Existence of manserin, a secretogranin II-derived neuropeptide, in the rat inner ear: relevance to modulation of auditory and vestibular system

Manserin is a 40-amino acid neuropeptide derived from rat brain. Manserin has been shown to distribute in the neuroendocrine system, such as the pituitary and adrenal glands, but it has been little studied in other organs. In this study, the authors examined localization of manserin in the inner ear of the adult Wistar rat using immunohistochemical analyses. Manserin immunoreactivity was detected in the neuronal terminals of the organ of Corti and type II spiral ganglion cells. In addition to being identified in the auditory system, manserin was detected at the synapses of the vestibular system, such as saccule, utricle, and semicircular canal. These results suggest that inner ear manserin may be involved in the function of peripheral auditory and vestibular systems.     

Secretogranin II and its Derivative Peptide,Manserin, are Differentially Localized in Purkinje Cells and Unipolar Brush Cells in the Rat Cerebellum

The cerebellum has long been recognized as the primary center of motor coordination in the central nervous system. Cerebellar neuropeptides have been postulated to be involved in such motor coordination, though this role is not fully understood. We herein investigated the localization of novel neuropeptide, “manserin” in the adult rat cerebellum. Punctate signals of manserin immunoreactivity were observed in the granular layer of the rat cerebellum. Manserin signals were also observed in the fibers and fiber terminals in the granular layer as well as the molecular layer. Manserin did not localize in Purkinje cells. Interestingly, cerebellar manserin was preferentially colocalized with unipolar brush cells, a class of excitatory granular layer interneuron, which are known to be involved in vestibullocerebellar functions. These results indicate that manserin plays pivotal roles in the cerebellar functions.     

Manserin, a secretogranin II-derived peptide, distributes in the rat endocrine pancreas colocalized with islet-cell specific manner

Manserin is a recently characterized 40-amino acid neuropeptide derived from secretogranin II, a protein belonging to the chromogranin family. Although the physiological roles of manserin have not been elucidated to date, manserin has been shown to distribute in not only the brain but also the endocrine system such as the pituitary and adrenal glands, suggesting its role in the endocrine system. The present study aimed to explore the occurrence and distribution of manserin in the rat pancreas using an immunohistochemical technique with a polyclonal antibody against rat manserin. Immunoreactivity for manserin was readily detected in almost whole islets of Langerhans whereas not at all in the exocrine pancreas. Manserin-expressing cells were not colocalized with the glucagon-secreting cells (α cells), whereas they colocalized with insulin-secreting cells (β cells) and somatostatin-secreting cells (δ cells), although their intracellular distribution was different. These results indicate that manserin, occurring in the endocrine pancreas, may have a potential role in the endocrine system.     

Secretoneurin: A new peptide in the human enteric nervous system

Secretoneurin is a functional neuropeptide derived from secretogranin II (chromogranin C). This proprotein is processed to varying degrees in neuroendocrine tissues. In the present study we established by gel filtration high performance liquid chromatography that in human intestinal wall and mucosa an antiserum against secretoneurin detects as the major immunoreactive moiety the free peptide secretoneurin. In the mucosa some larger immunoreactive peptides were also present, however, a significant amount of the intact proprotein secretogranin II could not be detected. By immunohistochemistry we studied the distribution of secretoneurin within the gut. Antibodies to protein gene product 9.5 and chromogranin A were used to identify all neurons and endocrine cells, respectively, whilst those to the peptides substance P. CGRP and somatostatin were used for the further characterization of individual secretoneurin-positive structures. Secretoneurin immunoreactivity was found in nerve fibres in all layers of the gut wall. In both myenteric and submucous plexuses, nerve fibres and the majority of ganglion cells were secretoneurin-immunoreactive. In the mucosa, some secretoneurin-positive nerve processes ran parallel to the basal membrane of epithelial cells, occasionally invading the epithelial layer. Secretoneurin immunoreactivity was found in endocrine cells, mostly D cells, in the following regions in descending order of density: stomach/duodenum; rectum; colon; ileum. Thus, secretoneurin is a new major peptide within the human enteric neuroendocrine system. Its presence in abundant myenteric ganglion cells may imply a role in the modulation of gastrointestinal motility. The chemotactic properties of secretoneurin and its possible localization in sensory fibres suggest that this peptide may be involved in the genesis of intestinal inflammation.     

