Immunohistochemical characteristics and distribution of neurons in the intramural ganglia supplying the urinary bladder in the male pig

2013 ◽  
Vol 16 (4) ◽  
pp. 629-638 ◽  
Author(s):  
Z. Pidsudko
Keyword(s):  
Urinary Bladder ◽  
Large Population ◽  
Double Labelling ◽  
Chemical Coding ◽  
Visceral Peritoneum ◽  
Calcitonin Gene ◽  
Oxide Synthase ◽  
Cryostat Sections ◽  
Labelling Immunofluorescence ◽  
Intramural Ganglia

Abstract This study investigated the distribution and chemical coding of neurons in intramural ganglia of the urinary bladder trigone (UBT-IG) and cervix (UBC-IG) in the male pig using combined retrograde tracing and double-labelling immunohistochemistry. Additionally, immunoblotting was used to confirm the presence of marker enzymes for main populations of autonomic neurons. Retrograde fluorescent tracer Fast Blue (FB) was injected into the wall of both the left and right side of the bladder trigone, cervix and apex during laparotomy performed under thiopental anaesthesia. Twelve μm-thick cryostat sections were processed for double-labelling immunofluorescence with antibodies against tyrosine hydroxylase (TH), dopamine β-hydroxylase (DBH), neuropeptide Y (NPY), somatostatin (SOM), galanin (GAL), vasoactive intestinal polypeptide (VIP), nitric oxide synthase (NOS), calcitonin gene-related peptide (CGRP), substance P (SP) and vesicular acetylcholine transporter (VAChT). UBT-IG and UBC-IG neurons in both parts of the organ formed characteristic clusters (from few to tens of neuronal cells) found under visceral peritoneum or in the outer muscular layer. Immunohistochemistry revealed several subpopulations in UBT-IG and UBC-IG neurons, namely noradrenergic (ca. 76% and 76%), cholinergic (ca. 22% and 20%), non-adrenergic/non-cholinergic nerve cells (ca. 1.5% and 3.8%), NPY- (ca. 66% and 58%), SOM- (ca. 39% and 39 %), VIP- (ca. 5% and 0%) and NOS- immunoreactive (IR) (ca. 1.5% and 3.8%), respectively. Immunoblotting using antibodies to TH and VAChT showed the presence of studied proteins as revealed by the presence of protein bands of the correct molecular weight. This study has revealed a relatively large population of differently coded UBT- and UBC- IG neurons, which constitute an important element of the complex neuroendocrine system involved in the regulation of the male urogenital organs function.

scholarly journals Immunohistochemical characterisation of neurons in the mandibular ganglion and nerve fibres supplying the porcine mandibular gland

2017 ◽  
Vol 61 (No. 7) ◽  
pp. 361-373 ◽  
Author(s):  
M. Klimczuk ◽  
P. Podlasz ◽  
W. Sienkiewicz ◽  
A. Franke-Radowiecka ◽  
A. Dudek ◽  
...  
Keyword(s):  
Mandibular Gland ◽  
Double Labelling ◽  
Nerve Terminals ◽  
Rt Pcr ◽  
Nerve Fibres ◽  
Chemical Coding ◽  
Pcr Product ◽  
Calcitonin Gene ◽  
Oxide Synthase ◽  
Cryostat Sections

The present study was designed to investigate the chemical coding of neurons in the mandibular ganglion (MGn) and nerve fibres supplying the porcine mandibular gland (MGl) with the use of immunofluorescence and RT-PCR. The cryostat sections from MGn and MGl were processed for double-labelling immunohistochemistry using antisera against vesicular acetylcholine transporter (VAChT), choline acetyltransferase (ChAT), dopamine β-hydroxylase (DβH), neuronal nitric oxide synthase (nNOS), vasoactive intestinal polypeptide (VIP), neuropeptide Y (NPY), galanin (GAL), substance P (SP) and calcitonin gene-related peptide (CGRP). The MGl was found to be richly supplied by VAChT-positive nerve fibres that surrounded intra- and interlobular salivary ducts. A large number of VAChT-immunoreactive (VAChT-IR) nerve terminals were also observed around acini. Many periductal and periacinar nerve fibres stained positive for DβH. Immunoreactivity to GAL, NPY or VIP was observed in an intermediate number of nerve terminals which were associated with both salivary ducts and acini. Double-immunostaining revealed that in MGn nearly all neurons stained positive for VAChT/ChAT (98.45 ± 0.59%, mean ± SEM) and nNOS (99.71 ± 0.18%). An intermediate number of the nerve cell bodies displayed immunoreactivity to NPY or VIP (18.67 ± 0.52% and 8.11 ± 0.36%, respectively). Single GAL-IR and CGRP-positive neurons were also observed. RT-PCR revealed the presence of transcripts of ChAT, VAChT, nNOS, NPY, VIP and GAL. For SP and DβH very weak signals were observed. RT-PCR with primers targeting CGRP did not generate any PCR product.

