Number, volume and size distribution of nucleoli in rat neurosecretory cells with suppressed and stimulated secretion

The nucleoli in the neurons of the supraoptic nucleus were investigated in the rat. This was performed after the secretion of vasopressin had been suppressed by water load, after the secretion had been stimulated by water deprival and in normal rats which had water ad libitum. The cells with high secretory activity had large single nucleoli. The cells with low secretory activity had less nucleolar volume, and a substantial part of the cells contained two nucleoli.     

Electron microscopy and morphometry of nucleoli in rat neurosecretory cells with stimulated and suppressed secretion

The nucleoli in the neurons of the supraoptic nucleus in the rat were analyzed by electron microscopy and morphometry after the secretion of vasopressin had been fully suppressed by water load, after the secretion had been stimulated by water deprivation and in normal rats which had water ad libitum. Suppression of the secretion increased the proportion of fibrillar centres in the nucleoli 2-fold. Stimulation of the secretion increased the proportion of the granular component by 22%. The overall nucleolar organization did not change very much with the secretory activity. The results show that an increased proportion of fibrillar centres in nucleoli is an indicator of decreased secretory activity and, moreover, that an increased volume of the nucleolar fibrillar and granular components per cell indicates an increased secretory activity.     

Morphometric electron-microscopic investigation of the neurons of the supraoptic nucleus in water-loaded, normal and water-deprived rats

The neurons of the supraoptic nucleus (SON) in the rat have been analysed by electron microscopy and morphometry, when the secretion of the antidiuretic hormone was fully suppressed by water loading. The water was supplied through a catheter inserted in the external jugular vein for 1.5, 2.5 and 24 h, respectively. The SON was also examined in normal rats and in rats that had been deprived of water for 72 h. The rats were fixed through chronically implanted catheters, so that at the time of fixation, the animal was uninfluenced by anaesthesia and surgery. The morphology of the granular endoplasmic reticulum and the Golgi complex showed that the water load suppressed the synthetic activity and the water deprivation stimulated it. The total volumes of the vasopressin-containing neurosecretory granules (NG) were 1.6, 2.8 and 5.0 X 10(4) micron3 after a 24-hour water load, in the normal state and after 72-hour water deprivation, respectively. In steady states there was a positive correlation between the secretory activity and the content of NG in the perikaryon.     

The secretory activity of rat nasal glands and the effect of cholinergic drugs

Summary The secretory behaviour of rat nasal glands, under normal conditions and after the application of cholinergic drugs, has been studied using morphological and radiobiochemical techniques.Autoradiography and electrophoresis provide evidence for the selective incorporation of ³H-arginine into the glycoprotein containing fraction of the nasal glandular secretion. Radiobiochemical experiments show that labelled arginine is rapidly incorporated into the acinar cells of unstimulated glands, although it takes approximately 4 h before the labelled secretory proteins leave the cells. The secretion of proteins is stimulated by the parasympathetic agonist pilocarpine, whose main action is to promote discharge. Histological sections show a depletion of secretory granules after pilocarpine treatment. The cholinergic antagonist atropine inhibits the secretion; the acinar cells are completely filled with secretory granules following this treatment. The time course of the events following atropine administration suggests that there is no feed-back system controlling glycoprotein synthesis.The techniques employed here therefore appear to be useful for studying the effects of drugs that interfere with the secretory activity of the nasal glands.     

The Root Cap as a Test System for the Evaluation of Golgi Inhibitors: II. EFFECT OF POTENTIAL INHIBITORS ON SLIME DROPLET FORMATION AND STRUCTURE OF THE SECRETORY SYSTEM

