Novel insights into the nervous system affected by prolonged hyperglycemia.

Multiple molecular pathways including the receptor for advanced glycation end-products-diaphanous related formin 1 (RAGE-Diaph1) signaling are known to play a role in diabetic peripheral neuropathy (DPN). Evidence suggests that neuropathological alterations in type 1 diabetic spinal cord may occur at the same time as or following peripheral nerve abnormalities. We demonstrated that DPN was associated with perturbations of RAGE-Diaph1 signaling pathway in peripheral nerve accompanied by widespread spinal cord molecular changes. More than 500 differentially expressed genes (DEGs) belonging to multiple functional pathways were identified in diabetic spinal cord and of those the most enriched was RAGE-Diaph1 related PI3K-Akt pathway. Only seven of spinal cord DEGs overlapped with DEGs from type 1 diabetic sciatic nerve and only a single gene cathepsin E (CTSE) was common for both type 1 and type 2 diabetic mice. In silico analysis suggests that molecular changes in spinal cord may act synergistically with RAGE-Diaph1 signaling axis in the peripheral nerve. KEY MESSAGES: Molecular perturbations in spinal cord may be involved in the progression of diabetic peripheral neuropathy. Diabetic peripheral neuropathy was associated with perturbations of RAGE-Diaph1 signaling pathway in peripheral nerve accompanied by widespread spinal cord molecular changes. In silico analysis revealed that PI3K-Akt signaling axis related to RAGE-Diaph1 was the most enriched biological pathway in diabetic spinal cord. Cathepsin E may be the target molecular hub for intervention against diabetic peripheral neuropathy.

The applicability of Calcofluor White (CWS) and Fluorescent Brightener (CFB) dyes for confocal laser microscopic analysis (CLSM) of various ß-glucans in selected dairy products and water.

The aim of this study was to determine the applicability of commonly used dyes (CWS and CFB) to stain different types of ß-glucans in water and selected, most popular, dairy products for CLSM imaging analysis. Structurally different pure ß-glucan preparations: curdlan-CU, oat ß-glucan-OG and scleroglucan-SC were tested. Our study showed for the first time that CWS is a dye with a specific affinity for all analyzed ß-glucans, while CFB is suitable for curdlan labeling only. Despite the technological processes, ß-glucans structures in dairy products are similar to those in aqueous suspension forms; each of the ß-glucans forms a different and characteristic structure, ranging from spindle-shaped and branched till granular. The presented results for the first time systematize the knowledge on CWS and CFB applicability in ß-glucan CLSM staining and allow future researchers to correctly analyze simultaneously stained ß-glucan, fat, and protein in a complex matrix as dairy products.

Role of RAGE in the Pathogenesis of Neurological Disorders.

This review reflects upon our own as well as other investigators' studies on the role of receptor for advanced glycation end-products (RAGE), bringing up the latest information on RAGE in physiology and pathology of the nervous system. Over the last ten years, major progress has been made in uncovering many of RAGE-ligand interactions and signaling pathways in nervous tissue; however, the translation of these discoveries into clinical practice has not come to fruition yet. This is likely, in part to be the result of our incomplete understanding of this crucial signaling pathway. Clinical trials examining the therapeutic efficacy of blocking RAGE-external ligand interactions by genetically engineered soluble RAGE or an endogenous RAGE antagonist, has not stood up to its promise; however, other trials with different blocking agents are being considered with hope for therapeutic success in diseases of the nervous system.

The Involvement of RAGE and Its Ligands during Progression of ALS in SOD1 G93A Transgenic Mice.

Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease characterized by a progressive degeneration of upper and lower motor neurons that causes paralysis and muscle atrophy. The pathogenesis of the disease is still not elucidated. Receptor for Advanced Glycation End Product (RAGE) is a major component of the innate immune system and has implications in ALS pathogenesis. Multiple studies suggest the role of RAGE and its ligands in ALS. RAGE and its ligands are overexpressed in human and murine ALS motor neurons, astrocytes, and microglia. Here, we demonstrated the expression of RAGE and its ligands during the progression of the disease in the transgenic SOD1 G93A mouse lumbar spinal cord. We observed the highest expression of HMGB1 and S100b proteins at ALS onset. Our results highlight the potential role of RAGE and its ligands in ALS pathogenesis and suggest that some of the RAGE ligands might be used as biomarkers in early ALS diagnosis and potentially be useful in targeted therapeutic interventions at the early stage of this devastating disease.

Cocaine and Amphetamine Regulated Transcript (CART) Expression Changes in the Stomach Wall Affected by Experimentally Induced Gastric Ulcerations.

Cocaine- and amphetamine-regulated transcript (CART) is a peptide suggested to play a role in gastrointestinal tract tissue reaction to pathology. Gastric ulceration is a common disorder affecting huge number of people, and additionally, it contributes to the loss of pig livestock production. Importantly, ulceration as a focal disruption affecting deeper layers of the stomach wall differs from other gastrointestinal pathologies and should be studied individually. The pig's gastrointestinal tract, due to its many similarities to the human counterpart, provides a valuable experimental model for studying digestive system pathologies. To date, the role of CART in gastric ulceration and the expression of the gene encoding CART in porcine gastrointestinal tube are completely unknown. Therefore, we aimed to verify the changes in the CART expression by Q-PCR (gene encoding CART in the tissue) and double immunofluorescence staining combined with confocal microscopy (CART immunofluorescence in enteric nervous system) in the porcine stomach tissues adjacent to gastric ulcerations. Surprisingly, we found that gastric ulcer caused a significant decrease in the expression of CART-encoding gene and huge reduction in the percentage of CART-immunofluorescent myenteric perikarya and neuronal fibers located within the circular muscle layer. Our results indicate a unique CART-dependent gastric response to ulcer disease.

Inferior vagal ganglion galaninergic response to gastric ulcers.

Galanin is a neuropeptide widely expressed in central and peripheral nerves and is known to be engaged in neuronal responses to pathological changes. Stomach ulcerations are one of the most common gastrointestinal disorders. Impaired stomach function in peptic ulcer disease suggests changes in autonomic nerve reflexes controlled by the inferior vagal ganglion, resulting in stomach dysfunction. In this paper, changes in the galaninergic response of inferior vagal neurons to gastric ulceration in a pig model of the disease were analyzed based on the authors' previous studies. The study was performed on 24 animals (12 control and 12 experimental). Gastric ulcers were induced by submucosal injections of 40% acetic acid solution into stomach submucosa and bilateral inferior vagal ganglia were collected one week afterwards. The number of galanin-immunoreactive perikarya in each ganglion was counted to determine fold-changes between both groups of animals and Q-PCR was applied to verify the changes in relative expression level of mRNA encoding both galanin and its receptor subtypes: GalR1, GalR2, GalR3. The results revealed a 2.72-fold increase in the number of galanin-immunoreactive perikarya compared with the controls. Q-PCR revealed that all studied genes were expressed in examined ganglia in both groups of animals. Statistical analysis revealed a 4.63-fold increase in galanin and a 1.45-fold increase in GalR3 mRNA as compared with the controls. No differences were observed between the groups for GalR1 or GalR2. The current study confirmed changes in the galaninergic inferior vagal ganglion response to stomach ulcerations and demonstrated, for the first time, the expression of mRNA encoding all galanin receptor subtypes in the porcine inferior vagal ganglia.

Nerve structures of the heart and their immunohistochemical characterization in 10-week-old porcine foetuses.