Biochemical characterization and immunocytochemical localization of EM66, a novel peptide derived from secretogranin II, in the rat pituitary and adrenal glands.

Characterization of secretogranin II (SgII) mRNA in various vertebrates has revealed selective conservation of the amino acid sequences of two regions of the protein, i.e., the bioactive peptide secretoneurin and a flanking novel peptide that we named EM66. To help elucidate the possible role of EM66, we examined the occurrence as well as the cellular and subcellular distribution of EM66 in rat pituitary and adrenal glands by using a polyclonal antibody raised against the recombinant human EM66 peptide. High-performance liquid chromatography (HPLC) analysis of rat pituitary and adrenal extracts combined with a radioimmunoassay resolved EM66-immunoreactive material exhibiting the same retention time as recombinant EM66. In the rat pituitary, double-labeling immunohistochemical (IHC) studies showed that EM66 immunoreactivity (IR) was present in gonadotrophs, lactotrophs, thyrotrophs, and melanotrophs, whereas corticotrophs were devoid of labeling. EM66-IR was also observed in nerve endings in the neural lobe. Immunocytochemical staining at the electron microscopic level revealed that EM66-IR is sequestered in the secretory granules within gonadotrophs and lactotrophs. In the adrenal medulla, double IHC labeling showed that EM66-IR occurs exclusively in epinephrine-synthesizing cells. At the ultrastructural level, EM66-IR was seen in chromaffin vesicles of adrenomedullary cells. These results demonstrate that post-translational processing of SgII generates a novel peptide that exhibits a cell-specific distribution in the rat pituitary and adrenal glands where it is stored in secretory granules, supporting the notion that EM66 may play a role in the endocrine system.     

Immunohistochemical Localization of Manserin,a Novel Neuropeptide Derived from Secretogranin II,in Rat Adrenal Gland,and its Upregulation by Physical Stress

We recently identified a novel 40-amino acid neuropeptide designated manserin from the rat brain (Yajima in NeuroReport 15: 1755–1759, 2004). Manserin is highly expressed in pituitary and hypothalamic nuclei, which suggests that it plays a role in the endocrine system. In this study, we employed immunohistochemical methods to investigate the presence of manserin in rat adrenal glands, as well as its regulation by physical stress. Immunohistochemical analysis using anti-manserin antibody showed that manserin is present in the rat adrenal medulla but not in the cortex. When the colocalization of manserin and phenylethanolamine N-methyltransferase (PNMT), an epinephrine-synthesizing enzyme, was examined, virtually all PNMT-positive cells expressed manserin. Interestingly, the immunoreactivity of manserin was significantly increased when the rats were exposed to water-immersion restraint stress. These results demonstrate for the first time that adrenal manserin, a novel neuropeptide, may have a potential physiological role under stress-inducing conditions.     

八肽胆囊收缩素在猪,大鼠和豚鼠肠道的免疫组织化学定位和分布

实验采用ＡＢＣ免疫组织化学方法研究了八肽胆囊收缩素样免疫反应物质在猪、大鼠和豚鼠肠道的定位与分布,并用相邻切片免疫双标记法观察了它与５－羟色胺的关系,结果表明,ＣＣＫ－８－ＩＲ细胞主要位于肠腺的底部,少数位于绒毛上皮。节段性分布上,在猪可见于从十二指肠到结肠全长的粘膜,大鼠和豚鼠ＣＣＫ－８－ＩＲ细胞则见于从十二指肠至回肠的粘膜,但均以十二指肠密度最高;免疫双标记法证实,在三种动物肠道中均有ＣＣＫ＝     

Disorganization of stroma alters epithelial differentiation of the glandular stomach in adult mice

Summary The response of adult epithelium in contact with heterologous mesenchymes/stromas was studied in three digestive organs (forestomach, glandular stomach, and duodenum). After various tissues were implanted beneath the epithelial layer of adult mice, the epithelial differentiation was examined after sacrifice of animals at intervals up to 24 weeks. In the forestomach and duodenum, the epithelial differentiation was not affected at all by the tissue implantation. In the glandular stomach, in contrast, epithelial cells exhibited altered differentiation in which chief and parietal cells disappeared and were replaced by columnar epithelial cells with PAS-positive granules. These epithelial cells often formed immature villi. Such differentiation-altered columnar epithelium (DACE) was induced by implanting any type of tissue and even by sham operation, indicating that it was induced by disorganization of the tissue-implanted stroma. The size of DACE was significantly influenced by the stage of implanted tissue; 14.5-day fetal mesenchyme induced the largest DACE, and was followed by 16.5-day fetal mesenchyme, adult stroma, and sham operation. These results suggest the importance of stromal organization in maintaining epithelial differentiation in the glandular stomach.     