The effects of four potential inhibitors of dictyosome activityon the root cap secretory system were monitored by visual estimationof slime droplet reformation rates and by quantitative microscopyof the secretory cells. Only monensin was found to affect bothdroplet reformation and cell structure. While some of our structuralobservations on the effects of this drug, such as swelling ofvesicles and dictyosome cisternae, agreed with those made previously,others did not. We are able to confirm a real increase in vesiclenumber, in addition to the numerical increase in vesicle profilesthat follows from an increase in vesicle size. Formation ofcup-shaped dictyosomes and separation of cisternae were foundto be just as prevalent in the normal and in the solvent controls,especially when fixed with permanganate. Scopoletin, tunicamycin and 2, 6-dichlorobenzonitrile all affecteddroplet formation but had no significant effect on cell structure.It is suggested that these chemicals were affecting water flowinto the slime droplet, rather than directly inhibiting Golgi-activityor release of carbohydrates by the secretory vesicles. The problems of using the root cap system for the identificationof specific Golgi inhibitors are discussed. Key words: Maize, Root cap secretion, Golgi activity inhibitors, Dichlorobenzonitrile, Monensin, Scopoletin, Tunicamycin     

Secretion of the intercalated cells of the fibrillar epithelium of the mucous membrane of the maxillary sinus]

Cytological and histochemical methods were used in order to establish that the intercalated cells of the ciliated epithelium of the rabbit's maxillary sinus mucosa performed gland function giving secretion formed in the shape of granules and represented by neutral mucopolysaccharides. Analysis of the morphology of intercalated cells, the amount of secretion in them, the shape and disposition of the Golgi apparatus and the nucleus allowed establishing 4 successive phases in their secretory cycle: the phase of synthesis, accumulation, aquosity and sectering. Under normal conditions of life the secretory activity of intercalated cells was asynchronous. In experiments with synchronization of the secretory activity by administration of pilocarpine it was established that the duration of the intercalated cells secretory cycle was 16-18 hours. Active participation of alkali phosphatase in the synthesis of secretion in the ciliated epithelium was noted.     

Persistent epithelial dysfunction and bacterial translocation after resolution of intestinal inflammation

Epithelial secretion may play an important role in reducing bacterial colonization and translocation in intestine. If so, secretory dysfunction could result in increased susceptibility to infection and inflammation. We investigated whether long-term colonic secretory dysfunction occurs after a bout of colitis and if this is accompanied by an increase in bacterial colonization and translocation. Rats were studied 6 wk after induction of colitis with trinitrobenzene sulfonic acid when inflammation had completely resolved, and epithelial permeability was normal. Intestinal loops were stimulated with either Clostridium difficile toxin A or a phosphodiesterase inhibitor. In vitro, colonic tissue from previously sensitized rats was exposed to antigen (ovalbumin). Secretory responses to all three stimuli were suppressed in rats that had previously had colitis. These rats exhibited increased (16-fold) numbers of colonic aerobic bacteria and increased (>3-fold) bacterial translocation, similar to results in rats studied after resolution of enteritis. Postcolitis bacterial translocation was prevented by daily treatment with an inhibitor of inducible nitric oxide synthase. This study demonstrates that intestinal inflammation results in prolonged impairment of colonic epithelial secretion, which may contribute to increases in bacterial load and bacterial translocation. Epithelial dysfunction of this type could underlie an increased propensity for further bouts of inflammation, a hallmark of diseases such as inflammatory bowel disease.     

Morphometric electron-microscopic investigation of the neuronal processes in the neurohypophysis in water-loaded, normal and water-deprived rats

The neuronal processes in the neurohypophysis of the rat were analyzed by electron microscopy and morphometry after the secretion of antidiuretic hormone had been fully suppressed by water load. The water was supplied through a catheter inserted in the external jugular vein for 1.5, 2.5 and 24 h, respectively. The neurohypophysis was also examined in normal rats and rats that had been water-deprived for 72 h. The rats were fixed through chronically implanted catheters, so that at the time of fixation the animal was uninfluenced by anaesthesia and surgery. Water load increased the density of the neurosecretory granules in the endings in the zone nearest the basal lamina of the pericapillary space. The interpretation of this is that water load fills up the readily releasable pool of the neurosecretory granules. Water-deprival increased the density of dispersed microvesicles in the endings, especially in the zone near the basal lamina, and it is suggested that the dispersed microvesicles are involved in membrane recapture.     

