Immunohistochemical localization of a gap junction protein (connexin43) in the muscularis externa of murine,canine, and human intestine

Electron-microscopic studies have revealed a heterogeneous distribution of gap junctions in the muscularis externa of mammalian intestines. This heterogeneity is observed at four different levels: among species; between small and large intestines; between longitudinal and circular muscle layers; and between subdivisions of the circular muscle layer. We correlated results obtained with two immunomethods, using an antibody to the known gap-junctional protein (connexin43) with ultrastructural findings, and further evaluated the respective sensitivity of these two approaches. For comparative reasons we also included the vascular smooth muscle of coronary arteries into our study. Two versions of the immunotechnique (peroxidase-antiperoxidase and fluorescence methods) were applied to frozen sections of murine, canine, and human small and large intestines, as well as to pig coronary artery. In the small intestine of all three species a very strong reactivity marked the outer main division of the circular muscle layer, while the longitudinal muscle layer as well as the inner thin division of the circular muscle layer were negative. In murine and human colon both muscle layers were negative, while in canine colon the border layer between the circular muscle and the submucosa reacted strongly, and scattered activity was found in the portion of the circular muscle layer (one tenth of its thickness) closest to the submucosa. The remainder of the circular muscle layer and the entire longitudinal muscle layer were negative in the canine colon. In the coronary artery we could not confirm the positive, specific labeling reported by other investigators (l.c.). In conclusion, we found close correlations at all four above-mentioned levels in the distribution of gap junctions in the gut musculature, as determined by binding of anticonnexin43 in comparison to conventional ultrastructural studies. Since no significant immunostaining was found in (i) the outer border of the circular muscle layer of the canine colon and (ii) the border layer between the submucosa and the circular muscle layer of human colon, where rare gap junctions have been identified at the ultrastructural level, we conclude that the electron-microscopic analysis is the more sensitive of the two methods.     

Comparison of the biological activity of canine and porcine motilin in rabbit

The biological activity of porcine and canine motilin was studied in rabbits by establishing dose-response curves of both peptides using two different methods. The dissociation constant, obtained from the displacement of iodinated porcine motilin by canine motilin was 0.6 +/- 0.3 nM, versus 1.2 +/- 0.4 nM for porcine motilin. For the 13-norleucine and 13-leucine analogues of porcine motilin a value of 0.8 +/- 0.3 nM was obtained. Both motilins were almost equipotent in stimulating the in vitro contractile response of longitudinal smooth muscle strips: half-maximal effect was achieved at a concentration of 1.0 +/- 0.1 nM for canine versus 1.3 +/- 0.2 nM for the 13-norleucine analogue of porcine motilin. We conclude that porcine and canine motilin have a comparable bioactivity in the rabbit, although canine motilin is slightly more effective. The motilin receptor is probably specific for the N-terminal portion which is identical in both molecules.     

Work partitioning of transversally loaded muscle: experimentation and simulation

Skeletal muscles are surrounded by other muscles, connective tissue and bones, which may transfer transversal forces to the muscle belly. Simple Hill-type muscle models do not consider transversal forces. Thus, the aim of this study was to examine and model the influence of transversal muscle loading on contraction dynamics, e.g. on the rate of force development and on the maximum isometric muscle force (F_im). Isometric experiments with and without transversal muscle loading were conducted on rat muscles. The muscles were loaded (1.3 N cm^? 2) by a custom-made plunger which was able to move in transversal direction. Then the muscle was fully stimulated, the isometric force was measured at the distal tendon and the movement of the plunger was captured with a high-speed camera. The interaction between the muscle and the transversal load was modelled based on energy balance between the (1) work done by the contractile component (CC) and (2) the work done to lift the load, to stretch the series elastic structures and to deform the muscle. Compared with the unloaded contraction, the force rate was reduced by about 25% and F_im was reduced by 5% both in the experiment and in the simulation. The reduction in F_im resulted from using part of the work done by the CC to lift the load and deform the muscle. The response of the muscle to transversal loading opens a window into the interdependence of contractile and deformation work, which can be used to specify and validate 3D muscle models.     

