Immunohistochemical detection of neuronal plexuses and nerve cells within the upper urinary tract of pigs (original) (raw)
Immunohistochemical demonstration of nerves and nerve cells in human and porcine ureters
Annals of Anatomy - Anatomischer Anzeiger, 1993
There are divergent opinions about the origin of ureteral motility. It is either a myogenic or a neutral phenomenon. Furthermore, the existence of nerve cells in the ureter is also a controversial question. In the present study we applied immunohistochemical methods to human and pocine ureters in an attempt to clarify the mattem. As neuronal markers we used anti-PGP 9.5 and anti-NSE, and as a glial marker anti-S-100. In the wohle mount preparations of pig ureter we observed two neuronal plexuses on both sides of the tunica muscularis. The inner plexus consisted of both nerve bundles and nerve cells, whereas the outer one did not contain any nerve cells. In the human ureter we found a ganglion with nerve cells beneath the tunica muscularis and the tunica adventitia.
Transitory inner medullary nerve terminals in the cat kidney
Neuroscience Letters, 1990
An indirect immunohistochemical method was used to visualize nerves immunoreactive for tyrosine hydroxylase (THI) and dopamine-fl-hydroxylase (DBHI) in kidney sections of cats 6 weeks and 2 and 3 months of age. THI and DBHI nerve terminals innervate the renal pelvis, interlobar veins and arterial tree including medullary vascular bundles of cats of each age studied. In kidneys of 6-week-old cats, THI and DBHI axons form elaborate plexuses that are distributed throughout much of the inner medulla, whereas some medullary axons appear to degenerate at 2 months and no inner medullary plexuses were visualized in 3-month-old cats. Transitory inner medullary nerves in the cat kidney may influence cellular development and play a role in salt and water balance.
European Journal of Histochemistry, 2013
The urinary bladder trigone (UBT) is a limited area through which the majority of vessels and nerve fibers penetrate into the urinary bladder and where nerve fibers and intramural neurons are more concentrated. We localized the extramural post-ganglionic autonomic neurons supplying the porcine UBT by means of retrograde tracing (Fast Blue, FB). Moreover, we investigated the phenotype of sympathetic trunk ganglia (STG) and caudal mesenteric ganglia (CMG) neurons positive to FB (FB+) by coupling retrograde tracing and double-labeling immunofluorescence methods. A mean number of 1845.1±259.3 FB+ neurons were localized bilaterally in the L1-S3 STG, which appeared as small pericarya (465.6±82.7 µm 2) mainly localized along an edge of the ganglion. A large number (4287.5±1450.6) of small (476.1±103.9 µm 2) FB+ neurons were localized mainly along a border of both CMG. The largest number (4793.3±1990.8) of FB+ neurons was observed in the pelvic plexus (PP), where labeled neurons were often clustered within different microganglia and had smaller soma cross-sectional area (374.9±85.4 µm 2). STG and CMG FB+ neurons were immunoreactive (IR) for tyrosine hydroxylase (TH) (66±10.1% and 52.7±8.2%, respectively), dopamine beta-hydroxylase (DbH) (62±6.2% and 52±6.2%, respectively), neuropeptide Y (NPY) (59±8.2% and 65.8±7.3%, respectively), calcitonin-gene-related peptide (CGRP) (24.1±3.3% and 22.1±3.3%, respectively), substance P (SP) (21.6±2.4% and 37.7±7.5%, respectively), vasoactive intestinal polypeptide (VIP) (18.9±2.3% and 35.4±4.4%, respectively), neuronal nitric oxide synthase (nNOS) (15.3±2% and 32.9±7.7%, respectively), vesicular acetylcholine transporter (VAChT) (15±2% and 34.7±4.5%, respectively), leuenkephalin (LENK) (14.3±7.1% and 25.9±8.9%, respectively), and somatostatin (SOM) (12.4±3% and 31.8±7.3%, respectively). UBT-projecting neurons were also surrounded by VAChT-, CGRP-, LENK-, and nNOS-IR fibers. The possible role of these neurons and fibers in the neural pathways of the UBT is discussed. We have chosen the domestic pig as experimental animal because it has been regarded as a more suitable model for studying human lower urinary tract innervation than rodents or carnivores. 15-18 Materials and Methods All the procedures described below were carried out in accordance with the European Communities Council Directive of 24 November 1986 (86/609/EEC) and the Italian legislation regarding experimental animals, after approval by the Scientific Ethics Committee for Experiments on Animals of the University of Parma (Prot. Rif. 68/09). All possible efforts were made to minimize the number of animals used and their suffering. Four intact male crossbreed (Large white x Landrace x Duroc) pigs (aged 3 months, mean body weight: 34±3 Kg, range 30-38 Kg) were maintained on a diet and water ad libitum for 1 week before the experiment. For 24 h prior to surgery, the animals were not given any food and preventive antibiotic therapy with Ceftiofur (Naxcel 5 mg/Kg i.m., Pfizer, Sandwich Kent, UK) was administered. The animals were sedated by intramuscular injection of azaperone (4-10 mg/10 Kg,
Qualitative and quantitative morphology of renal nerves in C57BL/6J mice
2002
