RESUMO
The localization of neuropeptide Y and somatostatin in the retina of six prenatal human specimens was determined by the utilization of light microscopic immunoperoxidase and immunofluorescence methods. The age of the prenatal fetuses ranged from 15 to 40 weeks of gestation. At 15 weeks, round or pear-shaped neuropeptide Y-immunopositive cells were observed in the inner nuclear layer and somatostatin immunoreactivity was only detected in the ganglion cell layer. Positive neuropeptide Y ganglion cells were observed by 17 weeks of gestation and by 28 weeks neuropeptide Y-immunopositive cells were demonstrated in the ganglion cell layer and inner nuclear layer. At 26-28 weeks, neuropeptide Y-immunopositive cells exhibited approximately the same shape as at 15 weeks of gestation, but the appearance of one or two processes was detected extending from the somata. By 38-40 weeks of gestation, neuropeptide Y-immunopositive cells were detected in the ganglion cell layer, inner nuclear layer and an immunopositive fibrous configuration in the inner plexiform layer. During this same period (38-40 weeks), somatostatin-positive cells were located in the ganglion cell layer, inner nuclear layer and the inner segments layers. Due to the difficulty of obtaining fetal materials, the exact time of initiation in the expression of neuropeptide Y and somatostatin is presently hard to delineate; however, it is safe to state that the peptides appear early in development.
Assuntos
Proteínas do Olho/análise , Neuropeptídeo Y/análise , Retina/química , Somatostatina/análise , Idade Gestacional , Humanos , Retina/citologia , Retina/embriologia , Células Ganglionares da Retina/química , Células Ganglionares da Retina/ultraestrutura , Especificidade da EspécieRESUMO
The motor innervation of the smooth muscle coat of the human vas deferens is predominantly noradrenergic in type while a less dense and differently distributed presumptive cholinergic innervation is also in evidence, although the precise role of the latter is undetermined. Immunohistochemical studies have confirmed the presence of catecholamine-synthesizing enzymes tyrosine hydroxylase (TH) and dopamine beta hydroxylase (DbetaH) in the majority of fine, varicose intramuscular nerves, about two-thirds of which also contain neuropeptide Y (NPY). Minor populations of noradrenergic nerves contain enkephalin (ENK), galanin (GAL), somatostatin (SOM), or nitric oxide synthase (NOS). The presumptive cholinergic intramuscular nerves contain vasoactive intestinal polypeptide (VIP) and NPY. The subepithelial nerves of the vas deferens are assumed to have a secretomotor function and are rich in acetylcholinesterase and NPY, many also containing either VIP or NOS. The muscle coat of the human vas deferens is poorly differentiated until after birth, the intramuscular nerves in the fetus being relatively thick and non-varicose. Development of a subepithelial nerve plexus lags behind that in the muscle coat but its density in the neonatal vas deferens resembles that seen in the adult. Observations on specimens of human vas deferens obtained at vasovasostomy carried out 1 to 15 years after vasectomy have shown a marked reduction in the density of noradrenergic nerves in the muscle coat of the testicular portion while that in the urethral portion remains unaltered. Furthermore, the subepithelial secretomotor nerves degenerate in the testicular portion. These long-term changes in the pattern of innervation of the vas deferens consequent upon vasectomy may have profound effects upon the outcome of vasovasostomy with respect to subsequent sperm maturation, transport, and viability.
Assuntos
Sistema Nervoso Autônomo/fisiologia , Ducto Deferente/anatomia & histologia , Ducto Deferente/inervação , Sistema Nervoso Autônomo/química , Monóxido de Carbono/fisiologia , Humanos , Imuno-Histoquímica , Masculino , Músculo Liso/inervação , Neuropeptídeos/análise , Neuropeptídeos/fisiologia , Óxido Nítrico/fisiologia , Purinas/metabolismo , Ducto Deferente/embriologia , Ducto Deferente/crescimento & desenvolvimento , VasectomiaRESUMO
Autonomic ganglia of the human pelvic plexus contain sympathetic and parasympathetic neurons which innervate the internal reproductive organs and the lower urinary tract while the urinary bladder also receives innervation from small intramural ganglia embedded in the detrusor muscle. Previous studies have used the immunocytochemical demonstration of tyrosine hydroxylase (TH), either alone or in combination with dopamine beta-hydroxylase, to identify noradrenergic neurons in these ganglia. However until recently a reliable marker for cholinergic neurons in the human autonomic nervous system was not available since antibodies to choline acetyltransferase do not react in this tissue. The present immunohistochemical study has used an antibody to human vesicular acetylcholine transporter (VAChT) to identify cholinergic neurons in the pelvic plexus and intramural bladder ganglia in a series of specimens from human male neonates and children. Immunostaining for TH was also carried out on the same sections and the results showed that while the vast majority of pelvic ganglion neurons were either cholinergic or noradrenergic (as seen by the presence of VAChT or TH respectively), approximately 50% of the neurons in the intramural ganglia were labeled with both immunomarkers. The presence of TH in cholinergic neurons may be due to the immaturity of the tissues examined since previous data on intramural bladder ganglia in the adult have shown that a much smaller proportion of the neurons contain TH than was observed in the present study. It is concluded that the presence of TH alone cannot be regarded as a specific marker for noradrenergic neurons in the genitourinary system of the human neonate and child.