The aim of the present study was to investigate the distribution of neuronal perikarya and nerve fibres and their chemical coding in the heart of 10-week-old porcine foetuses. The foetuses, obtained from a local slaughterhouse, were fixed by immersion in 4% buffered (pH 7.4) paraformaldehyde. Cryostat sections of the hearts were processed for single- or double-labelling immunofluorescence methods using antibodies against protein gene product (PGP) dopamine beta-hydroxylase (DßH), acetylcholine vesicular transporter (VAChT) and calcitonin gene-related peptide (CGRP). Numerous clusters of the PGP-positive nerve cells were observed throughout the heart wall. The majority of the clusters were found beneath the epicardium around the root of the aorta, pulmonary trunk and main veins. Some single PGP-positive neurons or small clusters of nerve cells were observed in the epicardium of heart atria and ventricles. The richest network of PGP-positive nerve fibres was observed in the base of the heart near the main cardiac blood vessel openings. Many bundles of PGP-positive nerve fibres were observed throughout the four cardiac chambers. Double-immunohistochemical staining revealed that the majority of the neurons contained immunoreactivity to VAChT, some of them stained for CGRP and the single neuronal somata were DßH-positive. The nerve fibres supplying the heart expressed immunoreactivity to DßH, VAChT or CGRP. The distribution and neurochemical coding of the nerve structures in ten-week-old foetuses are different from those observed in the hearts of juvenile pigs.

L'objectif du présent étude était d'examiner la disposition des cellules et des fibres nerveuses, ainsi que de leur codage, aux cœurs de fœtus de porc de 10 semaines. Les fœtus, obtenus dans un abattoir locale, ont été fixés par immersion dans la solution de paraformaldéhyde 4% (pH 7,4). Des coupes de cœurs, obtenues à l'aide du cryostat, ont subi une coloration immunohistochimique simple ou double afin de dépister, dans des structures nerveuses, la présence du produit génétique protéiné (PGP), de la dopamine bêta-hydroxylase (DßH), du transporteur vésiculaire de l'acétylcholine (VAChT) et du peptide relié au gène calcitonine (CGRP). De nombreuses concentrations de cellules nerveuses PGP-positives ont été observées dans la paroi cardiaque. La plupart des concentrations ont été retrouvées en dessous de l'épicarde, autour de la racine de l'aorte, du tronc pulmonaire et des veines principales. Des neurones singulaires PGP-positifs ou des petites concentrations de cellules nerveuses ont été observées dans l'épicarde des oreillettes et des ventricules du cœur. Le plus riche réseau de fibres nerveuses PGP-positives a été observé dans la base du cœur, près de l'acheminement des vaisseaux principaux. Dans les quatre chambres du cœur, de nombreux faisceaux de fibres nerveuses PGP-positives ont été observées. Des colorations immunohistochimiques doubles ont démontré que la plupart de neurones présentaient une immunoréactivité par rapport à VAChT, certains par rapport à CGRP, et des corps singulaires de cellules nerveuses étaient positives par rapport à DßH. Des fibres nerveuses alimentant le cœur présentaient une immunoréactivité par rapport à DßH, VAChT ou CGRP. La distribution et le codage neurochimique des structures nerveuses chez des fœtus de 10 semaines diffèrent de ceux observés dans les cœurs de jeunes porcs.

Innervation of internal female genital organs in the pig during prenatal development.

This study investigated the innervation of internal genital organs in 5-, 7- and 10-week-old female pig foetuses using single and double-labelling immunofluorescence methods. The structure and topography of the organs was examined using a scanning electron microscope (SEM). The investigations revealed differences in the innervation between the three developmental periods. Immunostaining for protein gene product 9.5 (PGP; general neural marker) disclosed solitary nerve fibres in the external part of the gonadal ridge and just outside of the mesenchyme surrounding mesonephric ducts in 5-week-old foetuses. Double-labelling immunohistochemistry revealed that nerve fibres associated with the ridge expressed dopamine ß-hydroxylase (DßH; adrenergic marker) or vesicular acetylcholine transporter (VAChT; cholinergic marker). In 7-week-old foetuses, the PGP-positive nerve terminals were absent from the gonad but some of them ran outside and along, and sometimes penetrated into the mesenchyme surrounding the tubal and uterine segments of the paramesonephric ducts and uterovaginal canal. Few axons penetrated into the mesenchyme. DßH-positive fibres were found in single nerve strands or bundles distributed at the edge of the mesenchyme. VAChT-positive nerve terminals formed delicate bundles located at the edge of the mesenchyme, and the single nerves penetrated into the mesenchyme. DßH was also expressed by neurons which formed cell clusters comprising also DßH- or VAChT-positive nerve fibres. In 10-week-old foetuses, PGP-positive nerve fibres were still absent from the ovary but some were distributed in the mesenchyme associated with the uterovaginal canal and uterine and a tubal segment of the paramesonephric ducts, respectively. DßH- or VAChT-positive nerve fibres were distributed at the periphery of the mesenchyme associated with the uterovaginal canal. Some DßH- and many VAChT-positive nerve fibres were evenly distributed throughout the mesenchyme. The clusters of nerve cells comprised DßH-positive perikarya and DßH- or VAChT-positive nerve fibres. The investigations revealed no DßH/VAChT-positive nerve fibres or neurons as well as no nerve structures stained for calcitonin gene-related peptide and/or substance P (sensory markers) associated with the genital organs in the studied prenatal periods.