The detailed morphology of the renal nerves in mice has not been reported previously. The aims of this study were to describe the general morphology of the extrinsic renal nerve in C57BL/6 mice, and determine its morphometric parameters. The major renal nerve innervating the left kidney was isolated in five mice. Thin sections of the nerve segments were then examined by transmission electron microscopy. The renal nerve averaged 35.4 Ϯ 3.6 (S.E.M.) m in diameter and 741 Ϯ 104 m in area. The renal nerve contained an average of 830 Ϯ 169 unmyelinated fibers and only 4.6 Ϯ 1.7 myelinated fibers. The axon diameter of myelinated and unmyelinated fibers averaged 2.2 Ϯ 0.3 m and 0.76 Ϯ 0.02 m, respectively. The diameter of the unmyelinated fibers ranged from 0.3 to 2.0 m, and the distribution histogram was unimodal. The majority of fibers (85%) had diameters of 0.6-1.0 m. These results are similar to those obtained for renal nerves of rats with respect to the predominance of unmyelinated fibers. However, the diameter of unmyelinated fibers is larger in rats and the distribution histogram of rat unmyelinated fibers is bimodal, in contrast to the unimodal distribution in mice. The morphological description of the renal nerves in mice provides baseline data for further investigations of the structural basis of altered autonomic reflexes. The results will be useful in analyses of genes that influence the development and structure of sympathetic and sensory innervation of the kidney in genetically manipulated mice. Anat Rec 268:399-404, 2002.
Neurourology and Urodynamics, 2008
Aims: The morphology and functional importance of the autonomic nervous system in the upper urinary tract is still not completely understood. Previous histological studies investigating the innervation of the urinary tract have mainly used conventional sections in which the three-dimensional structure of the intramural innervation is difficult to achieve. In contrast, the whole-mount preparation technique is a suitable method for visualizing the distribution of the mesh-like neuronal networks within the urinary tract. Methods: The distribution and regional variation of neurofilament (NF), tyrosine hydroxylase (TH), choline acetyltransferase (ChAT), and substance P-immunoreactive (SP-IR) neurons, as well as acetylcholinesterase (AChE) and nicotinamide adenine dinucleotide phosphate-diaphorase (NADPH-d)-positive neurons were investigated using whole-mount preparations of the guinea pig upper urinary tract. Results: Two distinct nervous plexuses were detected within the muscle layers containing NF, TH, ChAT, and SP-IR nerves. AChE-positive nerves were seen in all layers. Only moderate NADPH-d-positive innervation was found. Renal pelvis, upper and lower part of the ureter showed an overall increased innervation compared to the middle portion of the ureter. Ganglia were found at the pelviureteric border displaying NF and TH immunoreactivity. Conclusion: The whole-mount preparation technique provides an elegant method for assessing the three-dimensional architecture of ureteral innervation. The guinea pig upper urinary tract is richly supplied with adrenergic, cholinergic, nitrergic, and sensory nerves which suggest that the autonomous nervous system plays an important role in controlling ureteral motility and blood flow.
Polish Journal of Veterinary Sciences, 2023
Combined retrograde tracing and double-labelling immunofluorescence were used to investigate the distribution and chemical coding of neurons in testicular (TG) and aorticoerenal (ARG) ganglia supplying the urinary bladder trigone (UBT) in juvenile male pigs (n=4, 12 kg. of body weight). Retrograde fluorescent tracer Fast Blue (FB) was injected into the wall of the bladder trigone under pentobarbital anesthesia. After three weeks all the pigs were deeply anesthetized and transcardially perfused with 4% buffered paraformaldehyde. TG and ARG, were collected and processed for double-labelling immunofluorescence. The expression of tyrosine hydroxylase (TH) or dopamine beta-hydroxylase (DBH), neuropeptide Y (NPY), somatostatin (SOM), galanin (GAL), nitric oxide synthase (NOS) and vesicular acetylcholine transporter (VAChT) were investigated. The cryostat sections were examined with a Zeiss LSM 710 confocal microscope equipped with adequate filter blocks. The TG and ARG were found to contain many FB-positive neurons projecting to the UBT (UBT-PN). The UBT-PN were distributed in both TG and ARG. The majority of them were found in the right ganglia, mostly in TG. Immunohistochemistry disclosed that the vast majority of UBT-PN were noradrenergic (TH-and/or DBH-positive). Many noradrenergic neurons contained also immunoreactivity to NPY, SOM or GAL. Most of the UBT-PN were supplied with VAChT-, or NOS-IR (immunoreactive) varicose nerve fibres. This study has revealed a relatively large population of differently coded prevertebral neurons projecting to the porcine urinary bladder. As judged from their neurochemical organization these nerve cells constitute an important element of the complex neuro-endocrine system involved in the regulation of the porcine urogenital organ function.