Assuntos
Proteínas de Transporte/metabolismo , Proteínas de Membrana Transportadoras , Neurônios/metabolismo , Tirosina 3-Mono-Oxigenase/metabolismo , Bexiga Urinária/inervação , Proteínas de Transporte Vesicular , Criança , Pré-Escolar , Gânglios/citologia , Gânglios/metabolismo , Humanos , Imuno-Histoquímica/métodos , Lactente , Recém-Nascido , Masculino , Coloração e Rotulagem , Proteínas Vesiculares de Transporte de AcetilcolinaRESUMO
The infiltration of T lymphocytes and macrophages was examined in the retinae of traumatized and non-traumatized contra-lateral eyes of mice. Traumatization was produced by channeling a small hole in the limbic region of a single eye in each animal while it was under Nembutal anesthesia and a small piece of iron wire was inserted into the eye. After the T lymphocytes and macrophages were made apparent by immuno-histochemistry, they were counted on sectioned strips of the peripheral retinae 250 micrometer in length in both traumatized and non-traumatized eyes. During the first two weeks after the induced disturbance, both the retinae of the traumatized and non-traumatized eyes showed infiltration of T lymphocytes and macrophages. Although this increase in T lymphocyte infiltration was observed in both damaged eye and the undamaged one, the former displayed an enhanced increase. The contrary was noticed for macrophages infiltrating the undamaged eye. A bilateral response was therefore evident through the cellular infiltration in the traumatized and un-traumatized eyes. Morphometrical observations also revealed nuclear size changes in the retina during this acute phase of traumatization.
Assuntos
Ferimentos Oculares Penetrantes/patologia , Macrófagos/patologia , Oftalmia Simpática/patologia , Retina/patologia , Subpopulações de Linfócitos T/patologia , Doença Aguda , Animais , Núcleo Celular/ultraestrutura , Reação a Corpo Estranho/patologia , Inflamação , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Células Ganglionares da Retina/patologia , Corpo Vítreo/patologiaRESUMO
OBJECTIVE: To determine the spatial and temporal distribution of nitric oxide synthase (NOS) in the urogenital organs of a series of human male fetuses, using an immunohistochemical technique. MATERIAL AND METHODS: Thirteen pre-natal specimens ranging in gestational age from 13 to 30 weeks were acquired following abortion or miscarriage. The distribution of NOS, which catalyses the production of nitric oxide (NO), was revealed using an indirect immunolabelling technique and compared with the overall innervation of each specimen visualized using the general nerve-marker protein gene product 9.5 (PGP). RESULTS: At 13 weeks of gestation the majority of nerves supplying the developing prostate gland expressed NOS while similar nerves formed a very minor proportion of the total innervation to the urinary bladder and intramural ureters. With increasing gestational age, NOS-containing nerves became more numerous in the lower urinary tract, the majority occurring at the bladder neck and around the prostatic urethra. In contrast, NOS-containing nerves were not detected in the muscle coat of the vas deferens and seminal vesicle until 23 weeks of gestation and at 30 weeks still only formed a small proportion of the intramuscular nerves. From 23 weeks onwards NOS-containing nerves were present occasionally in the dense subepithelial nerve plexuses which developed in the bladder, prostate, vas deferens and seminal vesicle. Also from 23 weeks onwards, many of the epithelial cells lining the vas deferens, seminal vesicle and ejaculatory ducts showed immunoreactivity to NOS but no immunoreactivity was observed in the epithelial lining of the urinary bladder and the intramural ureters. CONCLUSION: Based on the comparative density of NOS-containing nerves and the difference in their temporal development among the various urogenital organs it is apparent that NO plays an increasingly important role in the autonomic control of the lower urinary tract during fetal development but that its involvement in the functional control of the vas deferens and seminal vesicle is relatively minor before birth.
Assuntos
Genitália Masculina/inervação , Óxido Nítrico Sintase/metabolismo , Sistema Urinário/inervação , Vias Autônomas/embriologia , Vias Autônomas/enzimologia , Técnica Direta de Fluorescência para Anticorpo , Genitália Masculina/embriologia , Genitália Masculina/enzimologia , Humanos , Imuno-Histoquímica , Masculino , Próstata/embriologia , Próstata/enzimologia , Próstata/inervação , Glândulas Seminais/embriologia , Glândulas Seminais/enzimologia , Glândulas Seminais/inervação , Ureter/embriologia , Ureter/enzimologia , Ureter/inervação , Bexiga Urinária/embriologia , Bexiga Urinária/enzimologia , Bexiga Urinária/inervação , Sistema Urinário/embriologia , Sistema Urinário/enzimologia , Ducto Deferente/embriologia , Ducto Deferente/enzimologia , Ducto Deferente/inervaçãoRESUMO