scholarly journals Immunohistochemical characterization of efferent neurons innervating the oviduct in the pig located in the sympathetic chain ganglia

2012 ◽  
Vol 47 (No. 4) ◽  
pp. 85-91
Author(s):  
K. Czaja
Keyword(s):  
Nerve Cells ◽  
Sympathetic Chain ◽  
Double Labelling ◽  
Retrograde Labelling ◽  
Calcitonin Gene ◽  
Ovum Transport ◽  
Efferent Neurons ◽  
Oxide Synthase ◽  
Labelling Immunofluorescence

The present study was aimed at disclosing the pattern(s) of putative co-incidence of tyrosine hydroxylase (TH), dopamine -hydroxylase (DH), neuropeptide Y (NPY), substance P (SP), calcitonin gene-related peptide (CGRP) and nitric oxide synthase (NOS) within the porcine &ldquo;oviductal&rdquo; efferent neurons using combined retrograde tracing and double-labelling immunohistochemistry. The fluorescent retrograde tracer Fast Blue (FB) was injected into the wall of the ampullar (n = 5) and isthmal (n = 5) part of the organ in ten sexually immature female pigs. After a survival period of three weeks sympathetic chain ganglia (SCG) were collected. 10 &micro;m-thick cryostat sections of the ganglia were examined for the presence of FB-positive (FB<sup>+</sup>) nerve cells under the fluorescent microscope. Tracered neurons were processed for double-labelling immunofluorescence according to the method of Wessendorf and Elde. Retrograde labelling revealed a population of &ldquo;oviductal&rdquo; efferent neurons located in the thoracic (T) and lumbar (L) SCG at the level of T<sub>14</sub> to L<sub>5</sub>. Double-labelling immunofluorescence allowed several subpopulations of the studied perikarya to be distinguished. The largest one consisted of TH<sup>+</sup>/DH<sup>+</sup> (immunopositive) nerve cells. The moderate number of FB<sup>+</sup> nerve cells expressed TH/NPY- immunoreactivity (IR). The tracered neurons did not show SP, CGRP and NOS immunoreactivity. Because identically coded nerve fibres have been observed within the wall of the porcine oviduct it can be assumed that TH<sup>+</sup>/DH<sup>+</sup> or TH<sup>+</sup>/NPY<sup>+</sup> neurons are involved in the control the oviductal tonus and ovum transport.

Immunohistochemical properties of motoneurons supplying the trapezius muscle in the rat

2011 ◽  
Vol 14 (2) ◽  
pp. 199-205 ◽  
Author(s):  
A. Dudek ◽  
W. Sienkiewicz ◽  
M. Marczak ◽  
J. Kaleczyc
Keyword(s):  
Trapezius Muscle ◽  
Retrograde Tracing ◽  
Male Rats ◽  
Double Labelling ◽  
Fluorescent Tracer ◽  
Fast Blue ◽  
Tissue Sections ◽  
Calcitonin Gene ◽  
Oxide Synthase ◽  
Labelling Immunofluorescence

