Serotonin Cells Research Articles

Granin proteins (chromogranin A and secretogranin II C23-3 and C26-3) in the intestine of reptiles

The occurrence, distribution and the possible cellular co-localizations of chromogranin A (CgA) and of two synthetic secretogranin II-peptides (SgIIC23-3 and SgIIC26-3) with several enteric neuropeptides and serotonin have been investigated immunohistochemically in turtles, lizards and snakes. The distribution of CgA-immunoreactivity was restricted only to the enteroendocrine cells in all the reptiles studied. SgII-immunoreactivity--absent in turtle--revealed nerve cells and fibers, besides enteroendocrine cells in lizard and snake guts. Moreover, the two antisera (C23-3 and C26-3) raised against the different regions of the SgII-molecule yielded distinct distribution patterns of immunoreactivity both in the lizard and snake organs. Small amounts of enteric serotonin cells co-stored CgA or SgIIC23-3 in lizards and snakes and only SgIIC26-3-peptide in snakes. CgA was found co-stored with somatostatin in a few enterocytes of the turtle duodenum. In the same gut segment of lizards and throughout the snake organ, neurotensin and the SgIIC23-3-peptide co-existed in a small number of endocrine cells. The pancreatic polypeptide-containing cells were devoid of immunoreactivity both for CgA and SgII. Bombesin immunopositive cells were absent throughout the intestines of the reptiles investigated. The above findings entirely support the heterogenous distribution of granins in neuroendocrine organs and tissues and also within the same neuroendocrine cell population. They further support the concept of a good conservation of granins during phylogeny.

Autoinhibition of serotonin cells: an intrinsic regulatory mechanism sensitive to the pattern of usage of the cells.

After periods of high-frequency firing, the normal rhythmically active serotonin (5HT)-containing neurosecretory neurons of the lobster ventral nerve cord display a period of suppressed spike generation and reduced synaptic input that we refer to as "autoinhibition." The duration of this autoinhibition is directly related to the magnitude and duration of the current injection triggering the high-frequency firing. More interesting, however, is that the autoinhibition is inversely related to the initial firing frequency of these cells within their normal range of firing (0.5-3 Hz). This allows more active 5HT neurons to resume firing after shorter durations of inhibition than cells that initially fired at slower rates. Although superfused 5HT inhibits the spontaneous firing of these cells, the persistence of autoinhibition in saline with no added calcium, in cadmium-containing saline, and in lobsters depleted of serotonin suggests that intrinsic membrane properties account for the autoinhibition. A similar autoinhibition is seen in spontaneously active octopamine neurons but is absent from spontaneously active gamma-aminobutyric acid cells. Thus, this might be a characteristic feature of amine-containing neurosecretory neurons. The 5HT cells of vertebrate brain nuclei share similarities in firing frequencies, spike shapes, and inhibition by 5HT with the lobster cells that were the focus of this study. However, the mechanism suggested to underlie autoinhibition in vertebrate neurons is that 5HT released from activated or neighboring cells acts back on inhibitory autoreceptors that are found on the dendrites and cell bodies of these neurons.

Co-localization of synaptophysin with different neuroendocrine hormones in the human gastrointestinal tract.

Colocalisation of synaptophysin has been studied in different neuroendocrine cell types in histologically normal mucosa from human gastrointestinal tract (corpus, antrum, duodenum, ileum and colon) using double-immunofluorescence stainings. Numerous synaptophysin immunoreactive cells were seen in the antrum, while a smaller number were found in the intestinal tract. Synaptophysin immunoreactivity was strong in the antrum but weak in the intestine. In the intestinal colocalisation studies the synaptophysin immunoreactivity was enhanced by using the tyramide amplification method. Synaptophysin and chromogranin A were colocalised but the latter occurred mainly basally, whereas synaptophysin was found to occur diffusely throughout the cytoplasm. Synaptophysin immunoreactivity occurred in the serotonin cells throughout the gastrointestinal tract, and in the antral gastrin and somatostatin cells. In the intestinal tract only a small fraction of somatostatin, gastrin, cholecystokinin, enteroglucagon, enteroglucagon/ peptide tyrosine tyrosine displayed synaptophysin immunoreactivity. In the gastrointestinal tract (except the antrum), chromogranin A is a better general neuroendocrine marker than synaptophysin. The functional role of synaptophysin is unclear but it may be involved in the intracellular transport and release of hormones. Based on the distribution background of synaptophysin, it seems to be of greater importance in the antrum than in the intestinal tract as a whole.