Involvement of hypothalamic dopamine in the regulation of prolactin secretion

The neuroendocrine control of prolactin (PRL) secretion is known to be a multifactorial process, but dopamine (DA) secreted by the tuberoinfundibular dopaminergic (TIDA) neurons of the hypothalamus is believed to exert a predominant inhibitory control on the secretion of PRL. The secretory activity of the TIDA neurons, including the rate of biosynthesis of DA and the rate of release of the neurohormone into hypophysial portal blood, can be readily evaluated in the rat. In most conditions in which an altered secretion of PRL has been documented, an altered secretory activity of the TIDA neurons has been found. When an acute reduction in the secretion of DA is observed, an increased secretion of PRL is associated, with an inverse relationship between DA and PRL concentrations in hypophysial portal and systemic blood, respectively. However, the secretion of PRL can be regulated by PRL itself through stimulation of the secretory activity of the TIDA neurons, and consequently hyperprolactinemia can be observed concomitantly with a sustained high secretion of DA, as seen after treatment with estrogen. The short loop feedback of PRL secretion seems to be impaired in the aging rat, since a sustained reduced hypothalamic secretion of DA is observed in spite of long-term hyperprolactinemia.     

Electron microscopy of two types of gonadotrophs in the anterior pituitary glands of persistent estrous and diestrous rats

Summary Persistent estrus and diestrus was produced in rats by the administration of estrone for either 5 days or 30 days, respectively, immediately after birth. Female rats without any treatment were used for control. After these rats grew up, the anterior pituitaries were examined by electron microscopy. The identification criteria for two types of gonadotrophs, FSH-and LH-cells, proposed by Barnes were adopted. In the persistent estrous rats, FSH-gonadotrophs were almost normal, but LH-gonadotrophs were filled with an abundance of secretory granules which were probably suppressed in discharge. On the other hand, in the persistent diestrous rats, FSH-cells were few in number and strongly atrophic, containing a few secretory granules, while LH-cells were almost normal or rather slightly activated. These electron microscopic findings well coincide with the results of light microscopy of ovaries, which suggested that in the persistent estrous rats FSH secretion might be almost normal but the secretion of LH might be inhibited, while in the persistent diestrous rats FSH secretion might be almost totally abolished but LH might be moderately secreted. From these findings, identification of FSH-and LH-gonadotrophs in the anterior pituitary of the rat well coincides with that proposed by Barnes in mice.     

Effect of N-linked glycosylation on hepatic lipase activity

Hepatic lipase (HL) is a secretory protein synthesized in hepatocytes and bound to liver endothelium. Previous studies have suggested that HL N-linked glycans are required for catalytic activity. To directly test this hypothesis, Xenopus laevis oocytes were used to express native rat HL or HL lacking one or both N-linked glycosylation sites. The expressed and secreted native HL had an apparent molecular mass of 53 kDa, consistent with purified rat liver HL. The mutant lacking both glycosylation sites, while poorly secreted, had an apparent molecular mass of 48 kDa, the same size observed for HL after enzymatic removal of N-linked oligosaccharides. Mutants lacking one of the two sites were intermediate in size and showed reduced secretion. Each of these expressed and secreted proteins had full catalytic activity that was inhibited by antisera to rat HL. Thus, N-linked glycosylation of rat HL, while important to lipase secretion, is not essential for the expression of lipase activity.     

Enterocyte chloride and water secretion into the small intestine after enterotoxin challenge: Unifying hypothesis or intellectual dead end?