Gastric ulcer induced changes in substance P and Nk1, Nk2, Nk3 receptors expression in different stomach localizations with regard to intrinsic neuronal system.

Gastric ulceration, a focal tissue damage accompanied by inflammation, can influence other parts of the stomach. Substance P and its receptors are strongly involved in regulation of gastrointestinal motility, secretion and inflammation. The enteric nervous system is one of the regulators of gastrointestinal functioning and contributes to tissue response to the pathology. The pig, an omnivorous animal, is a valuable species for gastrointestinal experiments. Thus, the objective of the study was to verify whether the antral ulceration induces changes in the expression of substance P and tachykinin receptors in the neighboring (antrum) and distanced (corpus, pylorus) porcine gastric tissues and therein localized myenteric and submucosal perikarya as well as in the intrinsic descending neurons supplying pyloric sphincter. The experiment was performed on healthy pigs and pigs with experimentally induced gastric ulcers. Stomach samples from the corpus, antrum (adjacent to the ulcer in experimental pigs) and pylorus were analyzed by: (1) double immunofluorescence for changes in the number of SP-positive myenteric and submucosal neurons (2) Real-Time PCR for changes in expression of mRNA encoding SP and Nk1, Nk2, Nk3 receptors. Additionally, gastric descending neurons supplying pyloric sphincter were immunostained for SP. In experimental animals, only the number of SP-positive myenteric perikarya significantly increased in all stomach localizations studied. Q-PCR revealed increased expression for: SP, Nk1, Nk3 in the corpus; Nk2 and Nk3 in the pylorus; In the antrum, expression of Nk3 was increased but Nk2-decreased. Antral ulcers induced significant changes in the expression of SP and tachykinin receptors in the wide stomach area indicating sophisticated tissue reaction.

Neuropeptides and lymphocyte populations in the porcine ileum and ileocecal lymph nodes during postnatal life.

The maturation-related changes in the concentrations of galanin (Gal), vasoactive intestinal polypeptide (VIP), substance P (SP) and somatostatin (Som), as well as in subpopulations of lymphocytes expressing antigens CD2 (lymphocytes T), CD4 (T helper), CD8 (T cytotoxic), CD21 (B lymphocytes), CD5-/CD8+ (NK cells) and TCRgamma/delta (gut mucosal/intraepitelial cells) were studied in the ileal Peyer's patches and ileo-cecal lymph nodes in female pigs aged 3 days, 2 weeks, 4 weeks and 4 months. As regards neuropeptide concentrations statistically significant changes in the ileum and lymph nodes were found only in case of Gal and VIP. The concentrations of neuropeptides were significantly higher only in new-born animals. As regards the changes in subpopulations of lymphocytes, statistically significant changes were noticed only in 4-months old animals and were dealing only with CD2+ and TCRgamma/delta cells in the ileum as well as CD4+, CD8+, CD21+ and TCRgamma/delta in lymph nodes. The highest number of CD8+, CD21+ and TCRgamma/delta lymphocytes occurred in 4-months old animals.