Gastrointestinal endocrine cells in an animal model for human type 2 diabetes.

Endocrine cells were investigated in various parts of the gastrointestinal tract in an animal model for human type 2 diabetes, namely the homozygous obese mouse. As controls, age- and sex-matched homozygous lean mice were used. The different endocrine cell types were stained by using immunocytochemistry and quantified by computerized image analysis. The numbers of PYY- and enteroglucagon-immunoreactive (IR) cells were decreased in the colon of obese diabetic mice vis-à-vis controls. Serotonin-IR cells were significantly decreased in numbers in the duodenum and colon. Furthermore, colonic serotonin cells had a high cell secretory index (CSI), but an unchanged nuclear area, indicating an impaired cellular release of this amine. There was no statistical difference between obese diabetic mice and lean controls regarding the numbers and CSI of antral gastrin/CCK-, somatostatin- and serotonin-; and duodenal secretin-, gastric inhibitory peptide (GIP)-, CCK/gastrin-, and somatostatin-IR cells; nor was there any difference regarding nuclear area, with the exception of the antral somatostatin- and duodenal GIP-IR cells. It is concluded that the abnormalities in gut endocrine cells observed in the present study might explain the gut dysmotility reported in animal models for human diabetes. They may also be of importance in gastrointestinal dysfunction occurring in diabetes patients.

Pax 4 and 6 regulate gastrointestinal endocrine cell development

The mechanisms behind the cell-specific and compartmentalized expression of gut and pancreatic hormones is largely unknown. We hereby report that deletion of the Pax 4 gene virtually eliminates duodenal and jejunal hormone-secreting cells, as well as serotonin and somatostatin cells of the distal stomach, while deletion of the Pax 6 gene eliminates duodenal GIP cells as well as gastrin and somatostatin cells of the distal stomach. Thus, together, these two genes regulate the differentiation of all proximal gastrointestinal endocrine cells and reflect common pathways for pancreatic and gastrointestinal endocrine cell differentiation.

The nature and implication of intestinal endocrine cell changes in coeliac disease.

Coeliac disease is associated with intestinal lesion. This lesion causes architectural derangement of the mucosa in the form of villus atrophy, increased crypt length and increased volume of the lamina propria. Several changes in the intestinal endocrine cells have been reported over the years, e.g. the number of secretin cells and increased numbers of GIP, CCK/gastrin, motilin, and serotonin cells. There is no consensus about the nature of the changes in somatostatin-cells. It has been postulated that the changes in the endocrine cells are a selective process to meet the new demands exerted by the dramatic decrease in intestinal absorptive area. It has been speculated further that the changes in the endocrine cells would cause an incomplete digestion of the ingested food and its rapid elimination from the intestine. These changes may be responsible for the diarrhoea and steatorrhoea that occur in patients with coeliac disease.

Colonic endocrine cells in patients with carcinoma of the colon.