Immunohistochemical properties of motoneurons supplying the trapezius muscle in the rat Combined retrograde tracing (using fluorescent tracer Fast blue) and double-labelling immunofluorescence were used to study the distribution and immunohistochemical characteristics of neurons projecting to the trapezius muscle in mature male rats (n=9). As revealed by retrograde tracing, Fast blue-positive (FB+) neurons were located within the ambiguous nucleus and accessory nucleus of the grey matter of the spinal cord. Immunohistochemistry revealed that nearly all the neurons were cholinergic in nature [choline acetyltransferase (ChAT)-positive]. Retrogradely labelled neurons displayed also immunoreactivities to calcitonin gene-related peptide (CGRP; approximately 60% of FB+ neurons), nitric oxide synthase (NOS; 50%), substance P (SP; 35%), Leu5-Enkephalin (LEnk; 10%) and vasoactive intestinal polypeptide (VIP; 5%). The analysis of double-stained tissue sections revealed that all CGRP-, VIP- and LEnk-immunoreactive FB+ perikarya were simultaneously ChAT-positive. The vast majority of the neurons expressing SP- or NOS-immunoreactivity were also cholinergic in nature; however, solitary somata were ChAT-negative. FB+ perikarya were surrounded by numerous varicose nerve fibres (often forming basket-like structures) immunoreactive to LEnk or SP. They were also associated with some CGRP-, NOS- and neuropeptide Y-positive nerve terminals.

scholarly journals Proliferative enteropathy (PPE)-induced changes in the expression of DBH, VAChT and NOS in the neurons of intramural ganglia of the porcine ileum

2008 ◽  
Vol 53 (No. 10) ◽  
pp. 533-542 ◽  
Author(s):  
Z. Pidsudko ◽  
K. Wasowicz ◽  
J. Kaleczyc ◽  
M. Majewski ◽  
M. Lakomy
Keyword(s):  
Enteric Neurons ◽  
Double Labelling ◽  
Lawsonia Intracellularis ◽  
Large White ◽  
Submucous Plexus ◽  
Chemical Coding ◽  
Proliferative Enteropathy ◽  
Induced Changes ◽  
Oxide Synthase ◽  
Intramural Ganglia

As enteric neurons are regarded to be highly adaptive in their response to various pathological states, including inflammation, it appears to be of interest to study the chemical coding of neurons in the intramural ganglia of the ileum wall in the course of porcine proliferative enteropathy (PPE) evoked by <I>Lawsonia intracellularis.</I> The study was performed on 12 juvenile pigs of the Large White Polish breed. The pigs were divided into the control (C, <I>n</I> = 6) group and the group consisting of pigs with clinically diagnosed <I>Lawsonia intracellularis</I> infection (E, <I>n</I> = 6). In E group animals the infection was confirmed with a PCR-based test. All the animals were sacrificed and segments of the ileum being pathologically changed were processed for double-labelling immunofluorescence using antibody against protein gene-product 9.5 (PGP 9.5) combined with antibody for dopamine &beta;-hydroxylase (D&beta;H), vesicular acetylcholine transporter (VAChT) or nitric oxide synthase (NOS). Immunohistochemistry revealed in the inner submucous plexus (ISP) and outer submucous plexus (OSP) an increase of the number of neurons containing D&beta;H and VAChT in the E group. Interestingly, a decrease in the number of D&beta;H- and VAChT-positive neurons in meynteric plexus (MP) ganglia of the E group animals was observed. The most remarkable difference in the chemical coding of enteric neurons between the control and PPE-suffering pigs was a significant increase of the number of NOS-positive nerve cells in the MP and OSP of the infected animals. The present results show that acetylcholine, catecholamines and NO may be involved in the regulation of functions of the porcine enteric nerve pathways not only under physiological, but also pathological conditions.

Distribution and chemical coding patterns of cocaine- and amphetamine-regulated transcript-like immunoreactive (CART-LI) neurons in the enteric nervous system of the porcine stomach cardia

2015 ◽  
Vol 18 (3) ◽  
pp. 515-522 ◽  
Author(s):  
W. Rękawek ◽  
P. Sobiech ◽  
S. Gonkowski ◽  
K. Żarczyńska ◽  
A. Snarska ◽  
...  
Keyword(s):  
Nervous System ◽  
Enteric Nervous System ◽  
Neuronal Nitric Oxide Synthase ◽  
Nerve Fibers ◽  
Vesicular Acetylcholine Transporter ◽  
Double Labelling ◽  
Dense Network ◽  
Chemical Coding ◽  
Oxide Synthase ◽  
Labelling Immunofluorescence