Morphological and neuroanatomical study of the mammary gland in the immature and mature European beaver (Castor fiber).

This study investigated general morphology and immunohistochemical properties of nerve fibres supplying the mammary gland (MG) in the European beaver. The microscopic analysis of the beaver mammary gland revealed the presence of morphological structures which are characteristic for mammals. There were no distinct differences in the morphological features of the mammary gland between the juvenile and non-pregnant mature beaver. The nerve fibres were visualized using antibodies against protein gene product 9.5 (PGP) and biologically active substances including ß-hydroxylase tyrosine (DßH), neuropeptide Y (NPY), calcitonine gene-related peptide (CGRP) and substance P (SP). The study has revealed that the MG in the juvenile and mature beaver is richly supplied with PGP-immunoreactive (PGP-IR) nerve fibres. The most abundant innervation was observed in the nipple and less numerous nerve terminals supplied the glandular tissue. Double-labelling immunohistochemistry disclosed that the majority of PGP-IR nerve fibres associated with blood vessels and smooth muscle cells in both the nipple and glandular tissue were also DßH-IR. However, these nerve terminals were less numerous in the glandular tissue than in the nipple. Most of the DßH-IR axons associated with arteries and smooth muscle cells in the entire gland also stained for NPY. Small number of DßH/NPY-IR fibres supplied veins. CGRP-IR fibres were more abundant than those expressing SP. No distinct differences in the distribution and immunohistochemical characteristic of nerve fibres were observed between the juvenile and adult animals. The distribution and immunohistochemical properties of nerve fibres supplying the gland in the beaver remind those previously described in other mammalian species.

The Influence of Gastric Antral Ulcerations on the Expression of Galanin and GalR1, GalR2, GalR3 Receptors in the Pylorus with Regard to Gastric Intrinsic Innervation of the Pyloric Sphincter.

Gastric antrum ulcerations are common disorders occurring in humans and animals. Such localization of ulcers disturbs the gastric emptying process, which is precisely controlled by the pylorus. Galanin (Gal) and its receptors are commonly accepted to participate in the regulation of inflammatory processes and neuronal plasticity. Their role in the regulation of gastrointestinal motility is also widely described. However, there is lack of data considering antral ulcerations in relation to changes in the expression of Gal and GalR1, GalR2, GalR3 receptors in the pyloric wall tissue and galaninergic intramural innervation of the pylorus. Two groups of pigs were used in the study: healthy gilts and gilts with experimentally induced antral ulcers. By double immunocytochemistry percentages of myenteric and submucosal neurons expressing Gal-immunoreactivity were determined in the pyloric wall tissue and in the population of gastric descending neurons supplying the pyloric sphincter (labelled by retrograde Fast Blue neuronal tracer). The percentage of Gal-immunoreactive neurons increased only in the myenteric plexus of the pyloric wall (from 16.14±2.06% in control to 25.5±2.07% in experimental animals), while no significant differences in other neuronal populations were observed between animals of both groups. Real-Time PCR revealed the increased expression of mRNA encoding Gal and GalR1 receptor in the pyloric wall tissue of the experimental animals, while the expression(s) of GalR2 and GalR3 were not significantly changed. The results obtained suggest the involvement of Gal, GalR1 and galaninergic pyloric myenteric neurons in the response of pyloric wall structures to antral ulcerations.

The Influence of Antral Ulcers on Intramural Gastric Nerve Projections Supplying the Pyloric Sphincter in the Pig (Sus scrofa domestica)-Neuronal Tracing Studies.