To investigate the colonic endocrine cells in patients with colon carcinoma in order to establish a possible abnormality. Twelve patients with colon adenocarcinoma (eight women and four men; mean age 69 years; range 52-88 years) were studied. As controls, macroscopically and histologically normal tissues from the colon of 12 patients (eight women and four men; mean age 66 years; range 34-86 years) were examined. These patients suffered from polyps, prolapsus, chronic diverticulitis, volvulus or haemorrhoids. Macroscopically and histologically normal tissues from the colon of the patients, about 10 cm from the tumour, and of the controls were examined. Endocrine cells were immunostained with the avidin-biotin-complex method, and were quantified by computer image analysis using an automatic standard sequence analysis operation. Three parameters were used: (1) the number of endocrine cells per mm3 of epithelial cells; (2) the cell secretory index (CSI); and (3) the nuclear volume. The numbers of somatostatin- and serotonin-immunoreactive cells were significantly reduced in patients with colon carcinoma (P = 0.03 and 0.009, respectively). There was no difference between patients and controls regarding the numbers of PYY-, PP-, and enteroglucagon-immunoreactive cells. The CSI of somatostatin- and serotonin-immunoreactive cells were significantly decreased. There was no difference in the CSI between the patients and controls regarding PYY-, PP- or enteroglucagon-immunoreactive cells. The nuclear volumes of PYY- and enteroglucagon-immunoreactive cells increased significantly in the patients. The nuclear volume of PP-, somatostatin- and serotonin cells did not differ from those of the controls. The present results support the assumption that a disturbance in the colonic neuroendocrine system occurs in patients with colon carcinoma, which might affect the development of the tumour. The decrease in the number and CSI of somatostatin cells may account for the decrease of the colonic content of this peptide observed previously in these patients. The decrease in the number and CSI of serotonin-immunoreactive cells in patients with colon adenocarcinoma might be a method by which the body defends itself against this mitogenic substance.

Homeobox gene product Nkx 6.1 immunoreactivity in nuclei of endocrine cells of rat and mouse stomach.

The homeobox gene product Nkx 6.1 is of unknown function but is expressed in the pancreas and the antropyloric mucosa of the stomach. In the adult pancreas, Nkx 6.1 possesses an insulin cell-restricted distribution, whereas its localization in the stomach is unknown. We now show that the vast majority of serotonin-producing enterochromaffin cells of the antropyloric mucosa contain Nkx 6. 1-immunoreactive nuclei. In addition, a subpopulation of cells co-storing serotonin and gastrin display Nkx 6.1-positive nuclei. Such cells have been postulated to represent precursors of mature gastrin and serotonin cells. The nuclei of the co-storing cells have previously also been found to be positive for another homeodomain protein, Pdx-1. Pdx-1-deficient animals were therefore investigated and were found to be devoid of Nkx 6.1-positive nuclei. Our data show that Pdx-1 is needed for Nkx 6.1 expression and suggest a role for Nkx 6.1 in the maturation of gastrin- and serotonin-positive precursor cells.

Eagle is required for the specification of serotonin neurons and other neuroblast 7-3 progeny in the Drosophila CNS

During development of the Drosophila nerve cord, neuroblast 7-3 gives rise to a pair of mitotic sister serotonin neurons in each hemisegment. Here we show that the zinc finger gene eagle, which is expressed in neuroblast 7-3, is essential for specifying the fate of serotonin neurons. We find that loss-of-function eagle mutations produce an unusual differential phenotype with respect to the sister serotonin cells and that eagle is necessary for the maintenance of engrailed and zfh-2 expression in the serotonin neurons. We present a model that uniquely identifies all progeny neurons in the neuroblast 7-3 lineage based on the expression of specific molecular markers, position within the nerve cord and the effect of eagle loss-of-function mutations. Although serotonin is an important neurotransmitter conserved throughout the animal kingdom, we show that hypomorphic alleles of eagle can produce viable adults that have a dramatic reduction in the number of serotonin-producing neurons.

GTP-cyclohydrolase-I like immunoreactivity in rat brain

GTPCH-I immunoreactive structures in the rat brain were studied using a polyclonal antibody raised in the chick. General mapping was made using the avidin–biotin–peroxidase technique and compared with the distribution of tyrosine hydroxylase and serotonin immunoreactivities. Double immunofluorescence was performed in order to establish real intracellular colocalization. GTPCH-I immunoreactivity was generally found to be low. Immunostained neurons were present in all the serotonin cell groups. In catecholaminergic neurons, although tyrosine hydroxylase immunoreactivity was always very high, GTPCH-I immunoreactivity was extremely variable, from relatively strong (substantia nigra, ventral tegmental area) to low (locus coeruleus, caudal part of the hypothalamus), extremely low (rostral hypothalamus, ventral brainstem) or almost absent (dorsal brainstem, some hypothalamic nuclei). When feasible, double immunolabeling revealed that all the serotonin cells and most of the tyrosine hydroxylase cells were also expressing GTPCH-I. Our results argue in favor of a regulation of tyrosine hydroxylase activity by the intracellular synthesis of BH4.