Endothelin system in intestinal villi: A possible role of endothelin-2/vasoactive intestinal contractor in the maintenance of intestinal architecture

The endothelin system consists of three ligands (ET-1, ET-2 and ET-3) and at least two receptors (ETA and ETB). In mice ET-2 counterpart is a peptide originally called "vasoactive intestinal contractor" (VIC) for this reason, this peptide is frequently named ET-2/VIC. In intestinal villi, fibroblasts-like cells express endothelin's receptors and response to ET-1 and ET-3 peptides, changing their cellular shape. Several functions have been attributed to these peptides in the "architecture" maintenance of intestinal villi acting over sub-epithelial fibroblasts. Despite this, ET-2/VIC has not been analyzed in depth. In this work we show the intestine gene expression and immunolocalization of ET-1, ET-2 and the ETA and ETB receptors from duodenum to rectus and in the villus-crypt axis in mice, allowing a complete analysis of their functions. While ET-1 is expressed uniformly, ET-2 had a particular distribution, being higher at the bottom of the villi of duodenum, ileum and jejunum and reverting this pattern in the crypts of colon and rectus, where the higher expression was at the top. We postulated that ET-2 would act in a cooperative manner with ET-1, giving to the villus the straight enough to withstand mechanical stress.     

凹耳蛙消化道组织学和嗜银细胞形态观察

为了揭示凹耳蛙(Odorrana tormota)消化道的基本特征,运用石蜡切片法和龙桂开银浸法对凹耳蛙消化道组织学结构及嗜银细胞的形态与分布密度进行了观察。结果显示:①凹耳蛙的胃壁具明显的纵行皱襞和胃小凹,胃腺发达,小肠可分为十二指肠和回肠,杯状细胞分散在十二指肠上皮细胞之间,十二指肠中未见十二指肠腺分布。②凹耳蛙嗜银细胞见于消化道全长,呈毛笔头样、锥体形、梭形、椭圆形和长条形等;幽门腺上皮和十二指肠绒毛上皮中的嗜银细胞具指向腺泡腔或肠腔的突起,提示其可能具有腔分泌的功能。嗜银细胞的分布密度胃幽门部最高,十二指肠和胃体其次,食道最低。据此认为胃既是凹耳蛙的主要消化器官,也是消化道中主要的内分泌器官;十二指肠是凹耳蛙消化道中的主要吸收部位,同时也具有内分泌功能;消化道嗜银细胞具有内分泌的功能,还可能具有腔分泌的功能。     

Comparative immunohistolocalization of carbonic anhydrase isozymes I, II and III in the equine and bovine digestive tract

Summary Immunohistochemical localizations of carbonic anhydrase isozymes (CA-I, CA-II and CA-III) in equine and bovine digestive tracts were studied. In the horse, epithelial cells in both the oesophagus and non-glandular part of the stomach lacked all three isozymes. In contrast, surface epithelial and parietal cells in the glandular region of the stomach showed reactivity for CA-II. In the small intestine, absorptive columnar cells covering the villi in the duodenum were positive for CA-II. The epithelium of the jejunum and ileum lacked all three isozymes. In the large intestine, CA-II was detected in the columnar cells in the upper part of the crypt. In cattle, epithelial cells of the oesophagus showed reactions for CA-I and CA-III but not for CA-II. Although the absorptive epithelial cells of the small intestine lacked CA-I, CA-II and CA-III, those of the upper part of large intestine crypts were heavily stained for all three isozymes.     