BACKGROUND: Gastric ulcerations in the region of antrum pylori represent a serious medical problem in humans and animals. Such localization of ulcers can influence the intrinsic descending nerve supply to the pyloric sphincter. The pyloric function is precisely regulated by intrinsic and extrinsic nerves. Impaired neural regulation could result in pyloric sphincter dysfunction and gastric emptying malfunction. The aim of the study was to determine the effect of gastric antral ulcerations on the density and distribution of intramural gastric descending neurons supplying the pyloric sphincter in pigs. METHODOLOGY/PRINCIPAL FINDINGS: The experiment was performed on 2 groups of pigs: healthy gilts (n=6) and gilts with experimentally induced peptic ulcers in the region of antrum pylori (n=6). Gastric neurons supplying pyloric sphincter were labeled using the retrograde neuronal tracing technique (20µl of Fast Blue tracer injected into the pyloric sphincter muscle). After a week survival period the animals were sacrificed and the stomachs were collected. Then, the stomach wall was cross-cut into 0.5cm thick sections taken in specified intervals (section I - 1.5cm; section II - 3.5cm; section III - 5.5cm; section IV - 7.5cm) starting from the sphincter. Consecutive microscopic slices prepared from each section were analyzed under fluorescent microscope to count traced neurons. Obtained data were statistically analyzed. The total number of FB-positive perikarya observed within all studied sections significantly decreased from 903.3 ± 130.7 in control to 243.8 ± 67.3 in experimental animals. In healthy pigs 76.1 ± 6.7% of labeled neurons were observed within the section I, 23.53 ± 6.5% in section II and only occasional cells in section III. In experimental animals, as many as 93.8 ± 2.1% of labeled cells were observed within the section I and only 6.2 ± 2.2% in section II, while section III was devoid of such neurons. There were no traced perikarya in section IV observed in both groups of pigs. CONCLUSIONS/SIGNIFICANCE: Obtained results revealed for the first time significant impact of antral ulcerations on intramural descending nerve pathways supplying the pyloric sphincter in pigs, animals of increasing value in biomedical research and great economic importance.

Innervation pattern of polycystic ovaries in the women.

The aim of the present study was to determine the changes in both the distribution pattern and density of nerve fibers containing dopamine ß-hydroxylase (DßH), vesicular acetylcholine transporter (VAChT), neuronal nitric oxide synthase (nNOS), substance P (SP), calcitonin gene related peptide (CGRP), neuropeptide Y (NPY), vasoactive intestinal peptide (VIP), somatostatin (SOM), galanin (GAL) and pituitary adenylate cyclase-activating polypeptide (PACAP) in the human polycystic ovaries. In the polycystic ovaries, when compared to the immunoreactions pattern observed in the control gonads, following changes were revealed: (1) an increase in the number of DßH-, VAChT-, VIP- or GAL-immunoreactive (IR) nerve fibers within the stroma as well as in the number of DßH-IR fibers near primordial follicles and medullar veins and venules; (2) a reduction in the number of nerve fibers containing nNOS, CGRP, SOM, PACAP within the stroma and in the numbers of CGRP-IR fibers around arteries; (3) an appearance of SP- and GAL-IR fibers around medullar and cortical arteries, arterioles, veins and venules, with except of GAL-IR fibers supplying medullar veins; and (4) the lack of nNOS-IR nerve fibers near primordial follicles and VIP-IR nerves around medullar arteries and arterioles. In conclusion, our results suggest that the changes in the innervation pattern of the polycystic ovaries in human may play an important role in the pathogenesis and/or course of this disorder.

Extrinsic primary afferent neurons projecting to the pylorus in the domestic pig--localization and neurochemical characteristics.

The pig, as an omnivorous animal, seems to be especially valuable species in "gastrointestinal" experiments. The importance of the pylorus in the proper functioning of the digestive tract is widely accepted. Although it is commonly known that sensory innervation plays an important role in the regulation of gastric activity and gastrointestinal tissue resistance, there is complete lack of data on the extrinsic afferents projecting to the swine pylorus. The present experiment has been designed to discover the precise localization and neurochemical properties of the primary sensory neurons projecting to the porcine pylorus. Combined retrograde tracing technique and double immunocytochemistry were applied in five piglets. An additional RT-PCR reaction was used to confirm the presence of all investigated neurotransmitters in the studied ganglia. Traced neurons were localized within the bilateral nodose ganglia of the vagus and bilateral dorsal root ganglia spreading from Th4 to L1. Fast Blue-positive afferents expressed immunoreactivity to substance P, calcitonin gene-related peptide, neuronal isoform of nitric oxide synthase, and galanin. In the vagal and spinal ganglia, the percentages of traced neurons immunoreactive to these substances were 54.8, 10.7, 49.6, 7.4 % and 22.2, 75.5, 95.2 %, respectively, and the solitary perikarya were Gal immunoreactive. The presence of all substances studied in the ganglion tissue was confirmed by RT-PCR technique.

