflamed preparations persisted in the presence of hexamethonium, suggesting that the deficit involved the inhibitory motor neuron and/or smooth muscle. Nerve fibre density was not altered in the circular muscle, and pre-contracted rings of inflamed colon relaxed normally to ATP, suggesting that the deficit involves altered ATP release and/or degradation. The P2Y(1) receptor antagonist MRS2179 slowed propulsive motility indicating that decreased purinergic neuromuscular transmission could contribute to the inflammation-induced motor deficit. We conclude that purinergic inhibitory neuronal input to the circular muscle is selectively reduced in regions of the colon in experimental colitis where the mucosa is damaged, and this is likely to contribute to altered motility in colitis by diminishing downstream relaxation during the peristaltic reflex.

Anisakid Larvae in Atlantic Salmon (Salmo salar L.) Grilse and Post-Smolts: Molecular Identification and Histopathology

The molecular identification and histopathology are described for the parasitic larvae of a nematode species present in the abdominal cavity of Atlantic salmon ( Salmo salar ) grilse caught in fish traps on their natal river in the west of Ireland and post-smolts collected during experimental trawls on the continental shelf edge of the northeast Atlantic Ocean. Larvae in the adult and juvenile salmon were identified as Anisakis simplex sensu stricto by PCR amplification and RFLP and sequencing of the ITS gene and PCR amplification and sequencing of the cox2 gene. Parasitic nematode larvae in the grilse were either encapsulated in the abdominal mesentery associated with the pyloric ceca or on the serosal surface of the liver and in the vent region. In some fish, larvae were found in the parenchyma of the liver and muscularis circularis of the intestine. In general, the larvae induced a limited cellular response apart from the occurrence of focal melanin macrophage aggregates and individual eosinophilic granular cells in the connective tissue capsule. Melanin macrophage aggregates were also present among the hepatocytes adjacent to encapsulated larvae in the liver. The reaction to the parasites was more severe in the wall of the intestine. Encapsulated nematode larvae caused displacement, vacuolation, and necrosis of the circular muscle fibers. The stratum compactum was also disrupted with focal areas of degeneration. Overall, the intestinal wall had a hypercellular appearance with extensive cellular infiltration comprising eosinophilic granular cells, macrophages, lymphocytes, and fibrocytes. The post-smolts were caught in May during the early oceanic phase of their life cycle. In these fish, A. simplex sensu stricto larvae were found lying free on the serosal surface of the intestine and liver without any apparent histologic changes. This is the earliest in the marine migration of Atlantic salmon that A. simplex sensu stricto infection has been recorded.