Functional links between canine height and jaw gape in catarrhines with special reference to early hominins

This study tests the hypothesis that decreased canine crown height in catarrhines is linked to (and arguably caused by) decreased jaw gape. Associations are characterized within and between variables such as upper and lower canine height beyond the occlusal plane (canine overlap), maximum jaw gape, and jaw length for 27 adult catarrhine species, including 539 living subjects and 316 museum specimens. The data demonstrate that most adult male catarrhines have relatively larger canine overlap dimensions and gapes than do conspecific females. For example, whereas male baboons open their jaws maximally more than 110% of jaw length, females open about 90%. Humans and hylobatids are the exceptions in that canine overlap is nearly the same in both the sexes and so is relative gape (ca. 65% for humans and 110% for hylobatids). A correlation analysis demonstrates that a large portion of relative gape (maximum gape/projected jaw length) is predicted by relative canine overlap (canine overlap/jaw length). Relative gape is mainly a function of jaw muscle position and/or jaw muscle‐fiber length. All things equal, more rostrally positioned jaw muscles and/or shorter muscle fibers decrease gape and increase bite force during the power stroke of mastication, and the net benefit is to increase the mechanical efficiency during chewing. Similarly, more caudally positioned muscles and/or longer muscle fibers increase the amount of gape and decrease bite force. Overall, the data support the hypothesis that canine reduction in early hominins is functionally linked to decreased gape and increased mechanical efficiency of the jaws. Am J Phys Anthropol, 2013. © 2012 Wiley Periodicals, Inc.     

The pharyngoesophageal transition

Estimated are the origin areas of the cricopharyngeal part of the constrictor pharyngis inferior muscle, of the longitudinal fibre system of the oesophagus, the cricopharyngeal ligament and the upper border of the circular fibres of the oesophagus. Measured were also the border of the oesophagus which is to be seen sidewards of the trachea. The results of the preparations and measurements are discussed with investigations of earlier researchers.     

Muscle force depends on the amount of transversal muscle loading

Skeletal muscles are embedded in an environment of other muscles, connective tissue, and bones, which may transfer transversal forces to the muscle tissue, thereby compressing it. In a recent study we demonstrated that transversal loading of a muscle with 1.3 N cm⁻² reduces maximum isometric force (F_im) and rate of force development by approximately 5% and 25%, respectively. The aim of the present study was to examine the influence of increasing transversal muscle loading on contraction dynamics.     

Deformation and three-dimensional displacement of fibers in isometrically contracting rat plantaris muscles

In this study, the deformation of different fibers of the rat m. plantaris during "isometric" contractions at different muscle lengths was considered. Because the m. plantaris has an obviously inhomogeneous architecture, its fibers on the medial side of the muscle belly are judged to be shorter than those on the lateral side of it. It was expected that longitudinal deformation of different fibers would vary accordingly. A 3D video analysis of contracting muscle showed that longitudinal strain of fibers as a function of muscle length does not differ between fibers on different sides of the muscle. Apart from longitudinal shortening, the fibers were also displaced laterally during a contraction. The fibers displaced during a contraction in a direction perpendicular to their longitudinal axis. The displacement of the fibers occurred asymmetrically, resulting in a helical deformation of the whole muscle. It is concluded that the asymmetric displacement and the helical deformation must result from transversal forces between the fibers. It is hypothesized that these transversal forces cancel out differences in longitudinal strains that might exist between fibers.     

Autoantibodies in canine masticatory muscle myositis recognize a novel myosin binding protein-C family member

Inflammatory myopathies are a group of autoimmune diseases that affect muscles. In humans, the most common inflammatory myopathies are polymyositis, dermatomyositis, and inclusion body myositis. Autoantibodies may be found in humans with inflammatory myopathies, and these play an important role in diagnosis and disease classification. However, these Abs are typically not muscle specific. Spontaneously occurring canine inflammatory myopathies may be good parallel disorders and provide insights into human myositis. In dogs with inflammatory myopathy, muscle-specific autoantibodies have been found, especially in masticatory muscle myositis. We have identified the major Ag recognized by the autoantibodies in canine masticatory muscle myositis. This Ag is a novel member of the myosin binding protein-C family, which we call masticatory myosin binding protein-C (mMyBP-C). mMyBP-C is localized not only within the masticatory muscle fibers, but also at or near their cell surface, perhaps making it accessible as an immunogen. The gene for mMyBP-C also exists in humans, and mMyBP-C could potentially play a role in certain human inflammatory myopathies. Understanding the role of mMyBP-C in this canine inflammatory myopathy may advance our knowledge of mechanisms of autoimmune inflammatory muscle diseases, not only in dogs, but also in humans.     