Immunohistochemical methods were used to study the developing peptidergic innervation of the human fetal prostate gland in a series of specimens ranging in gestational age from 13 to 30 wk. The overall innervation of each specimen was visualised using protein gene product 9.5 (PGP), a general nerve marker. The onset and development of specific neuropeptide-containing subpopulations were investigated using antisera to neuropeptide Y (NPY), vasoactive intestinal peptide (VIP), substance P (SP), calcitonin gene-related peptide (CGRP), bombesin (BOM), somatostatin (SOM), leu-enkephalin (l-ENK) and met-enkephalin (m-ENK). In addition the occurrence and distribution of presumptive noradrenergic nerves was studied using antisera to dopamine-beta-hydroxylase (D beta H) and tyrosine hydroxylase (TH). At 13 wk numerous branching PGP-immunoreactive (-IR) nerves were observed in the capsule of the developing prostate gland and surrounding the preprostatic urethra but the remainder of the gland was devoid of nerves. The majority of nerves in the capsule contained D beta H and TH and were presumed to be noradrenergic in type while other nerves (in decreasing numbers) contained NPY, l-ENK, SP and CGRP. Nerves associated with the preprostatic urethra did not contain any of the neuropeptides under investigation. At 17 wk the density of nerves in the capsule had increased and occasional m-ENK-, VIP- and BOM-IR nerve fibres were also observed. In addition PGP, D beta H-, TH-, NPY- and l-ENK-IR nerves occurred in association with smooth muscle bundles which at 17 wk were present in the outer part of the gland. Occasional PGP-IR nerves were also present at the base of the epithelium forming some of the prostatic glands. At 23 wk some of the subepithelial nerves showed immunoreactivity for NPY, VIP or l-ENK. At 26 wk smooth muscle bundles occurred throughout the gland and were richly innervated by PGP, D beta H and TH-IR nerves while a less dense plexus was formed by NPY- and l-ENK-IR nerves together with a few m-ENK-IR nerves. Occasional smooth muscle-associated varicose nerve fibres showed immunoreactivity for SP, CGRP, VIP or BOM although the majority of these types of nerve formed perivascular plexuses. Also at 26 wk numerous varicose nerve fibres were observed in association with the prostatic acini, the majority of such nerves containing NPY with a few showing immunoreactivity to VIP, l-ENK, SP or CGRP.(ABSTRACT TRUNCATED AT 400 WORDS)
Assuntos
Desenvolvimento Embrionário e Fetal , Neuropeptídeos/análise , Próstata/embriologia , Próstata/inervação , Bombesina/análise , Peptídeo Relacionado com Gene de Calcitonina/análise , Dopamina beta-Hidroxilase/análise , Encefalina Leucina/análise , Encefalina Metionina/análise , Epitélio/química , Feminino , Humanos , Imuno-Histoquímica , Masculino , Neuropeptídeo Y/análise , Gravidez , Segundo Trimestre da Gravidez , Próstata/química , Somatostatina/análise , Substância P/análise , Tioléster Hidrolases/análise , Tirosina 3-Mono-Oxigenase/análise , Ubiquitina Tiolesterase , Uretra/química , Peptídeo Intestinal Vasoativo/análiseRESUMO
Triple label immunohistochemistry was used to study the coexistence of the catecholamine-synthesising enzymes dopamine beta-hydroxylase (DBH) and tyrosine hydroxylase (TH) and several neuropeptides including neuropeptide Y (NPY), vasoactive intestinal polypeptide (VIP), substance P (SP), calcitonin gene-related peptide (CGRP), somatostatin (SOM) and galanin (GAL) as well as nitric oxide synthase (NOS) in developing pelvic paraganglion cells in a series of human male fetal, neonatal and infant specimens ranging in age from 13 wk of gestation to 3 y postnatal. 13-20 wk old fetal specimens possessed large clusters of paraganglion cells lying lateral to the urinary bladder and prostate gland which were intensely DBH-immunoreactive (-IR) but lacked TH, NOS and the neuropeptides investigated. With increasing fetal age small clusters of paraganglion cells were observed in the muscle coat of the urinary bladder. At 23 wk of gestation occasional paraganglion cells were NOS or NPY-IR while at 26 wk of gestation the majority of paraganglion cells were TH-IR and a few were SOM or GAL-IR. Some postnatal paraganglia within the bladder musculature contained cells which were all VIP, SP or CGRP-IR while others displayed coexistence of NOS and NPY, SP and CGRP, or NPY and VIP. The presence of NOS in certain paraganglion cells indicates their capacity to generate nitric oxide (NO). These results show that human paraganglion cells develop different phenotypes possibly dependent upon their location within the bladder wall. A delicate plexus of branching varicose nerves was observed in the fetal paraganglia which increased in density with increasing gestational age. The majority of these nerves were VIP-IR while others were CGRP, SP, NPY, NOS or GAL-IR. The presence of nerve terminals adjacent to the paraganglion cells implies a neural influence on the functional activity of the paraganglia. Some paraganglia in the late fetal and early postnatal specimens contained Timofeew's sensory corpuscles, resembling pacinian corpuscles in their morphology. The central nerve fibre of these corpuscles displayed immunoreactivity for SP, CGRP and NOS, the latter indicating a possible role for NO in afferent transmission from the urinary bladder. In addition, a few corpuscles were penetrated by a noradrenergic nerve fibre immunoreactive for NPY and TH, which may have a modulatory role on the sensory receptor.