Effect of hypophysectomy on endocrine cell types in rat gastrointestinal mucosa.

The effect of hypophysectomy on the gastrointestinal tract was studied in the rat 8 wk after operation, particularly regarding the frequency and distribution of serotonin, somatostatin and gastrin-immunoreactive cells. Body weight, the length of the intestine and the thickness of the mucosa of the antrum and small intestine were all reduced in the hypophysectomised rats compared with sham-operated and untreated controls. In the hypophysectomised animals the serotonin-immunoreactive cells were fewer in the antrum and caecum, whereas they were more numerous in the proximal large intestine. There were fewer gastrin-immunoreactive cells in the antrum, while the somatostatin-immunoreactive cells were more numerous in the antrum and caecum. The significant influence of hypophysectomy on the gastrointestinal tract could be direct, but could also be associated with the marked effect of pituitary deficiency on endocrine cells, known to exert both trophic and antitrophic actions. However, it could also be an indirect effect on metabolism, resulting in lower food intake, other endocrine cell systems, and growth factors.

Serotonin (5-HT): An enteroendocrine marker of rejection after small bowel allotransplantation in the pig

The histopathologic findings of mucosal epithelial cell injury in acute and chronic rejection in small bowel transplantation are well known. Several studies have been done on the pig either as autotransplantation, allotrans-plantation, orthotopic, or enterotopic transplantation, with or without immunosuppression. Very few investigations on the enteroendocrine system have been done. In particular, none have been done on rejection in pigs after intestinal transplantation. On the other hand, changes in the enteroendocrine markers (somatostatin and chromogranin A) have been examined in detail after auxiliary small bowel transplantation in dogs without immunosuppression, showing no apparent changes on the 7th postoperative day, even if in a severe rejection status. Recent studies on rats have evaluated the presence of serotonin after small bowel allotransplantation with different times of preservation, and after reperfusion. The authors concluded that the number of serotonin-positive cells decreased with a preservation time of more than 1 hour in Ringer lactate at 4°C. We too have examined the modifications of argentaffin as well as serotonin-positive cells in the harvested small bowel of pigs perfused with Belzer solution (4°C) and preserved in Ringer lactate at 4°C, after preservation and reperfusion in the combined liver-small bowel allotransplantation. We noticed that the argentaffin and serotonin cells were preserved up to 20 hours and that after reperfusion the most important injury was due to the disappearance of these cells from the crypts. In this study, we studied the modifications of the intestinal enterochromaffin cells after combined liver-small bowel and small bowel alone allotransplantation in pigs, with the aim of evaluating a correlation with rejection.

Serotonin (5-HT): An enteroendocrine marker of reperfusion injury after liver-small bowel allotransplantation in the pig

HISTOMORPHOLOGIC and functional aspects of preservation/reperfusion injury, after small bowel transplantation, have been well investigated after different modalities and types of perfusion solutions. Experimental and clinical studies on bowel preservation damage have well defined the modifications of enteric and Goblet cells, vascular structures, myoenteric nerves, and ganglia. The enteroendocrine system is not so well studied. In recent studies of Lai et al, 1 on auxiliary small bowel allotransplantation in the dog without immunosuppression, biopsies of the small bowel were done daily and at time of death; specimens were stained with immunohistochemical methods for chromogranin A and somatostatin. Also, in animals with severe rejection, this type of enteroendocrine cell did not present any evident modification. Kaihara et al 2 suggested serotonin as a good marker of ischemia/reperfusion injury after allotransplantation of the small bowel in the rat. In their experience, the release of serotonin in the lumen bowel after reperfusion inversely correlated with the number and quality of serotonin cells. Our previous experiences with normothermic preservation in Ringer lactate solution, up to 20 hours in the pig, showed good preservation of argentaffin and serotonin cells up to 6 hours with a progressive marked reduction in the following hours. 3 The bowel perfused and preserved ipothermically with 4°C Wisconsin solution up to 36 hours, showed normal argentaffin cells up to 20 hours, after which time these cells were numerically reduced and degranulated. On the basis of these previous observations, we decided to analyze the modifications of enteroendocrine cells in a pig model of orthotopic liver-small bowel allotransplantation with different preservation/reperfusion times.

