Influence of lactation on the prolactin secreting cells of the hypophysis of impala (Aepyceros melampus): an immunocytochemical and computer image analysis study.

Acute stress in the course of wildlife management has been intensively investigated. Chronic stress, on the contrary, has not been researched at all, probably due to the difficulty in measuring it as a result of the overriding effect of the physiological response to the restraining of wild animals. It was therefore decided to evaluate the use of immunocytochemistry, combined with computer image analysis to try and determine the magnitude of the structural changes of various hormone-secreting cells of the hypophysis. Since it was a pilot study to determine whether the combination of immunocytochemistry with computer image analysis could be of value to distinguish between two normally diverse groups, it was decided to compare the relative activity of prolactin secreting cells of lactating and non-lactating impala ewes. After transforming the prolactin immunoreactive area data by log10 to fall inside the parameters for kurtosis and skewness, a significant difference (P < 0.05, 5-% level, 2-tail) with the parametric t-test could be shown between the mean prolactin immunoreactive area of lactating (3.0751 micron 2) and non-lactating (3.0467 micron 2) ewes. However, the Pearson product moment (r = 0.03) showed that this difference may not be important for all practical reasons. This may be due to either sampling errors or limitations of computer image analysis, as it was often difficult to distinguish individual prolactin immunoreactive areas. Furthermore, a significant increase in the total prolactin immunoreactive areas of lactating ewes was also established. This technique, however, could distinguish between the hypophysis of lactating and non-lactating impala ewes, and with further refinement could be a useful tool in determining chronic stress in wildlife populations.

Distribution of endocrine cells in the gut of the impala (Aepyceros melampus).

Immunocytochemical methods were employed to demonstrate endocrine cells, containing peptides and serotonin, in the gut of the impala. Cells immunoreactive to serotonin, somatostatin, gastrin, cholecystokinin, glucagon, neurotensin, secretin, glucose-dependent insulinotropic peptide and motilin were detected. Antisera raised to substance P and pancreatic polypeptide failed to stain any cells. The distribution of these peptide-containing cells is more in line with the situation in sheep than other ruminants. In contrast, the distribution and abundance of serotonin cells in the gut of the impala parallels the situation seen in game herbivores.

Venom of a six-eyed crab spider, Sicarius testaceus (Purcell, 1908), causes necrotic and haemorrhagic lesions in the rabbit.

By employing haematological, histological and isotope studies on rabbit, it was shown that envenomation by the South African crab spider, Sicarius testaceus, results in tissue necrosis and a marked increase in the permeability of the blood vessels in the vicinity of envenomation. The venom also acts systemically, which results in petechial bleeding of some viscera, thrombocytopaenia and elevated fibrinogen levels. Rabbit envenomation by this species results in a dramatic drop of the platelet count, without haemolytic anaemia. Should these parameters be the same in humans, they may serve as a yardstick to distinguish presumptive S. testaceus envenomation from that of Loxosceles.

The endocrine pancreas of the Cape fur seal, Arctocephalus pusillus (Schreber, 1776): an immunocytochemical study.

The indirect peroxidase method was employed to study the endocrine pancreas of the Cape fur seal. Immunoreactivity to insulin was confined to the cores of the islets and the insulin cells were more abundant than the other endocrine cell types, which occurred mainly in the mantles of the islets. Of these, glucagon cells were the most numerous, followed by somatostatin and pancreatic polypeptide (PP) cells. The latter were observed in the mantles of the islets and scattered in the exocrine tissue of the duodenal lobe. The marked variation in the shape and the distribution of the endocrine cells in the mantles of the islets seen in the pancreas of the seal, seems to be typical of carnivorous species like the cat and dog.

The intestine and endocrine pancreas of the African elephant: a histological immunocytochemical and immunofluorescence study.