Assuntos
Desenvolvimento Embrionário e Fetal , Gânglios Sensitivos/embriologia , Neuropeptídeos/análise , Paragânglios não Cromafins/embriologia , Bexiga Urinária/embriologia , Bexiga Urinária/inervação , Peptídeo Relacionado com Gene de Calcitonina/análise , Pré-Escolar , Dopamina beta-Hidroxilase/análise , Galanina/análise , Gânglios Sensitivos/química , Gânglios Sensitivos/crescimento & desenvolvimento , Humanos , Imuno-Histoquímica , Lactente , Recém-Nascido , Masculino , Neuropeptídeo Y/análise , Óxido Nítrico Sintase/análise , Paragânglios não Cromafins/química , Paragânglios não Cromafins/crescimento & desenvolvimento , Somatostatina/análise , Substância P/análise , Tirosina 3-Mono-Oxigenase/análise , Bexiga Urinária/crescimento & desenvolvimento , Peptídeo Intestinal Vasoativo/análiseRESUMO
The purpose of this presentation is to describe the distribution of noradrenergic nerves in the human genitourinary system. The techniques which have been employed include formaldehyde-induced fluorescence and immunocytochemical methods to demonstrate dopamine beta-hydroxylase and tyrosine hydroxylase. These methods have been applied to human fetal, neonatal, infant, child and adult tissues removed either at post mortem examination or by surgical excision. The innervation of the fetal urinary bladder is well established by 13 weeks and, as in older specimens, the detrusor receives a sparse noradrenergic nerve supply. In contrast the smooth muscle of the terminal ureter is well supplied by this type of autonomic nerve. An additional incomplete muscle layer has been identified as a nomal component of the terminal ureter which is richly innervated by noradrenergic nerves. In some cases this muscle forms a complete collar which may be responsible for ureteric obstruction. By comparison with the detrusor, bladder neck smooth muscle receives a dense noradrenergic nerve supply particularly in the male. Unlike the detrusor, the structure and innervation of the vas deferens, seminal vesicle and prostate are poorly differentiated in the fetus. In the infant and child, the structure of the intramural smooth muscle of these organs remains immature although a rich noradrenergic nerve supply resembing the adult has been established in the fetus by 30 weeks. In the fetus, autonomic ganglia occur in association with noradrenaline rich paraganglia and surprisingly, with sensory nerve endings resembling pacinian corpuscles. Shortly after birth paraganglia are no longer associated with the autonomic ganglia of the genitourinary system. On the basis of size at least two types of autonomic neuron populate these autonomic ganglia. One type is relatively large and devoid of catecholamines but is closely associated with pericellular noradrenergic nerve fibres. The second type of neuron is small, contains noradrenaline and is arranged in clusters closely related to the capsule of the prostate gland. The significance of these observations will be considered with respect to the neurological control of the genitourinary system.
Assuntos
Sistema Nervoso Autônomo/anatomia & histologia , Norepinefrina/análise , Sistema Urogenital/inervação , Sistema Nervoso Autônomo/embriologia , Sistema Nervoso Autônomo/metabolismo , Humanos , Masculino , Próstata/inervação , Glândulas Seminais/inervação , Ureter/inervação , Uretra/inervação , Bexiga Urinária/inervação , Ducto Deferente/inervaçãoRESUMO
OBJECTIVE: To use immunohistochemical methods to study the developing autonomic innervation of the human fetal vas deferens and seminal vesicle. MATERIAL AND METHODS: Thirteen pre-natal specimens ranging in gestational age from 13 to 30 weeks were acquired following abortion or miscarriage. The overall innervation of each specimen was visualized using protein gene product 9.5 (PGP), a general nerve marker, while the onset and development of specific neuropeptide-containing sub-populations were investigated using antisera to neuropeptide Y (NPY), vasoactive intestinal peptide (VIP), substance P (SP), calcitonin gene related peptide (CGRP), bombesin (BOM), somatostatin (SOM), and met-enkephalin (ENK). In addition the occurrence and distribution of presumptive noradrenergic nerves was studied using antisera to dopamine-beta-hydroxylase (D beta H) and tyrosine hydroxylase (TH). RESULTS: At 13 weeks numerous PGP, D beta H, TH, NPY and ENK immunoreactive (-IR) nerve trunks were present in the adventitia of the vas deferens and seminal vesicle but at this stage nerve fibres were not present in the smooth muscle coat of either organ. By 17 weeks, fine PGP-, D beta H, and TH-IR nerve fibres had penetrated the outer aspect of the muscle coat of the seminal vesicle but not the vas deferens. At 20 weeks a branching network of PGP-, D beta H- and TH-IR nerve fibres occurred throughout the full thickness of the muscle coat of the seminal vesicle while similar nerves were present only in the outer half of the muscle coat of the vas deferens. At 23 weeks the full thickness of the muscle coat of the vas deferens was richly innervated by a branching plexus of PGP-IR nerves. Many of these adventitial and intramuscular nerves were immunoreactive for D beta H or TH while some were immunoreactive for either NPY or ENK. Occasional adventitial nerves were immunoreactive for SP or CGRP, these being first observed at 20 weeks. VIP-IR nerves were extremely rare in the muscle coat of either organ, being first observed at 17 weeks in the seminal vesicle and at 20 weeks in the vas deferens where they mainly formed perivascular plexuses. PGP-IR nerves were first observed in the submucosa of the seminal vesicle at 20 weeks and in the vas deferens at 21 weeks. Some of these nerves were perivascular in location while other formed a subepithelial plexus which increased in density with increasing gestational age. At 22 weeks of gestation some of the submucosal nerves were immunoreactive for SP or NPY, while at 30 weeks NPY-IR nerves formed the majority of subepithelial nerves. Occasional VIP-IR subepithelial nerves were first observed at 26 weeks but were extremely rare even at 30 weeks. Submucosal nerves immunoreactive for CGRP, D beta H, TH or ENK did not occur in any of the specimens examined. CONCLUSION: (i) From 13 weeks gestation autonomic nerves develop in the muscle coat of the fetal seminal vesicle and vas deferens, being denser in the seminal vesicle than the vas deferens up to 23 weeks gestation. (ii) The majority of the intramuscular nerves in either organ contain D beta H, TH, NPY and ENK and are presumably noradrenergic in type. (iii) A subepithelial nerve plexus develops around 20 weeks gestation and contains NPY but not VIP, unlike the adult organs. (iv) Scattered neuroendocrine cells immunoreactive for SOM are present in the mucosa of the seminal vesicle from 23 weeks of gestation.