Abstract The aim of this study was to determine the presence of cocaine- and amphetamine-regulated transcript-like immunoreactive (CART-LI) neurons and co-localisation of CART with vesicular acetylcholine transporter (VAChT), neuronal nitric oxide synthase (n-NOS), vasoactive intestinal polypeptide (VIP), substance P (SP) and leu-enkephalin (LENK) in the enteric nervous system of the porcine gastric cardia by using a double-labelling immunofluorescence technique. CART-LI neurons were observed in the myenteric plexus (18.2±2.6%). A dense network of CART-LI nerve fibers was mainly observed in the muscular layer. CART showed co-localization mainly with VAChT, n-NOS, VIP and to a lesser degree with LENK and SP. Distribution of CART and its co-localization with other neurotransmitters suggest that this peptide plays an important role in gastric motility in the pig.

Distribution pattern and chemical coding of neurons of the sympathetic chain ganglia supplying the descending colon in the pig

2010 ◽  
Vol 58 (2) ◽  
pp. 189-198 ◽  
Author(s):  
Cezary Skobowiat ◽  
Jarosław Calka ◽  
Krzysztof Wasowicz ◽  
Mariusz Majewski
Keyword(s):  
Great Majority ◽  
Retrograde Tracing ◽  
Sympathetic Chain ◽  
Double Labelling ◽  
Chemical Coding ◽  
Pituitary Adenylate Cyclase ◽  
Descending Colon ◽  
Oxide Synthase ◽  
Ventral Area ◽  
Labelling Immunofluorescence

Sympathetic chain ganglia (SChG) neurons projecting to the descending colon of the pig were studied by means of retrograde tracing (Fast Blue, FB) and double-labelling immunofluorescence methods. FB was injected into the gut wall and after three weeks survival time the animals were transcardially perfused with paraformaldehyde and the bilateral sympathetic trunks were collected. The FBpositive neurons were localised only in the lumbar (L 1 –L 5 ) ganglia of the sympathetic trunk and appeared either as small (30–50 μm in diameter) round-shaped perikarya forming clusters localised in caudal-ventral area or, rarely, as bigger (50–80 μm) and dispersed solitary irregular perikarya. Immunohistochemical staining revealed the catecholaminergic (tyrosine hydroxylase-/dopamine β-hydroxylase-immunoreactive) character of the great majority of FB-positive neurons which preferentially co-expressed neuropeptide Y. In addition, none of the FB-positive perikarya was immunopositive to galanin, somatostatin, choline acetyltransferase, vasoactive intestinal peptide, pituitary adenylate cyclase-activating peptide, leu 5 -enkephalin, nitric oxide synthase, substance P and calcitonin-generelated peptide.

Characterisation of cocaine- and amphetamine- regulated transcript-like immunoreactive (CART-LI) enteric neurons in the porcine small intestine

2012 ◽  
Vol 60 (3) ◽  
pp. 371-381 ◽  
Author(s):  
Joanna Wojtkiewicz ◽  
Sławomir Gonkowski ◽  
Marek Bladowski ◽  
Mariusz Majewski
Keyword(s):  
Nitric Oxide ◽  
Small Intestine ◽  
The Other ◽  
Enteric Neurons ◽  
Double Labelling ◽  
Immunofluorescence Technique ◽  
Submucous Plexus ◽  
Oxide Synthase ◽  
First Time ◽  
Labelling Immunofluorescence

The aim of this study was to investigate the distribution and the number of cocaine- and amphetamine-regulated transcript-like immunoreactive (CART-LI) neurons and the co-localisation of CART with substance P (SP), somatostatin (SOM), nitric oxide synthase (NOS) and vasoactive intestinal polypeptide (VIP) within the enteric nervous system (ENS) in the porcine small intestine. Accordingly, the myenteric plexus (MP), outer submucous plexus (OSP) and inner submucous plexus (ISP) of the small intestine (duodenum, jejunum and ileum) were studied by double-labelling immunofluorescence technique. CART-LI neurons were observed in all gut fragments and all types of intramural plexuses studied and amounted from 0.2 ± 0.1% in the ISP of ileum to 22.4 ± 2.4% in the MP of this segment. The co-localisation of CART and NOS or/and VIP was observed depending on the segment of the gut and the complexity of the intramural plexus. On the other hand, during this study the co-localisation of CART and SOM or/and SP was not observed. The present study, for the first time, presents a detailed description of the CART distribution pattern and co-localisation with other neuromodulators within the ENS of the porcine small intestine.