Many forms of diarrhoeal disease, particularly so called “secretory” diarrhoeal disease are thought to arise by the active secretion of chloride ion from the enterocytes, creating an osmotic gradient for fluid movement into the small intestinal lumen. This model implies that normally occurring intestinal secretion is catastrophically enhanced by bacterial enterotoxins. This review advocates that neither normal nor abnormal intestinal secretion from the enterocytes occurs and that no competent proof for chloride secretion exists. Prior to 1970, the physiological evidence failed to support the concept of the formation of intestinal juice as a normal intestinal event. support the concept of the formation of intestinal juice as a normal intestinal event. The concept was later revived to explain the high rate of fluid entry into the lumen after exposure to cholera toxin. Much evidence has been advanced for the chloride secretion hypothesis, the dominant secretory paradigm after 1974, but is the evidence sufficiently compelling for it to be regarded as proving the chloride secretory model? The evidence falls into four categories and a fifth conjectural argument that proposes that an abnormal chloride ion channel in cystic fibrotic sufferers confers a natural selective advantage by preventing diarrhoeal disease. Secretion is putatively demonstrated by 1) showing that mass transfer of fluid is into the lumen (secretion) and not merely a failure to transport out of the lumen (failed absorption). Support is offered by 2) chloride ion flux measurementsin vitro in Ussing chambers and by 3) shortcircuit current measurements that are consistent with and purport to show chloride ion movement into the lumen. In addition, 4) pharmacological agents are identified that affect short-circuit current and these are assumed to be anti-secretory, consistent with the biochemical mechanism for secretion, confirmed wherever possible by mouse knock-out models. Finally, the proxy methods used to study water movement such as elevated short-circuit current measurements show these to be absent in cystic fibrotic patients. The enterocyte secretion hypothesis is challenged here on the basis of an examination of the methods used to show secretion, particularly after exposing the small intestine to heat stable enterotoxin (STa) fromE. coli. STa is thought to be secretory because fluid entry into the lumen is claimed, enhanced isotopic flux of chloride ion towards the lumen occurs, an increase in short-circuit current is found, preventable by various drugs that are deemed likely to be anti-secretory and also because the short-circuit current changes after STa are not seen in cystic fibrotic patients. Using volume recoveryin vivo, STa is found not to be secretory but only anti-absorptive. Hence, other techniques used to show secretion are not fit for that purpose. If STa is identified as secretory and yet no secretion occurs, how reliable is the evidence for other toxins being secretory when these methods are used? This review concludes that chloride ion secretion is unproven. A review of the literature indicates that secretion occurs not because epithelial cells actively pump water but by interdiction of fluid absorption, increased conductivity through tight junctions and an increased hydrostatic driving force through elevated capillary pressure. The exclusive focus on chloride secretion may explain the failure to develop antisecretory drugs over the last three decades.     

Adaptation of the 24-h growth hormone profile to a state of sleep debt

In normal men, the majority of GH secretion occurs in a single large postsleep onset pulse that is suppressed during total sleep deprivation. We examined the impact of semichronic partial sleep loss, a highly prevalent condition, on the 24-h growth hormone profile. Eleven young men were studied after six nights of restricted bedtimes (0100-0500) and after 7 nights of extended bedtimes (2100-0900). Slow-wave sleep (SWS) was estimated as the duration of stages III and IV. Slow-wave activity (SWA) was calculated as electroencephalogram power density in the 0.5- to 3-Hz frequency range. During the state of sleep debt, the GH secretory pattern was biphasic, with both a presleep onset "circadian" pulse and a postsleep onset pulse. Postsleep onset GH secretion was negatively related to presleep onset secretion and tended to be positively correlated with the amount of concomitant SWA. When sleep was restricted, both SWS and SWA were increased during early sleep. Unexpectedly, the increase in SWA affected the second, rather than the first, SWA cycle, suggesting that presleep onset GH secretion may have limited SWA in the first cycle, possibly via an inhibition of central GH-releasing hormone activity. Thus neither the GH profile nor the distribution of SWA conformed with predictions from acute sleep deprivation studies, indicating that adaptation mechanisms are operative during chronic partial sleep loss.     

Biliary secretion in the rat after partial hepatectomy

The secretory efficiency of the liver increased in rats at 12 hr after partial hepatectomy. The secretory efficiency was seen to decrease at 24 hr after partial hepatectomy and increased again at 2-4 days following liver resection. These changes would correspond to the evolution of the hepatocyte proliferative process. The secretion of bile acids expressed per 100 g of body weight had returned to normal at 16 days after partial hepatectomy, although choleresis and the secretion of inorganic electrolytes remained lowered.     

Alterations in albumin secretion and total protein synthesis in livers of thyroidectomized rats.