Identification of 5-HT Receptor Sub-types in a Homogeneous Population of Presumptive Serotoninergic Neurones

The expression of the known rat 5-HT receptor sub-types has been analyzed in a presumptive serotoninergic cell line derived from the rat raphé nuclei, using reverse transcription followed by polymerase chain reaction. By manipulating the activity of the oncogene (ts-SV40T) product used to immortalize the serotoninergic precursors, it has been possible to compare the expression of the 5-HT receptors in either replicative or differentiating cells. 5-HT1B, 5-HT1D, 5-HT3, 5-HT6 and 5-HT7 receptor gene expression were all observed in the replicating cells. However, under differentiation conditions, expression of all except the 5-HT1B receptor was lost. Only one novel amplification product appeared during early differentiation, in the 5-HT2B lane; its smaller than expected size was suggestive of a previously undescribed alternate splicing of the mRNA in brain. The curtailment of 5-HT receptor expression in differentiating neurones in vitro may reflect the normal ongoing restriction in the phenotypic potential during embryogenesis in vivo.The serotonin cells, therefore, constitute a pristine cell line in which to study the receptor pharmacology of one or more 5-HT receptor sub-types in isolation. © 1997 Elsevier Science Ltd.

Constitutive expression of calmodulin-binding phosphoprotein GAP-43 in rat serotonergic and noradrenergic cell groups which project to the spinal cord.

In situ hybridization was combined with serotonin (5-hydroxytryptamine, 5-HT) or tyrosine hydroxylase immunocytochemistry and with Fluoro-Gold retrograde labeling of bulbo-spinal pathways in order to investigate the expression of GAP-43 mRNA in monoamine cell groups of the adult rat brain stem. Consistent with previous reports, GAP-43 mRNA was observed in serotonin and dopamine cell groups in the pons. In addition, GAP-43 expressing cells were observed in all the major monoamine cell groups in the medulla. Thus the B1, B2 and B3 serotonin cell groups all showed high GAP-43 expression in all contained many GAP-43 expressing serotonin cells with spinal cord projections. The A1, A2, A5 and A6 noradrenaline cell groups also showed high GAP-43 expression, although cells with spinal cord projections were largely restricted to the A5 group and A6 subcoeruleus region. In all areas, GAP-43 expressing cells with spinal cord projections were also observed which were not serotonergic or noradrenergic.

Pancreatic-duodenal homeobox 1 -role in gastric endocrine patterning

The gastrointestinal tract is subdivided into regions with different roles in digestion and absorption. How this patterning is established is unknown. We now report that the pancreatic-duodenal homeobox 1 gene (pdx1) is also expressed in cells of the distal stomach. Positive cells include subpopulations of the three main endocrine (gastrin, somatostatin and serotonin) cell types of this region. Pdx1 deficient mice were virtually devoid of gastrin cells, had normal numbers of somatostatin cells and increased numbers of serotonin cells. Pdx1 is thus important for development of the gastrin cells of the antropyloric mucosa of the stomach and probably acts by controlling the fate of gastrin/serotonin precursor cells.

Immunohistochemical examination of intraspinal serotonin neurons and fibers in the chicken lumbar spinal cord and coexistence with Leu-enkephalin.

Intraspinal serotonin-positive cells and fibers were examined in the chicken lumbar spinal cord following removal of descending serotonin fibers by spinal transection. Co-localization of Leu-enkephalin immunoreactivity in intraspinal serotonin cells was also examined using a double immunofluorescence labeling technique. By one or two weeks after spinal transection, virtually all supraspinal serotonin fibers were eliminated. Intraspinal serotonin cells were located ventral or ventrolateral to the central canal corresponding to laminae VII, VIII, and IX, and the anterior funiculus. Intraspinal serotonin cells sent fibers to (1) the pia mater on the ventral or ventrolateral surface of the spinal cord; (2) vessels in the spinal cord; (3) sympathetic preganglionic column of Terni; (4) other intraspinal serotonin neurons; (5) the central canal. Some 30%-50% of the intraspinal serotonin cells co-localized with Leu-enkephalin. Intraspinal serotonin fibers co-containing Leu-enkephalin were observed in the pia mater located on the most lateral surface of the spinal cord.