Histological, immunocytochemical and immunofluorescence methods were employed to study the intestine and endocrine pancreas of the elephant. The histological findings were in line with those in monogastric mammals. In the mucosa of intestine, endocrine cells were immunoreactive to somatostatin, gastrin, CCK, GIP, secretin, motilin, glucagon and NPY. Nerve cells immunoreactive to somatostatin, substance P, VIP, PHI, NPY, bombesin and CGRP were detected. No immunoreactivity to neurotensin was observed, islets of the pancreas had insulin cells in their cores and glucagon and somatostatin cells in their mantles. The antisera employed failed to demonstrate PP cells in the pancreas, but NPY-immunoreactive cells were present.

Screening of the venoms of two Parabuthus scorpion species on isolated cardiomyocytes.

Crude, diluted venoms of Parabuthus transvaalicus and P. granulatus were tested on isolated cardiomyocytes. Ventricular myocytes were subjected to hypercontractility tests and whole-cell voltage clamp. Both venoms had a profound influence on the hypercontractility of cardiomyocytes, which was markedly decreased when they were pre-absorbed with commercial antiserum. Whole-cell clamp results showed an increase in the sodium current, and a retardation of the time course of inactivation, implicating the presence of an alpha toxin in both venoms.

Putative cardiotoxicity of the venoms of three mamba species.

The venoms of all mamba species are known to contain potent neurotoxins. Some authors suggest that cardiotoxins from mamba venom are responsible for heart failure in envenomated patients. This study was undertaken to come to a better understanding of the mechanisms and possible effects of mamba venom on the hearts of animals. The venoms of three mamba species namely Dendroaspis polylepis, Dendroaspis angusticeps, and Dendroaspis jamesoni were screened for cardiotoxicity by the cardiomyocyte viability test on cardiomyocytes isolated enzymatically from guinea pig ventricle muscle. This analysis was followed by an electrophysiological evaluation of the effects of venoms (from the Dendroaspis species) on cardiac ion channels by employing the whole-cell clamp procedure. In this study the cardiomyocyte viability test indicates differences among the venoms of the three mamba species. The venom of D. jamesoni seems to be the most potent followed by D. angusticeps and then D. polylepis. The whole-cell clamp results indicate that the venoms have no affinity for cardiac potassium channels but have an inhibitory influence on cardiac L-calcium channels. Although this study provides evidence that mamba venoms have a specific effect on isolated myocytes of guinea pig, it is doubtful that it will have a profound influence on a human heart in case of envenomation.

Immunogold localization and quantification of cellular and subcellular abscisic acid, prior to and during drought stress.

An immunogold labeling procedure and experimental data are presented, which demonstrate that antibodies produced against a bovine serum albumin-abscisic acid conjugate can be used both to characterize the cellular and subcellular localization of abscisic acid (ABA), and to permit quantitative comparisons of this hormone in the subcellular compartments prior to and at times of drought stress. At the control leaf water potential (approximately -0.45 MPa), a quantitatively similar positive labeling pattern was observed in the chloroplasts and apoplast. A twofold drought stress-induced increase in the apoplastic ABA concentration was observed in the drought stressed leaf tissue (i.e., at a leaf water potential of approximately -1.55 MPa), while the ABA concentration in the chloroplasts did not differ from that of the controls. Three histochemical controls and the physiological observations validated the specificity of the procedure. Based on the labeling patterns we observed and literature cited, the validity of the hypothesis that drought stress induces a release of chloroplastic ABA is questioned. We interpreted our results as providing indirect evidence for a drought stress-induced root source origin for the increased apoplastic ABA concentrations.

Histological Studies of Ditylenchus africanus Within Peanut Pods.

Ditylenchus africanus entered the immature pegs and pods of peanut (Arachis hypogaea cv. Sellie) at the peg-connection and subsequently invaded the parenchymatous regions of the hull exocarp and endocarp, and eventually the seed testa. The nematode caused malformations of the cells of infected tissues, cell wall breakage, and cell collapse. The damage appeared to be due to enzymatic activity. In some testae the entire parenchyma region, which aids in protection of the seed, was destroyed. In immature pods, the nematodes moved across the fibrous region of the mesocarp into the hull endocarp. In mature pods, however, the fibrous mesocarp of the hull was lignified and apparently was a barrier to penetration of the inner pod tissues. In late-harvested pods, increased numbers of eggs and anhydrobiotes were found in the hull tissues, and eggs in the seed testa, suggesting the onset of winter survival mechanisms of the nematode.