扬子鳄消化道嗜银细胞的分布及形态学观察

该文用龙桂开浸银法对扬子鳄消化道嗜银细胞的分布及形态进行了观察。结果表明：嗜银细胞分布于整个消化道中,从食道到直肠。其中,十二指肠和回、直肠交接处密度很高,胃体及直肠很低。嗜银细胞形态多样。食道嗜银细胞位于上皮基部和固有膜中,呈椭圆形或不规则形。胃嗜银细胞位于胃腺部,圆形或椭圆形,有的可见明显的胞突。肠嗜银细胞位于上皮细胞之间,呈长柱形、纺锤形、长颈瓶形或锤状。多数细胞两端有较长胞突,分别与固有膜     

黑麂肝、小肠和大肠的组织学结构及Ghrelin的分布

研究了成年雌性黑麂(Muntiacus crinifrons)的肝、小肠和大肠的组织学结构及Ghrelin的分布。采用H.E染色法观察组织学结构,免疫组化PV-9000两步法并结合DAB显色技术确定Ghrelin的分布。结果表明,黑麂的肝组织分为被膜、肝小叶、肝中央静脉、门管区等结构。被膜为浆膜结构,肝小叶不明显。肝细胞以中央静脉为中心,呈放射状排列。肝血窦的形状不规则。肠黏膜上皮为单层柱状上皮,小肠、盲肠和结肠的黏膜肌层很薄,管壁皱襞与肠绒毛等形态在消化道各部也存在差异。免疫组化结果显示肝细胞中有Ghrelin阳性细胞的表达;在肠道,免疫阳性细胞在十二指肠、空肠、回肠、盲肠和直肠的黏膜、黏膜下层和肌层均有分布,尤其在肠绒毛上皮和黏膜下层分布较多。黑麂肝、小肠和大肠结构与哺乳动物基本相似,但无十二指肠腺;Ghrelin阳性细胞在肝、小肠和大肠均有分布,这表明Ghrelin可能对消化有一定的调节作用。     

Immunohistochemical detection of secretoneurin, a novel neuropeptide endoproteolytically processed from secretogranin II, in normal human endocrine and neuronal tissues

Summary An antiserum raised against a synthetic peptide derived from the primary amino sequence of rat secretogranin II (chromogranin C) was used for immunological (quantitative radioimmunoassay analysis) and immunohistochemical studies of normal human endocrine and nervous tissues. This antibody recognized a novel and biologically active neuropeptide which was coined as secretoneurin. In endocrine tissues, secretoneurin was mainly co-localized with chromogranin A and B with some exceptions (e.g., parathyroid gland). Secretoneurin was demonstrated immunohistochemically in the adrenal medulla, thyroid C cells, TSH- and FSH/LH-produting cells of the anterior pituitary, A and B cells of pancreatic islets, in endocrine cells of the gastrointestinal tract and the bronchial mucosa, and the prostate. Immunoreactivity determined by radioimmunoassay analysis revealed high secretoneurin levels in the anterior and posterior pituitary and lower levels in pancreatic and thyroid tissue. A strong secretoneurin immunoreactivity was also found in ganglion cells of the submucdsal and myenteric plexus of the gastrointestinal tract, and in ganglionic cells of dorsal root ganglia, peripheral nerves, and ganglion cells of the adrenal medulla. Thus, secretoneurin may serve as a useful marker of gangliocytic/neuronal differentiation.     

中华花龟消化道5-羟色胺免疫活性内分泌细胞的分布与形态学观察

应用5-HT抗血清,以ABC（avidin-biotin-peroxidase complex）免疫组织化学方法,探究中华花龟消化道内5-HT免疫反应阳性内分泌细胞的分布及形态。结果表明,5-HT阳性细胞从食管到直肠的消化道各段均有分布,以十二指肠部位分布密度为最高,胃幽门部次之,食管部最低;5-HT阳性细胞广泛分布于上皮细胞之间、上皮基部、腺泡上皮细胞之间、腺泡之间以及固有膜内,形态多样呈圆形、椭圆形、锥体形、梭形等,以锥体形为主。认为中华花龟消化道5-HT阳性细胞具有内、外分泌两种作用途径,并且5-HT阳性细胞的密度分布可能与其食性、生活环境等有关。     

Morphogenesis of intestinal villi. I. Scanning electron microscopy of the duodenal epithelium of the developing chick embryo