Assuntos
Sistema Nervoso Autônomo/embriologia , Glândulas Seminais/inervação , Ducto Deferente/inervação , Sistema Nervoso Autônomo/química , Idade Gestacional , Humanos , Imuno-Histoquímica , Masculino , Microscopia de Fluorescência , Músculo Liso/embriologia , Músculo Liso/inervação , Neuropeptídeos/análise , Glândulas Seminais/embriologia , Ducto Deferente/embriologiaRESUMO
OBJECTIVE: To use immunohistochemical techniques to determine the spatial and temporal distribution of a variety of neuropeptides in the human fetal and neonatal urinary bladder. MATERIALS AND METHODS: Thirteen pre-natal specimens ranging in gestational age from 17 to 35 weeks were acquired following abortion or miscarriage. In addition two post-natal specimens aged 8 and 12 weeks were obtained at post-mortem and were included in this study. The overall innervation of each specimen was visualized using the general nerve marker protein gene product 9.5 (PGP). Localization of dopamine-beta-hydroxylase (DBH) and tyrosine hydroxylase (TH) revealed putative noradrenergic nerves. The neuropeptides studied included neuropeptide Y (NPY), vasoactive intestinal polypeptide (VIP), substance P (SP), and calcitonin gene-related peptide (CGRP). RESULTS: At 17 weeks a rich plexus of PGP and NPY-containing nerves was present throughout the detrusor muscle coat. As gestational age increased, VIP, SP and CGRP-containing nerves were observed with increasing frequency although SP and CGRP were mainly confined to perivascular nerve plexuses. TH- and DBH-containing nerves were first observed in the intramural ureters at 30 weeks and the detrusor muscle at 35 weeks and were relatively numerous in the intramural ureters and muscle of the superficial trigone in the two post-natal specimens. PGP-containing nerves were first observed beneath the bladder epithelium at 23 weeks and gradually became more numerous with increasing age. Occasional NPY, VIP, SP and CGRP-containing nerves were observed in the submucosa but TH- and DBH-immunostained nerves were especially numerous in the mucosa of the trigone in the two post-natal specimens, many such nerves being unrelated to the vascular supply. CONCLUSIONS: The bladder detrusor possesses a rich autonomic innervation by 17 weeks of gestation and this presumptive cholinergic innervation is associated with NPY immunoreactivity. Presumptive noradrenergic nerves appear relatively late in pre-natal development and mainly supply the intramural ureters and superficial trigone. A submucosal plexus of nerves has been demonstrated, the functional significance of which remains uncertain.
Assuntos
Neuropeptídeos/análise , Bexiga Urinária/inervação , Peptídeo Relacionado com Gene de Calcitonina/análise , Dopamina beta-Hidroxilase/análise , Feminino , Humanos , Imuno-Histoquímica , Lactente , Recém-Nascido , Masculino , Neuropeptídeo Y/análise , Substância P/análise , Tioléster Hidrolases/análise , Ubiquitina Tiolesterase , Bexiga Urinária/química , Peptídeo Intestinal Vasoativo/análiseRESUMO
Double-label immunocytochemistry was used to investigate the co-localisation of neuropeptides and the enzyme nitric oxide synthase (NOS) with tyrosine hydroxylase (TH) in autonomic ganglia of the human postnatal male pelvic plexus. Postmortem specimens were obtained from six male infants and children ranging in age from 2 to 12 months who had died as a result of cot death or accidental trauma. On average, ganglia lying adjacent to the neck of the urinary bladder contained 45% of neurons which were TH-immunoreactive (-IR) while ganglia situated adjacent to the posterior and lateral aspects of the prostate gland contained 67% of neurons which were TH-IR. All the TH-IR neurons also contained dopamine beta-hydroxylase and were considered to be noradrenergic in type. On average, 61% of TH-IR neurons in bladder ganglia contained NOS, compared with 77% of non-TH-IR neurons (based on counts of over 1,000 cells in each case), while the percentages of TH- and non-TH-IR neurons containing neuropeptides were: calcitonin gene-related peptide (CGRP) (30%; 11%), neuropeptide Y (NPY) (66%; 92%), somatostatin (SOM) (70%; 29%), substance P (SP) (64%; 46%), vasoactive intestinal polypeptide (VIP) (64%; 83%). The equivalent values for TH- and non-TH-IR neurons in prostatic ganglia were NOS (38%; 59%), CGRP (55%; 18%), NPY (62%, 65%), SOM (14%, 20%), SP (13%, 8%) and VIP (42%; 82%). Varicose nerve fibers within the ganglia were seen forming pericellular arborizations around many of the ganglion cells, the most numerous containing TH-, CGRP-, NPY-, SOM- or VIP-immunoreactivity. Less common were pericellular varicosities containing SP-immunoreactivity while terminals containing NOS were not observed. No correlation could be detected between the peptide contents of the ganglion cells and of the associated pericellular terminals. However, the peptide content of the ganglion cells found in association with the urinary bladder and prostate gland correlates well with the previously documented coexistence of enzymes and neuropeptides in the intrinsic nerve fibers supplying these two regions of the human postnatal male genitourinary system.