Neuro-tracing approach to study kidney innervation: a technical note
Kidney research and clinical practice, 2017
Neuro-tracing approach is a great option to study innervation of the visceral organs including the kidneys. Important factors contributing to the success of this technique include the choice of a neuro-tracer, and delivery methods to result in successful labeling of peripheral sensory and motor ganglia. The neuro-tracer is usually applied directly to the kidney accessed via a surgical opening of the abdominal wall under deep anesthesia. A series of local microinjections of the dye are performed followed by a wound closure, and recovery period from the surgery. An extra care should be taken to prevent neuro-tracer spillage and accidental labeling of the surrounding organs during injections of the dye. Retrograde neuro-tracers like Fast Blue do not cross synapses, therefore, only neuronal bodies located within dorsal root ganglion neurons and major peripheral ganglia will be labeled by this approach. Retrogradely labeled peripheral neurons could be freshly isolated and dissociated for...
Innervation of congenitally hydronephrotic and normal porcine upper urinary tract
BJU International, 2002
Objective To investigate the intrinsic innervation of the upper urinary tract in congenitally hydronephrotic and normal Goettingen minipigs, using the wholemount preparation technique. Materials and methods Whole-mount preparations of hydronephrotic (two with bilateral ectopic ureters, one with left distal ureteric stenosis) and normal (three) porcine upper urinary tracts were examined by immunohistochemistry with tyrosine hydroxylase (TH) and neurofilament and by histochemical staining with NADPH-diaphorase and acetylcholinesterase. Staining results were evaluated using normal bright-field and confocal laser scanning microscopy. Results Neurofilament-, TH-immunoreactive and acetylcholinesterase-positive nerve fibres and neuronal networks were identified in the adventitial, muscle and subepithelial layers of the whole upper urinary tract. An NADPH-diaphorase-positive network was expressed in the subepithelial layer and less densely in the muscle layer. The general distribution of the identified neuronal networks was similar in hydronephrotic and normal upper urinary tracts, but the density of these neuronal networks was less in the former. The most striking observation was the absence or marked reduction of neuronal networks in the stenotic part of the ureter in the pig with left distal ureteric stenosis. Conclusion Whole-mount preparations provide a method for assessing the three-dimensional topography of neuronal networks in the different layers of the porcine upper urinary tract. Although the macroscopic differences between the hydronephrotic and normal porcine upper urinary tracts were striking, changes in the innervation pattern were less obvious, except in distal ureteric stenosis.
Journal of Comparative Neurology, 2012
Porcine lumbosacral dorsal root ganglion (DRG) neurons were neurochemically characterized by using six neuronal markers: calcitonin gene‐related peptide (CGRP), substance P (SP), neuronal nitric oxide synthase (nNOS), neurofilament 200kDa (NF200), transient receptor potential vanilloid 1 (TRPV1), and isolectin B4 (IB4) from Griffonia simplicifolia. In addition, the phenotype and cross‐sectional area of DRG neurons innervating the urinary bladder trigone (UBT) were evaluated by coupling retrograde tracer technique and immunohistochemistry. Lumbar and sacral DRG neuronal subpopulations were immunoreactive (IR) for CGRP (30 ± 3% and 29 ± 3%, respectively), SP (26 ± 8% and 27 ± 12%, respectively), nNOS (21 ± 4% and 26 ± 7%, respectively), NF200 (75 ± 14% and 81 ± 7%, respectively), and TRPV1 (48 ± 13% and 43 ± 6%, respectively), and labeled for IB4 (56 ± 6% and 43 ± 10%, respectively). UBT sensory neurons, which were distributed from L2 to Ca1 DRG, had a segmental localization, showing ...