The oesophagus and stomach of the African elephant: a histological, immunocytochemical and immunofluorescence study.

Histological, immunocytochemical and immunofluorescence methods were employed to study the oesophagus and stomach of the elephant. The histological findings were in line with the situation in monogastric species like swine and man. In the mucosa of the stomach, endocrine cells were immunoreactive to gastrin, somatostatin, chromogranin A and serotonin. Nerve cells immunoreactive to somatostatin, bombesin, VIP, PHI and CGRP were detected in the submucosal and myenteric plexus of the stomach. In the stomach, the absence of glucagon cells and the presence of endocrine cells immunoreactive to PYY, are in contrast to the situation in mammals and need further investigation. Small gastric ulcers were observed in some of the specimens.

A light microscopical study of the intestinal tract of the Nile crocodile (Crocodylus niloticus, Laurenti 1768).

Although the histology of the intestinal tract of Crocodylus niloticus is touched on in overall studies on reptilian intestinal tract, a more comprehensive light microscopical study on this area is lacking. Specimens for histological examination were taken from the duodenum, the jejunum, the ileum and the rectum. The data obtained revealed that the mucosa is thrown into folds and simple, slightly branched tubular intestinal glands. The mucosal folds diminished in height and eventually disappeared upon reaching the ileorectal junction. The epithelium covering the folds and crypts was of the simple columnar type. Clear marginal cells, goblet cells and argentaffin cells were observed throughout the intestinal tract. No Paneth cells were seen in this study. The lamina propria was rich in lymphocytic infiltrations while the muscularis mucosa consisted mainly of an outer longitudinal layer, the inner circular layer being rudimentary or absent. The submucosa was extremely narrow, and the circular and longitudinal layers of the tunica muscularis contained distinct layers of dense fibrous connective tissue. The histology of the intestinal tract of C. niloticus is shown to be in line with the situation in crocodilians and also exhibits a resemblance to that of carnivorous mammals.

Cardiotoxicity of the skin of the red-banded rubber frog, Phrynomerus bifasciatus (Smith 1847).

The skin of the red-banded rubber frog is said to secrete an unidentified toxin. Whole skin extract as well as HPLC fractions were tested for toxicity on viable, isolated cardiomyocytes. The skin extract and one of its fractions were shown to be cardiotoxic. Whole-cell clamping tests demonstrated that both extract and fraction interfered with the function of the potassium channels of the cardiomyocytes and that the toxic substance has an affinity for the IK, channels. Investigations are in progress to isolate and characterize the toxic substance.

Bioactive peptides and serotonin in the gut endocrine cells of the crocodile, Crocodylus niloticus (Laurenti 1768): an immunocytochemical study.

Little is known about peptide-storing endocrine cells in the gut of the Nile crocodile. As in the case of other reptiles, particularly the alligator, a limited range of peptide-storing cells was found in the gut of the crocodile. They were somatostatin, glucagon, gastrin, neurotensin and pancreatic polypeptide. The topographical distribution of cells immunoreactive to somatostatin and gastrin in the gut of the crocodile is comparable to the situation in the alligator. Glucagon and neurotensin immunoreactive cells have a much wider distribution in the gastro-intestinal tract of the crocodile compared to the alligator. Cholecystokinin and bombesin cells previously reported in the small intestine of the alligator were not detected in this study. This is the first report to demonstrate pancreatic polypeptide and serotonin immunoreactivity in the gut of a crocodilian specie.

Light, electron microscopical, and immunocytochemical investigation of the stomach wall of the tigerfish Hydrocynus forskahlii.