Free radical centers in the dog myocardial tissue in regional ischemia

The effect of regional ischemia on canine myocardial in situ free radical species was studied by the EPR method. Rapid fixation of heart muscle samples by freezeclamping was performed at the following physiological states: native myocardial blood circulation, regional ischemia with the presence of collateral circulation, total ischemia, and postischemic reperfusion. EPR spectra of the samples at -40 degrees C exhibited two free radical signals from the semireduced forms of ubiquinone and flavine coenzymes. Upon transition from normal blood supply to regional ischemia, an increase in the contribution of the flavine signal was registered, but reperfusion resulted in the recovery of the characteristics of EPR signals. It was found that the increase in the intensity of collateral circulation in the ischemic area led to an increase in the portion of ubisemiquinone in the integral EPR signal, whereas in total ischemia this signal was not registered. It was shown that the changes in spectral characteristics of integral free radical signals are accompanied by changes in their relaxation parameters.     

Galanin: an inhibitory neural peptide of the canine small intestine

Galanin injected intraarterially during phasic activity of the canine small intestine in vivo produced inhibition. Fifty percent inhibition occurred at 1.5 +/- 0.5 X 10(-10) mols lasting for 0.7 min. The inhibitory response was not decreased by treatment with atropine, hexamethonium, yohimbine or naloxone, suggesting that muscarinic, nicotinic, alpha 2 adrenergic or opiate receptors were not being stimulated. Since tetrodotoxin blockade of nerves did not reduce the response and galanin at 10(-10) mols was able to eliminate the smooth muscle response to intraarterial acetylcholine, we suggest that galanin acts to inhibit smooth muscle directly. Galanin 10(-9) M added to the muscle bath also inhibited phasic activity of the canine ileum circular muscle in vitro in the presence of tetrodotoxin. These results suggest that the neural peptide galanin may be a non-adrenergic, non-cholinergic, non-opioid neurotransmitter in the canine small intestine.     

Effect of sighs on breathing memory and dynamics in healthy infants.

Deep inspirations (sighs) play a significant role in altering lung mechanical and airway wall function; however, their role in respiratory control remains unclear. We examined whether sighs act via a resetting mechanism to improve control of the respiratory regulatory system. Effects of sighs on system variability, short- and long-range memory, and stability were assessed in 25 healthy full-term infants at 1 mo of age [mean 36 (range 28-57) days] during quiet sleep. Variability was examined using moving-window coefficient of variation, short-range memory using autocorrelation function, and long-range memory using detrended fluctuation analysis. Stability was examined by studying the behavior of the attractor with use of phase-space plots. Variability of tidal volume (VT) and minute ventilation (VE) increased during the initial 15 breaths after a sigh. Short-range memory of VT decreased during the 50 breaths preceding a sigh, becoming uncorrelated (random) during the 10-breath presigh window. Short-range memory increased after a sigh for the entire 50 breaths compared with the randomized data set and for 20 breaths compared with the presigh window. Similar, but shorter duration, changes were noted in VE. No change in long-range memory was seen after a sigh. Coefficient of variation and range of points located within a defined attractor segment increased after a sigh. Thus control of breathing in healthy infants shows long-range stability and improvement in short-range memory and variability after a sigh. These results add new evidence that the role of sighs is not purely mechanical.     