Assuntos
Gânglios Autônomos/metabolismo , Neurônios/metabolismo , Neuropeptídeos/metabolismo , Óxido Nítrico Sintase/metabolismo , Tirosina 3-Mono-Oxigenase/metabolismo , Dopamina beta-Hidroxilase/metabolismo , Gânglios Autônomos/citologia , Gânglios Autônomos/enzimologia , Humanos , Imuno-Histoquímica , Lactente , Masculino , Terminações Nervosas/enzimologia , Terminações Nervosas/metabolismo , Neurônios/fisiologia , Pelve/inervação , Sinaptofisina/metabolismoRESUMO
Double-label immunocytochemistry was used to investigate the colocalisation of various neuropeptides and the enzymes nitric oxide synthase (NOS) and tyrosine hydroxylase (TH) in intramural ganglia of the human male urinary bladder neck and trigone. Postmortem specimens were obtained from 7 male infants and children ranging in age from 2 mo to 3 y who had died as a result of cot death or accidental trauma. On average 60% of the intramural neurons were non-TH-immunoreactive (-IR) (i.e. presumptive cholinergic) and 40% were TH- and D beta H-IR (i.e. noradrenergic). Within the non-TH-IR population, calcitonin gene-related peptide (CGRP) was found in 65% of cells, neuropeptide Y (NPY) in 90%, nitric oxide synthase (NOS) in 45%, somatostatin (SOM) in 90%, and vasoactive intestinal polypeptide (VIP) in 40%. The corresponding values for the TH-IR neurons were CGRP (54%), NPY (70%), NOS (58%), SOM (73%) and VIP (40%). All the observed bombesin (BOM)-immunoreactivity was colocalised with TH while 90% of VIP and almost all the CGRP was colocalised with NPY. Less than 5% of neurons were immunoreactive for substance P (SP) or met-enkephalin (m-ENK) and some of these also contained TH. Varicose nerve fibres were seen in close proximity to some of the intramural neurons, the majority of such varicosities showing immunoreactivity to CGRP, VIP or TH. Less common were pericellular varicosities immunoreactive to NPY, SOM or SP. These results demonstrate the neurochemical heterogeneity of intramural neurons in the human bladder neck and provide indirect evidence for the complexity of the peripheral innervation of the human urinary bladder.
Assuntos
Gânglios/química , Óxido Nítrico Sintase/análise , Nervos Periféricos/química , Tirosina 3-Mono-Oxigenase/análise , Bexiga Urinária/inervação , Peptídeo Relacionado com Gene de Calcitonina/análise , Pré-Escolar , Dopamina beta-Hidroxilase/análise , Gânglios/enzimologia , Humanos , Imuno-Histoquímica , Lactente , Masculino , Nervos Periféricos/enzimologia , Substância P/análise , Peptídeo Intestinal Vasoativo/análiseRESUMO
OBJECTIVE: To determine the distribution and patterns of co-localization of nitric oxide synthase (NOS), neuropeptides and tyrosine hydroxylase (TH) in intrinsic nerves of the human post-natal vas deferens and seminal vesicle. MATERIALS AND METHODS: Double and triple immunolabelling methods were used in tissue from 10 male infants and children (age range 2 months to 3 years) obtained at post-mortem examinations carried out within 12 h of death. RESULTS: Most nerves supplying the muscle coat of either organ were TH-immunoreactive (-IR), most of which also contained neuropeptide Y (NPY) while a smaller proportion contained both NPY and NOS. Minor populations of the TH/NPY-IR intramuscular nerves contained calcitonin gene-related peptide (CGRP), galanin (GAL), met-enkephalin (m-ENK) or vasoactive intestinal polypeptide (VIP). Non-TH-IR intramuscular nerves were relatively infrequent and most contained NPY and either VIP or NOS. Presumptive secretomotor nerves formed subepithelial plexuses in both organs, most of which contained NPY co-localized with either VIP or NOS, with minor populations containing CGRP and/or GAL. TH- and substance P (SP) -IR nerves were not observed subepithelially. Perivascular nerve plexuses were mainly formed by TH-IR varicose nerves, most of which contained co-localized NPY and CGRP, with a smaller proportion containing NPY and NOS and minor populations containing VIP, m-ENK, SP or GAL. CONCLUSION: These results indicate that the autonomic control of the human vas deferens and seminal vesicle is provided by several immunohistochemically distinct nerve populations. Furthermore, NOS is present in a proportion of both the noradrenergic and non-noradrenergic nerves. Pharmacological studies are now required to elucidate the precise roles of nitric oxide and neuropeptides in the functional control of these organs.