The wall of the stomach of the tigerfish is described and compared with that of other vertebrates. Light microscopic and ultrastructural characteristics of the stomach wall correspond to a large extent to those of other vertebrates, although some differences are found. The mucosa contains (1) surface epithelium characterized by narrow columnar cells with abundant mucous granules; (2) gastric glands consisting of pepsinogenic cells of variable height, containing tubulovesicles and bearing microvilli; (3) five granulated cell types located basally in the epithelium (types 1-5); and (4) lamina propria and muscularis mucosae. Connective tissue separating smooth muscle fibers of the muscularis mucosae constitutes a stratum compactum. The submucosa contains a loose connective tissue, a tunica muscularis of inner circular and outer longitudinal layers, and a serosa of mesothelium and subjacent connective tissue. Immunocytochemical tests with antisera to five polypeptides show gastrin/cholecystokinin (CCK), vasoactive intestinal polypeptide (VIP) immunoreactivities in some cells of the gastric glands, and somatostatin in cells lying among epithelial cells lining the gastric luminal surface or gastric pits.

An immunohistochemical study of various peptide-containing endocrine cells and neurones at the equine ileocaecal junction.

The ileocaecal junctions of 5 horses and 2 donkeys were examined by using antisera to the following peptides: somatostatin, glucagon, gastrin, neurotensin, vasoactive intestinal peptide (VIP), peptide histidine isoleucine (PHI), calcitonin gene-related peptide (CGRP), substance P (SP) and neuropeptide Y (NPY). Antisera to somatostatin, neurotensin and NPY demonstrated endocrine cells in the ileal- and caecal parts of the ileocaecal junction, while immunoreactivity for glucagon was demonstrated in endocrine cells of the ileal part only. Nerve cell bodies showing immunoreactivity to SP, VIP, CGRP and PHI were demonstrated in the myenteric and submucosal plexuses and were associated with small blood vessels in the submucosa of all the regions tested. Ramified nerve fibres in the submucosa immunoreactive to SP, VIP, CGRP and PHI extended to the mucosa and to small blood vessels in the submucosa. Nerve fibres showing immunoreactivity to SP, VIP and PHI extended to the circular smooth muscle layer of the ileocaecal junction.

A light microscopic and immunocytochemical study of the gastrointestinal tract of the ostrich (Struthio camelus L.).

Although the histological features and endocrine cells of the gastro-intestinal tract of the chicken have been well studied, little is known about these features of the gut of the ostrich. The present study was undertaken to elucidate the histology and peptide-storing endocrine cells of the ostrich. As a rule the histological features of the gastro-intestinal tract of the ostrich corresponded to that of the fowl. However, certain differences were observed. The superficial proventricular glands were simple, branched tubular glands, while the deep proventricular glands were restricted to a slipper-shaped area and extended into the muscularis mucosae. The gizzard had a variably developed muscularis mucosae, a feature that seems to be unique to the ostrich. The villi of the small intestine were long and branched profusely, forming a labyrinthine surface. No Paneth cells were observed. The mucosa of the ceca and the first part of the rectum was thrown in large circular folds, forming a compressed spiral. Numerous melanocytes were seen in the submucosa and the connective tissue around the bloodvessels of the muscle layers at the tips of the ceca. A well developed subserosa was present throughout the gastro-intestinal tract. Endocrine cells immunoreactive to somatostatin, glucagon, gastrin, bombesin, neurotensin, substance P and pancreatic polypeptide were detected in the gastro-intestinal tract of the ostrich. The topographical distribution of those endocrine cells immunoreactive to glucagon, bombesin, neurotensin and substance P differed from that of the chicken. The results of this investigation inferred that at least one of the gut peptides of the ostrich (secretin) to be structurally different from its counterparts in mammal and chicken. Molecular heterogeneity of somatostatin was observed in endocrine cells situated in the deep ventricular glands of the ostrich.

Developmental profile of chromogranin, hormonal peptides, and 5-hydroxytryptamine in gastrointestinal endocrine cells.