Impact of transversal calf muscle loading on plantarflexion

Muscle compression commonly occurs in daily life (for instance wearing backpacks or compression garments, and during sitting). However, the effects of the compression on contraction dynamics in humans are not well examined. The aim of the study was to quantify the alterations of contraction dynamics and muscle architecture in human muscle with external transverse loads.The posterior tibialis nerve of 29 subjects was stimulated to obtain the maximal double-twitch force of the gastrocnemius muscle with and without transverse compression that was generated using an indentor. The muscle architecture was determined by a sonographic probe that was embedded within the indentor. Five stimulations each were conducted at 5 conditions: (1) pretest (unloaded), (2) indentor loading with 2 kg, (3) 4.5 kg, (4) 10 kg, and (5) posttest (unloaded).Compared to the pretest maximal force decreased by 9%, 13% and 16% for 2 kg, 4.5 kg and 10 kg, respectively. The half-relaxation time increased with increased transverse load whereas the rate of force development decreased from pretest to 2 kg and from 4.5 kg to 10 kg. The lifting height of the indentor increased with transverse load from 2 kg to 4.5 kg but decreased from 4.5 kg to 10 kg. Increases in pennation during the twitches were reduced at the highest transverse load.The results demonstrate changes of the contraction dynamics due to transversal muscle loading. Those alterations are associated with the applied pressure, changes in muscle architecture and partitioning of muscle force in transversal and longitudinal direction.     

Structural characterization of interstitial cells of Cajal in myenteric plexus and muscle layers of canine colon

We have carried out a detailed ultrastructural study of the interstitial cells near the myenteric plexus of the canine colon and defined the structural characteristics which distinguish them from other resident non-neural cells. We have also examined the interconnections of these interstitial cells with nerves, the longitudinal muscle, and the circular muscle. In addition, we sought connections between interstitial cells of the myenteric plexus and those described earlier at the inner border of the circular muscle in proximal and distal colon. The interstitial cells of the myenteric plexus were structurally distinctive, and made gap junctions with one another and occasionally with smooth muscle. There seemed to be two subsets of these interstitial cells, one associated with the longitudinal muscle and the other with the circular muscle. Cells of both subsets were often close (less than or equal to 20 nm) to nerve profiles. The interstitial cells near the longitudinal muscle layer penetrated slightly into the muscle layer, but those near the circular muscle did not and neither set contacted the other. Moreover, interstitial cells of Cajal located near the myenteric plexus were never observed to contact those at the inner border of circular muscle. The interstitial cells of Cajal at the canine colon myenteric plexus are structurally organized to provide independent pacemaking activities for the longitudinal and adjacent circular muscle. Their dense innervation suggests that they mediate neural modulation of intestinal pacemaker activities. Moreover, they lack direct contacts with the interstitial cell network at the inner border of circular muscle, which is essential for the primary pacemaking activity of circular muscle. The structural organization of interstitial cells in canine colon is consistent with their proposed role in pacemaking activity of the two muscle layers.     

Histochemical and stereological analysis of the levator ani (pubocaudal) muscle in nulliparous and multiparous beagles

Fiber-type composition and several stereological parameters of the levator ani (pubocaudal) muscle were evaluated in five nulliparous and five multiparous beagles using myosin ATPase-histochemistry and systematically selected muscle cross-sections. With respect to the narrow canine pelvic cavity, this study was also undertaken to determine whether vaginal birth of at least seven litters causes similar neuromuscular changes in the canine levator ani (pubocaudal) muscle analoguous to those seen in the pelvic floor muscles of women after vaginal delivery. The canine pubocaudal muscle is comprised of approximately equal amounts of slow twitch type I and fast twitch type II (IIA, IIS) fibers. The muscles of both the nulliparous and multiparous beagles did not display any signs indicative of denervation or myopathology. The multiparous dogs exhibited significantly increased mean absolute muscle (1720 mm(3)) and total fiber-type I volumes (850 mm(3)) as well as relevantly increased mean diameter of type I fibers (72.0 microm) when compared with the nulliparous group. The canine levator ani (pubocaudal) muscle is not pathologically affected by vaginal deliveries and seems to adapt to numerous successive pregnancies and births through fiber-type I hypertrophy.     