Development of villi in the duodenum of the chick was studied in stages ranging from 11 days of incubation to one week after hatching. Formation of definitive villi is preceded by development of a set of previllous ridges that run lengthwise along the duodenum. The first set of 16 previllous ridges (Set I) is complete by about 13 days of incubation; all ridges in the set are fairly uniform and proceed through their subsequent development in synchrony. Previllous ridges in Set I fold into a highly regular zigzag pattern between 14 and 16 days of incubation. Definitive villi develop from Set I ridges beginning at about 17 days when populations of distinct cells appear on the crests of the ridges between angles in the zigzag folds. Cells in these populations lack the rounded appearance of cells seen in earlier stages; their apical surfaces are densely covered with microvilli. A second set of villi (Set II) develops at about 16 days of incubation when about 16 rows of tongue-like flaps erupt between the previllous ridges of Set I. At hatching, Set II villi are still smaller than villi of Set I; this distinction disappears by about the fourth day after hatching. The significance of the morphological changes in epithelial cells is discussed in terms of several hypotheses bearing on the mechanisms of villus formation.     

Secretogranin II: Relative Amounts and Processing to Secretoneurin in Various Rat Tissues

Abstract: Secretoneurin is a 33-amino-acid peptide produced in vivo from secretogranin II. An antiserum raised against this peptide recognizes both the free peptide and its precursors. By HPLC and radioimmunoassay we characterized the immunoreactive molecules and determined the levels of immunoreactivity in various rat organs. In adrenal medulla and to a lesser degree in the anterior pituitary processing of secretogranin II to secretoneurin was very limited, whereas in all other organs studied (brain, intestine, endocrine pancreas, thyroid gland, and posterior pituitary) a high degree of processing was apparent. Thus, practically all of the immunoreactivity was present as free secretoneurin. This was also true for serum. When the total amount of secretoneurin immunoreactivity was calculated for the various organs, the largest pools in descending order were in the intestine, CNS, anterior pituitary, pancreas, and adrenal gland. This makes it likely that secretoneurin in serum is mainly derived from the intestine. The high degree of processing of secretogranin II in most organs is consistent with the concept that this protein acts as a precursor of a functional peptide, i.e., secretoneurin.     

Eimeria dispersa, E. adenoeides, and E. meleagrimitis: intestinal mucosal disruption in turkeys as seen with scanning electron microscopy.

Tissues from the digestive tract of turkeys infected with Eimeria dispersa, E. adenoeides, or E. meleagrimitis were compared with tissues from uninfected controls as seen with light microscopy (LM) and scanning electron microscopy (SEM). Six regions were examined--duodenum, jejunum, ileum, cecal neck, cecal pouch, and large intestine. Although LM showed large numbers of E. dispersa in the epithelial cells, SEM usually showed little mucosal disruption. Occasionally the surface of duodenum and jejunum, as seen with SEM, was convoluted and disrupted. In one bird, some parasite-induced damage was found with either LM or SEM in the duodenum and jejunum of turkeys given E. adenoeides oocysts. Although LM showed parasites in the rest of the digestive tract of all E. adenoeides infected birds, SEM showed only a localized sloughing of the mucosa in the cecal pouch. The most extensive damage to the villar surface was caused by E. meleagrimitis. Infections often disrupted the villar tips, especially in the small intestine and cecal neck. Localized areas of pitting were often found on individual villi. All 3 species produced oocyst extrusion sites, especially in the ileum and the ceca.     

Serotonin-containing epithelial cells in rat duodenum. I. Quantitative morphometric study of the distribution density

Summary Serotonin (5HT)-containing epithelial cells in rat duodenum were studied quantitatively by three-dimensional morphometric analysis. Longitudinal sections covering the whole length of rat duodenum were stained by either 5HT immunohistochemistry or by glyoxylic acid fluorescent histochemistry. Three-dimensional values for positive cell density, namely the number of 5HT cells per unit volume of the epithelium, were obtained by stereological morphometry with the aid of a computer-assisted image analyzer. This analytical method provides an absolute value for the distribution density of 5HT-containing cells regardless of thickness of sections, or which of the two histochemical procedures is used. The mean number of such cells per unit volume was higher in the crypts than in the villi but varied little along the duodenum. The density of 5HT cells in a given duodenal region, however, varied greatly among individual animals. The villi of the 10 to 16-mm segment from the pylorus were identified as having the smallest individual variation and therefore as being the most suitable for statistical evaluation in future pharmacohistochemical investigations.