In this immunocytochemical study, we have analyzed the developmental profile and phenotypic expression of the endocrine cell antigens chromogranin, 5-hydroxytryptamine, gastrin/cholecystokinin, cholecystokinin (9-20), somatostatin, somatostatin 28 (1-14), somatostatin cryptic peptide, glucagon, glucagonlike peptides 1 and 2, glicentin, peptide YY, glucose-dependent insulinotropic peptide, secretin, neurotensin, and substance P in human fetal stomach and intestine. All currently identifiable endocrine cell types were detected by 10 wk of gestation. Immunostaining for the endocrine cell marker chromogranin revealed abundant endocrine cells in the earliest specimens (8 wk of gestation) with a relatively higher frequency in both proximal duodenum and distal colon/rectum compared with other areas. Quantification of endocrine cells showed an increase with age that was roughly parallel to the growth of the gut as a whole. These studies show that the diversity of the endocrine component of the gut appears to be established by 10 wk of gestation and that gut activity is preceded by the development of a fully differentiated endocrine component, which may subserve or even initiate the onset of functional maturity.

Ontogeny of peptide- and amine-containing neurones in motor, sensory, and autonomic regions of rat and human spinal cord, dorsal root ganglia, and rat skin.

The developmental patterns of neurofilament triplet proteins, peptide and amine immunoreactivities were compared in motor (ventral spinal cord), sensory (dorsal spinal cord, dorsal root ganglia, epidermis), and autonomic (intermediolateral cell columns, dermis) regions in the rat and human. In the rat, neurofilament triplet proteins first appeared in motoneurones (embryonic day 13). In the youngest human fetuses studied (6 weeks), immunoreactivity was present throughout the spinal cord. Peptides and amines occurred later. Calcitonin gene-related peptide, galanin, somatostatin, neuropeptide Y and its C-flanking peptide (CPON) were the first to appear localized to motoneurones (embryonic days 15-17 rat; fetal weeks 6-14 human). Numbers of immunoreactive motoneurones decreased toward birth, but immunoreactive fibers increased in the ventral horn with enkephalin, thyrotrophin-releasing hormone, and the monoaminergic markers 5-hydroxytryptamine and tyrosine hydroxylase (all presumably of supraspinal origin) the last to appear perinatally. In the dorsal horn, particularly in the rat, a transient expression of substance P-, somatostatin-, and neuropeptide Y/CPON-immunoreactive cells was detected (embryonic days 15-17). A pronounced increase of calcitonin gene-related peptide-, galanin-, somatostatin- and substance P- immunoreactive fibers was found perinatally in both species. This coincided with an increased detection of cells in the dorsal root ganglia containing these peptides and the earliest appearance of calcitonin gene-related peptide-, somatostatin-, and substance P-immunoreactive fibers in the rat epidermis. Few antigens were localized to the intermediolateral cell columns before embryonic day 20 (rat), fetal week 20 (human), with thyrotrophin-releasing hormone-, 5-hydroxytryptamine-, tyrosine hydroxylase-, and vasoactive intestinal polypeptide-immunoreactive nerves appearing perinatally. In the rat dermis, tyrosine hydroxylase-immunoreactive fibers (sympathetic fibers) and fibers immunoreactive for neuropeptide Y/CPON and vasoactive intestinal polypeptide were detected from postnatal day 1. In conclusion, 1) peptide and amine immunoreactivity develops in motor before sensory or autonomic regions, 2) many peptide-containing cells are transient in fetal life, and 3) central terminals of dorsal root ganglion cells express peptides before terminals in the skin.

Viral hepatitis.

The natural history of and biochemical changes in 27 patients with viral hepatitis are outlined. Analysis is made of the symptomatology and physical signs at the time of presentation as well as a comparison between hepatitis B surface antigen-positive and negative patients. In general, hepatitis B surface antigen-positive patients have a more severe clinical illness as well as greater biochemical derangement and a longer duration of disease.