Axotomy-induced changes in the chemical coding pattern of colon projecting calbindin-positive neurons in the inferior mesenteric ganglia of the pig.

The present study examines the response of colon-projecting neurons localized in the inferior mesenteric ganglia (IMG) to axotomy in the pig animal model. In all animals (n = 8), a median laparotomy was performed under anesthesia and the retrograde tracer Fast Blue was injected into the descending colon wall. In experimental animals (n = 4), the descending colon was exposed and the bilateral caudal colonic nerves were identified and severed. All animals were euthanized and the inferior mesenteric ganglia were harvested and processed for double-labeling immunofluorescence for calbindin-D28k (CB) in combination with either tyrosine hydroxylase (TH), neuropeptide Y (NPY), somatostatin (SOM), vasoactive intestinal polypeptide (VIP), nitric oxide synthase (NOS), Leu-enkephalin (LENK), substance P, vesicular acetylcholine transporter, or galanin. Immunohistochemistry revealed significant changes in the chemical coding pattern of injured inferior mesenteric ganglion neurons. In control animals, Fast Blue-positive neurons were immunoreactive to TH, NPY, SOM, VIP, NOS, LENK, and CB. In the experimental group, the numbers of TH-, NPY-, and SOM-expressing neurons were reduced, whereas the number of neurons immunoreactive to LENK was increased. Our data indicate that the colon-projecting neurons of the porcine IMG react to the axotomy in a similar, but not an identical manner in a comparison to other species, especially rodents. Further studies are needed to elucidate the detailed factors/mechanisms involved in the response to nerve injury.

Localization and neurochemical characteristics of the extrinsic sympathetic neurons projecting to the pylorus in the domestic pig.

The pylorus, an important part of the digestive tract controlling the flow of chyme between the stomach and the duodenum, is widely innervated by intrinsic and extrinsic nerves. To determine the locations of postganglionic sympathetic perikarya that innervate the pylorus of the domestic pig, a retrograde tracing method with application of Fast Blue tracer was used. All positive neuronal cell bodies (ca. 1750) were found in the celiac-cranial mesenteric ganglion complex (CSMG), however, the coeliac poles of this complex provided the major input to the pylorus. Afterwards, the immunohistochemical staining procedure was applied to determine biologically active substances expressed in the FB-labeled perikarya. Approximately 77% of the FB-positive cell bodies contained tyrosine hydroxylase (TH), 87% dopamine ß-hydroxylase (DßH), 40% neuropeptide Y (NPY), 12% somatostatin (SOM) and 7% galanin (GAL). The presence of all these substances in the ganglion tissue was confirmed by RT-PCR technique. Double immunocytochemistry revealed that all of the TH-positive perikarya contained DßH, about 40% NPY, 12% SOM and 8% GAL. Additionally, all above-cited immunohistochemical markers as well as VIP, PACAP, ChAT, LEU, MET, SP and nNOS were observed within nerve fibers associated with the FB-positive perikarya. Immunocytochemical labeling of the pyloric wall tissue disclosed that TH+, DßH+ and NPY+ nerve fibers innervated ganglia of the myenteric and submucosal plexuses, blood vessels, both muscular layers and the muscularis mucosae; nerve fibers immunoreactive to GAL mostly innervated both muscular layers, while SOM+ nerve fibers were observed within the myenteric plexus. Presented study revealed sources of origin and immunohistochemical characteristics of the sympathetic postganglionic perikarya innervating the porcine pylorus.