Assuntos
Neuropeptídeos/análise , Óxido Nítrico Sintase/análise , Glândulas Seminais/inervação , Ducto Deferente/inervação , Biomarcadores/análise , Peptídeo Relacionado com Gene de Calcitonina/análise , Pré-Escolar , Encefalina Metionina/análise , Galanina/análise , Humanos , Imuno-Histoquímica , Lactente , Masculino , Neuropeptídeo Y/análise , Tirosina 3-Mono-Oxigenase/análise , Peptídeo Intestinal Vasoativo/análiseRESUMO
Single and double-label immunofluorescence methods were used to determine the distribution and patterns of colocalisation of various neuropeptides and nitric oxide synthase (NOS) with the catecholamine synthesising enzymes tyrosine hydroxylase (TH) and dopamine-beta-hydroxylase (DbetaH) in nerve fibres within specimens of adult human vas deferens obtained at vasectomy (age range 28 to 83 y). Cholinergic nerve fibres were immunolabelled with an antiserum to vesicular acetylcholine transporter (VAChT). Using the general nerve marker protein gene product 9.5 (PGP) the density of intramural nerve fibres was found to be similar irrespective of age. Many of these axons, especially in the outer 2 muscle layers were TH and DbetaH-immunoreactive (IR) and were thus confirmed as noradrenergic. Fewer such axons were seen in the inner longitudinal muscle layer. All the noradrenergic nerve fibres also displayed NPY-immunoreactivity with minor populations containing galanin (GAL) or somatostatin (SOM). Nerve fibres lacking TH and DbetaH-IR were immunoreactive for VAChT and were sparsely distributed throughout the 2 outer muscle layers but more numerous in the inner muscle layer. Nerves lacking TH and DbetaH were immunoreactive for NPY and some also contained NOS, VIP or CGRP. These results have been compared with those obtained previously from specimens of human neonatal and infant vas deferens where, in contrast to the present results, NOS and VIP were shown to be colocalised with TH in many of the intramuscular nerve fibres. It thus appears that NOS and VIP cease their coexistence with TH in intramuscular nerve fibres of the human vas deferens between the pre- and postpubertal states. In addition to the intramuscular nerve fibres a VAChT-IR subepithelial nerve plexus occurs in the vas deferens and may control the secretory activity of the lining epithelium. Most of these subepithelial nerve fibres were immunoreactive for NPY and many also contained VIP while minor populations were immunoreactive for NOS, GAL, SOM or SP although fibres containing CGRP were not observed. The neuropeptide content of the subepithelial nerve plexus was similar to that observed in the infant, except for an increased density of VIP-IR nerves, which may reflect greater activity of the lining epithelial cells in the adult vas deferens.
Assuntos
Catecolaminas/metabolismo , Fibras Nervosas/metabolismo , Neuropeptídeos/metabolismo , Óxido Nítrico Sintase/metabolismo , Ducto Deferente/inervação , Adulto , Idoso , Idoso de 80 Anos ou mais , Envelhecimento , Biomarcadores , Humanos , Imuno-Histoquímica , Masculino , Pessoa de Meia-IdadeRESUMO
Because doubt still remains concerning the distribution of nerves that are unequivocally cholinergic in the human genitourinary organs, we have used a specific marker, namely, an antibody to vesicular acetylcholine transporter (VAChT), to immunolabel cholinergic axons and cell bodies in specimens of urinary bladder, seminal vesicle, vas deferens, and prostate gland obtained from neonates and children post mortem. In addition some sections were double-immunolabeled with VAChT and either neuropeptide Y (NPY) or nitric oxide synthase (NOS). The results demonstrated a rich cholinergic innervation to the muscle coat of the bladder body with a much less prominent, but nonetheless significant, cholinergic innervation to the smooth muscle components of the seminal vesicle, vas deferens, and prostate. Small ganglia were scattered throughout the detrusor muscle of the urinary bladder, approximately 75% of the intramural neurons being VAChT immunoreactive, whereas approximately 95% contained NPY and approximately 40% contained NOS. VAChT immunoreactivity was observed in 40% of neurons in ganglia scattered throughout the pelvic plexus. Almost all these cholinergic neurons contained NPY and approximately 65% contained NOS. Almost all the cholinergic nerve fibers throughout the genitourinary organs also contained NPY. Although NOS was sparse in the cholinergic nerves of the bladder body, it occurred in the majority of cholinergic nerves at the bladder neck and was also present in a proportion of the cholinergic nerves in the other organs examined. VAChT-immunoreactive nerves were also observed in a sub-epithelial location in all the organs examined, the majority containing NPY, whereas a small proportion contained NOS. Although doubt remains about the function of sub-epithelial cholinergic nerves in the urinary bladder, the majority of similar nerves in the seminal vesicle, vas deferens, and prostate gland are considered to be secretomotor. Collectively these findings demonstrate that the cholinergic innervation of the male genitourinary system is well established in the neonate and child. Neurourol. Urodynam. 19:185-194, 2000.