Tenotomy of the avian anterior latissimus dorsi muscle. I. Effect of age on fiber-type transformation and regeneration from the stump in chicks

The chick's anterior latissimus dorsi muscle (ALD) was tenotomized at its origin at either 1 day or 4 weeks of age, and investigated histochemically and ultrastructurally at intervals thereafter to determine whether muscle fiber-type transformation from a slow to a twitch type is greater in young birds than older birds. No transformation of fiber type occurred in either procedure, but a new muscular connection regenerated between the scar tissue at the end of the original tenotomized stump and the former origin. This regenerated muscle had a mosaic pattern of fiber types, as demonstrated by myofibrillar ATPase activity, and contained predominantly fast fibers, as contrasted with controls or the tenotomized portion, which contained predominantly slow tonic muscle fibers. The regenerated portion contained muscle spindles. The original portion of the tenotomized muscle was indistinguishable from the control muscle. These responses of the chick ALD to tenotomy are quite different from those in the pigeon, which are reported in the following study.     

The microcirculation of cerebral arteries: a morphologic and morphometric examination of the major canine cerebral arteries

A morphologic and morphometric examination of the major cerebral blood vessels in the dog was carried out to determine whether there were vasa vasorum in these arteries and what features might be associated with them. True vasa vasorum confined to the media were not seen in any of the vessels examined. Microvessels confined to the adventitia, however, were found in the internal carotid and vertebral arteries but not in the basilar, middle cerebral, or anterior spinal arteries. Animal size, vessel size as determined by adventitial and medial area, and the number of smooth muscle cell lamellae were not associated with the presence of these adventitial vessels; they occurred only in arteries with both an intra- and extradural portion. It therefore appears that most canine cerebral arteries do not have vasa vasorum.     

On the anisotropy of the canine diaphragmatic central tendon

We studied the mechanical and anatomical anisotropy of the canine diaphragmatic central tendon (CT). Dumb-bell-shaped strips with effective dimensions of 10 x 2 mm (length x width) were cut from different regions of the canine diaphragmatic CT in two different orientations relative to the direction of neighboring muscle fibers. Specimens sampled with their long axial dimension oriented parallel to the neighboring muscle fibers were named Group-1 and those sampled with an orientation perpendicular to the neighboring muscle fibers were named Group-2. Results from one-dimensional stress-strain and tensile failure strength tests revealed that the CT is a nonlinear, inelastic, and anisotropic material. Group-1 specimens were found to have a higher stiffness, higher failure strength and higher strain energy density at failure than Group-2 specimens. Polarized microscopy showed that multiple sheets of collagen fiber bundles formed an orthogonal network in the tendon. Collagen fiber bundles along Group-1 direction formed parallel trajectory lines connecting the neighboring costal and crural muscles; bundles along Group-2 direction were observed to orient 90 degrees away. At the central apex region of the CT, collagen bundles of Group-1 formed a fan-like trajectory pattern. This collagen network architecture was compared favorably to the trajectories of an approximated principal stress field in the CT due to simulated contractile forces from its adjacent costal and crural muscles. These combined results suggest a structure-function relationship for the anatomical and mechanical anisotropy in the canine diaphragmatic CT.     

The effect of vasoactive intestinal polypeptide (VIP) on the canine gallbladder motility.

1. VIP at doses of 10(-9) to 10(-8) M was ineffective and at doses of 5 x 10(-8) to 10(-7) M exerted a slight inhibitory effect on the tone of the canine gallbladder muscle strip. However, VIP (0.1-1 micrograms/kg) injected intravenously (i.v.) in conscious dogs dose-dependently decreased the gallbladder pressure. 2. VIP did not influence significantly the acetylcholine (ACh)- or carbachol- induced contractions of canine gallbladder under in vitro or in vivo conditions, but it decreased the electrically-induced, atropine-sensitive contractions of gallbladder muscle strips. 3. VIP (5 x 10(-9) to 5 x 10(-8) M) did not influence significantly the dose-response curve for cholecystokinin octapeptide (CCK OP) of canine and guinea-pig gallbladder muscle strips. VIP injected i.v. (0.1-0.5 micrograms/kg) in conscious dogs greatly decreased the CCK OP-induced gallbladder pressure.