Effects of tetrahydroaminoacridine and nicotine in nucleus basalis and serotonin-lesioned rats

The present study was designed to investigate the hypothesis that concurrent degeneration of serotonin and acetylcholine cells may decrease the therapeutic effects of cholinergic drugs on cognitive functioning in Alzheimer dementia. Therefore, we compared the effects of pretraining injections of a cholinesterase inhibitor, tetrahydroaminoacridine (1, 3 and 5 mg/kg i.p.), and nicotine (0.03, 0.1 and 0.3 mg/kg i.p.) on spatial navigation (water maze) and passive avoidance in nucleus basalis- and nucleus basalis + p-chlorophenylalanine-lesioned rats. Nicotine (0.1 and 0.3 mg/kg) promoted passive avoidance performance of nucleus basalis-lesioned rats, but nicotine did not improve performance of combined-lesioned rats. Tetrahydroaminoacridine (3 mg/kg) facilitated passive avoidance performance of nucleus basalis- and combined-lesioned rats. However, tetrahydroaminoacridine-treated nucleus basalis + p-chlorophenylalanine-lesioned rats were not performing better than vehicle-treated nucleus basalis-lesioned rats. Spatial navigation of nucleus basalis and nucleus basalis + p-chlorophenylalanine-lesioned rats was slightly impaired during the first training day and tetrahydroaminoacridine 3 mg/kg restored the performance of combined-lesioned rats. Combined-lesioned rats performed as well as the controls during the other training days. The present results suggest that, in Alzheimer's disease, combined degeneration of nucleus basalis cholinergic and brainstem serotonergic cells decreases the therapeutic effect of nicotine, but not that of tetrahydroaminoacridine.

Tryptophan hydroxylase can be present in mast cells and nerve fibers of the rat dura mater but only mast cells contain serotonin

Tryptophan hydroxylase-immunopositive (TPH-I) but not serotonin-I nerve fibers were observed in the rat dura mater. This tissue also contained numerous serotonin and TPH-I mast cells. The TPH appeared to be located in granules and/or enclosed in a juxta-nuclear organite. Westernblots showed that the TPH located in the dura mater is similar to the TPH of pineal gland but different from raphe TPH. According to the animal, both nerve fiber and mast cell TPH immunoreactivity was highly variable in intensity and in number of labelled elements. This variability might be due to the complex regulatory mechanisms of TPH as indicated by the presence of two types of mast cells.

Distribution of serotonin cells projecting to the pontomedullary reticular formation in the cat

Using immunohistochemistry and retrograde transport techniques, this study demonstrates that serotonin (5-HT) cells in the dorsal tegmental gray of the pons at rostrocaudal from P1 to P6 levels are sources of bilateral serotonergic projections to the gigantocellular tegmental field of the medial pontine and medullary reticular formation in the cat. The cells observed were a relatively homogeneous population of small (30 μm) to medium-sized (40 μm) cells, oval, fusiform or spindle-shaped. Rostrally and caudally located 5-HT cells in the dorsal tegmental gray tended to project to the pontine and medullary reticular formation respectively. The dendrites of these cells spread in a plane perpendicular to the long axis of the brainstem. Thin 5-HT fibers with a series of fine varicosities (0.5–1.0 μm in diameter) surrounded reticular neurons, and some of these varicosities closely apposed the proximal dendrites and somata of the reticular neurons. Both large (50 μm) and small (20–30 μm) reticular neurons were uniformly innervated by 5-HT fibers. As revealed in the present study, the serotonergic system, together with the monoaminergic and cholinergic systems, seems to be involved in behavioral state regulation.