Assuntos
Proteínas de Transporte/análise , Fibras Colinérgicas/química , Genitália Masculina/inervação , Isoenzimas/análise , Proteínas de Membrana Transportadoras , Proteínas do Tecido Nervoso/análise , Neuropeptídeo Y/análise , Óxido Nítrico Sintase/análise , Sistema Nervoso Parassimpático/anatomia & histologia , Proteínas de Transporte Vesicular , Biomarcadores , Criança , Pré-Escolar , Feminino , Técnica Indireta de Fluorescência para Anticorpo , Gânglios Parassimpáticos/química , Genitália Masculina/química , Humanos , Lactente , Recém-Nascido , Masculino , Neurônios/química , Especificidade de Órgãos , Sistema Nervoso Parassimpático/química , Próstata/química , Próstata/inervação , Glândulas Seminais/química , Glândulas Seminais/inervação , Ureter/química , Ureter/inervação , Bexiga Urinária/química , Bexiga Urinária/inervação , Ducto Deferente/química , Ducto Deferente/inervação , Proteínas Vesiculares de Transporte de AcetilcolinaRESUMO
PURPOSE: The objective of this study was to examine the distribution of nitric oxide synthase (NOS) and the catecholamine-synthesizing enzyme tyrosine hydroxylase (TH) in nerve fibers supplying the human neonatal male genitourinary organs. MATERIALS AND METHODS: An indirect double label immunofluorescence technique was employed on specimens obtained from infants and children at postmortem examination. RESULTS: Many nerve fibers immunoreactive for both NOS and TH were observed in the muscle coat of the vas deferens and the seminal vesicle, within the fibromuscular stroma of the prostate gland and at the bladder neck, and also formed perivascular plexuses in each of these organs. Double-labeled nerves occurred less frequently in the intramural ureters and superficial trigone while similar nerves in the bladder body were relatively sparse. Numerous nerves immunoreactive for NOS but not TH were observed at the base of the epithelium of each organ examined. Four types of autonomic ganglion cell were observed in nearby pelvic ganglia: those which contained NOS and TH, those which contained NOS alone, those which contained TH alone and those which contained neither NOS nor TH. CONCLUSION: The results indicate that many of the noradrenergic nerves as well as non-noradrenergic nerves supplying the male genitourinary organs have the capacity to synthesize nitric oxide (NO) and that NO may play a significant role in the autonomic control of both the urinary and genital organs in the postnatal human male.
Assuntos
Fibras Nervosas/química , Óxido Nítrico Sintase/análise , Tirosina 3-Mono-Oxigenase/análise , Sistema Urogenital/química , Sistema Urogenital/inervação , Cadáver , Pré-Escolar , Humanos , Lactente , MasculinoRESUMO
OBJECTIVE: To examine the histological structure and autonomic innervation of the vesico-ureteric junction (VUJ) in three cases of primary obstructive megaureter occurring in association with ectopic ureteric insertion. PATIENTS AND METHODS: Specimens of VUJ were obtained from one male and two females with primary obstructive megaureter and ectopic ureteric insertions. Serial frozen sections were cut and stained with either Masson's trichrome or immunostained for protein gene product (PGP 9.5; a general nerve marker) or dopamine beta-hydroxylase (DbetaH), a marker for noradrenergic nerves. RESULTS: In each case examined the terminal ureter was encircled by a thick collar of smooth muscle on the inside of which lay normal narrow-diameter longitudinally orientated ureteric muscle bundles. Loose connective tissue separated the surrounding detrusor muscle from this abnormal muscle collar, which was formed of distinctive smooth muscle cells arranged in a meshwork. The autonomic innervation of the ureteric and detrusor muscle bundles was similar to that previously reported for the normal VUJ while the thick muscle collar was richly innervated by DbetaH-immunoreactive noradrenergic nerves. CONCLUSION: The presence of an additional smooth muscle collar surrounding the terminal ureter may impede the normal flow of urine from ureter to bladder and be the cause of megaureter in these patients. We propose that the presence of this muscular collar arises from the developmental anomaly and suggest that these cases represent a subgroup that requires early surgical treatment.
Assuntos
Ureter/inervação , Obstrução Ureteral/patologia , Bexiga Urinária/inervação , Criança , Dopamina beta-Hidroxilase/análise , Feminino , Humanos , Imuno-Histoquímica , Recém-Nascido , Masculino , Músculo Liso/metabolismo , Tioléster Hidrolases/análise , Ubiquitina Tiolesterase , Ureter/anormalidades , Obstrução Ureteral/metabolismoRESUMO
OBJECTIVE: To determine the histological structure and autonomic innervation of the vesico-ureteric junction (VUJ) in cases of primary ureteric reflux, to compare the results with those reported previously for non-refluxing VUJs and thus determine possible structural anomalies which could be responsible for vesico-ureteric reflux (VUR). MATERIALS AND METHODS: Nineteen specimens of VUJs with reflux were obtained from four males and 10 females undergoing ureteric reimplantation. Serial frozen sections were cut and stained either with Masson's trichrome or immunostained for protein gene product (PGP 9.5), a general nerve marker, dopamine beta hydroxylase (D beta H), a marker for noradrenergic nerves, or neuropeptide Y (NPY). RESULTS: Longitudinally orientated muscle bundles of narrow diameter accompanied the ureter to its termination at the ureteric orifice. Where they merged with those of the superficial trigone. In some specimens, an additional muscle component was identified, situated on the outer aspect of the ureteric muscle coat, consisting of an incomplete layer of distinctive muscle whose constituent cells formed an interlacing mesh-work. Occasionally, specimens included detrusor muscle which consisted of relatively large diameter compact bundles separated from the ureter by a connective tissue sheath. D beta H- and NPY-immunoreactive (-IR) nerves were numerous among the ureteric muscle bundles. Many NPY-IR nerves occurred within the detrusor muscle while D beta H-IR nerves were mainly perivascular. When present, the outer muscle component was richly innervated by both D beta H- and NPY-IR nerves. PGP immunoreactivity varied among regions in the same section, PGP-IR nerves frequently being less numerous than those containing D beta H or NPY. CONCLUSION: These results indicate that the morphology of the VUJ in cases of VUR is indistinguishable from the normal VUJ described previously. Furthermore, the density and distribution of D beta H- and NPY-IR nerves is identical to that in controls. However, the antigenicity of PGP in the reflux specimens appears to be masked in some regions by an unknown factor.