Distribution and chemical coding of sympathetic neurons in the caudal mesenteric ganglion projecting to the ovary in sexually mature gilts

2010 ◽  
Vol 58 (3) ◽  
pp. 389-403 ◽  
Author(s):  
Marlena Koszykowska ◽  
Anna Kozłowska ◽  
Joanna Wojtkiewicz ◽  
Cezary Skobowiat ◽  
Mariusz Majewski ◽  
...  
Keyword(s):  
Sympathetic Neurons ◽  
Physiological Role ◽  
Double Labelling ◽  
Nerve Fibres ◽  
Neuronal Tracing ◽  
Chemical Coding ◽  
Mesenteric Ganglion ◽  
Caudal Mesenteric Ganglion ◽  
Labelling Immunofluorescence ◽  
Sexually Mature

The distribution and co-localisation patterns of dopamine-β-hydroxylase (DβH), neuropeptide Y (NPY), somatostatin (SOM) and galanin (GAL) were investigated by use of retrograde neuronal tracing and double-labelling immunofluorescence techniques in the caudal mesenteric ganglion (CaMG) neurons supplying the ovary of adult pigs. The existence and density of nerve fibres that are immunoreactive (IR) for the above-mentioned neuroactive substances were also evaluated. Injections of a fluorescent tracer (Fast Blue; FB) into the ovaries revealed the presence of small- (76.38%) and large-sized (23.62%) FB-positive postganglionic neurons in the CaMG. Noradrenergic FB-positive cells were simultaneously NPY- (43.38%), SOM- (18.77%) and GAL- (18.31%) IR. Of the examined FB-positive neurons, 53.49% were DβH-IR but NPY-immunonegative (IN), 79.06% were DβH-IR but SOM-IN, and 77.16% were DβH-IR but GAL-IN. Small- or large-sized subsets of traced neurons were supplied by only one or a few nerve fibres, exhibiting DβH-, NPY-, SOM- and/or GAL-IR. Our data show the specific morphological as well as immunochemical structural organisation of the sympathetic neurons in the CaMG in adult gilts. The occurrence of an abundant population of noradrenergic perikarya in the CaMG may suggest their important physiological role in the regulation of gonadal function(s) in these animals.

scholarly journals Neurochemical Plasticity of nNOS-, VIP- and CART-Immunoreactive Neurons Following Prolonged Acetylsalicylic Acid Supplementation in the Porcine Jejunum

2020 ◽  
Vol 21 (6) ◽  
pp. 2157
Author(s):  
Dominika Rząp ◽  
Marta Czajkowska ◽  
Jarosław Całka
Keyword(s):  
Nitric Oxide ◽  
Nervous System ◽  
Acetylsalicylic Acid ◽  
Neuronal Plasticity ◽  
Adaptive Response ◽  
Neuronal Nitric Oxide Synthase ◽  
Double Labelling ◽  
Neurochemical Plasticity ◽  
Oxide Synthase ◽  
Labelling Immunofluorescence

Aspirin, also known as acetylsalicylic acid (ASA), is a commonly used anti-inflammatory drug that has analgesic and antipyretic properties. The side effects are well known, however, knowledge concerning its influence on gastric and intestinal innervation is limited. The enteric nervous system (ENS) innervates the whole gastrointestinal tract (GIT) and is comprised of more than one hundred million neurons. The capacity of neurons to adapt to microenvironmental influences, termed as an enteric neuronal plasticity, is an essential adaptive response to various pathological stimuli. Therefore, the goal of the present study was to determine the influence of prolonged ASA supplementation on the immunolocalization of neuronal nitric oxide synthase (nNOS), vasoactive intestinal peptide (VIP) and cocaine- and amphetamine- regulated transcript peptide (CART) in the porcine jejunum. The experiment was performed on 8 Pietrain × Duroc immature gilts. Using routine double-labelling immunofluorescence, we revealed that the ENS nerve cells underwent adaptive changes in response to the induced inflammation, which was manifested by upregulated or downregulated expression of the studied neurotransmitters. Our results suggest the participation of nNOS, VIP and CART in the development of inflammation and may form the basis for further neuro-gastroenterological research.

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