Perfused rat livers and isolated rat hepatocytes exhibited a 50% decrease in the secretion of both albumin and total secretory proteins after thyroidectomy. In contrast, synthesis of non-secretory proteins was decreased by only 20% from the rates observed in liver preparations from euthyroid rats. These observations suggested a disproportionate effect of thyroidectomy on the synthesis of secretory proteins compared with non-secretory proteins. Disproportionate decreases in the synthesis of albumin in other endocrine-deficient states such as hypophysectomy and diabetes had previously been shown to be associated with decreases of similar magnitude in the relative abundance of albumin-mRNA sequences. In contrast, thyroidectomy did not affect the activity or amount of albumin mRNA in total liver poly(A)-containing RNA when assayed by cell-free translation and by hybridization with complementary DNA, respectively. Furthermore, labelling experiments in vivo demonstrated that albumin synthesis represented 12.9 +/- 0.5% and 12.4 +/- 0.4% of total protein synthesis in livers of thyroidectomized and euthyroid rats respectively. Therefore the fall in secretion of albumin and total secretory protein after thyroidectomy did not appear to be a reflection of disproportionate decreases in the synthesis of these proteins. Instead, defects in steps involved in the post-synthetic processing and secretion of albumin are suggested. A number of comparisons, including ribosome half-transit times, the size distributions of total and albumin-synthesizing polyribosomes, and the fraction of RNA present as inactive ribosomes, provided evidence that the overall decrease in protein synthesis after thyroidectomy was not due to generalized alterations in translational processes. Instead, the decrease in total protein synthesis appeared to reflect the RNA content of the liver, which fell in proportion to th decrease in protein synthesis.     

Recovery of rat mast cells after secretion: a morphometric study

Granule reconstitution in rat peritoneal mast cells following massive secretion was studied by morphometric techniques. Immediately following secretion, the earliest identifiable mast cells showed a substantial decrease in cell volume associated with granule loss. Cell volume then increased almost to the original level over a period of a month. The size of the Golgi apparatus increased markedly in the week following secretion and then returned to its original size. The total volume of granules increased slowly after the secretory depletion and by 34 days had not returned to the original value although the number of granules had recovered fully. The reconstitution of mast cells after secretion is a prolonged process with several phases resulting in mast cells of varying appearance and content. This heterogeneity generated by reconstitution post secretion must be considered in studies of populations of mast cells in vivo.     

Changes in chloride secretion rate and vascular perfusion in the rectal gland of the European lesser-spotted dogfish in response to environmental and hormonal stimuli

Chloride secretion rates of rectal glands taken from the European lesser-spotted dogfish Scyliorhinus canicula adapting to 70% and 120% sea water (SW) were significantly greater and less than, respectively, those in the control 100% SW group. C-type natriuretic peptide (CNP) significantly increased chloride secretion rates above basal values in 100% SW although angiotenisn II (ANG II) had no effect. Perfusion of the secretory epithelia in rectal glands from 70% SW lesser-spotted dogfish was significantly higher than in rectal glands from 100% and 120% SW lesser-spotted dogfish. Perfusion of rectal glands with ANG II had no effect on perfusion of the secretory epithelia, although CNP perfusion induced significantly greater perfusion of the secretory epithelia than all other treatments. It remains to be determined if a reduction in environmental salinity induces an increase in plasma concentration of CNP and hence an increase in rectal gland activity.     

Localization of acid phosphatase activity in collagen-secreting and collagen-resorbing fibroblasts

The ultrastructural localization of acid phosphatase (ACPase) activity was examined in cultured human gingival fibroblasts in the formative and resorptive phases. In the collagen-secreting fibroblasts, weak ACPase activity was demonstrated in the lysosomes, inner Golgi cisternae, and condensing vacuoles, and none was found in the Golgi-associated endoplasmic reticulum-lysosome system (GERL), presecretory granules, or secretory granules. On the contrary, collagen phagocytosis induced strong ACPase activity in the GERL, which was in addition to the weaker activity found in the same sites as those in the collagen-secreting cells. At the same time, collagen secretion was suppressed, and dense elongated secretory bodies associated with ACPase activity accumulated within the cells. When collagen fibrils had been interiorized in whole or in part within the phagosomes, primary lysosomes derived from the Golgi-GERL complex then fused with them to form phagolysosomes. Collagen degradation occurred within these bodies. The observations indicate significant differences in ACPase activity used as a marker for lysosomal enzyme activities in the different functional phases of fibroblasts. These results suggest that fibroblasts work only one way at a given time, viz., collagen synthesis or collagen degradation.     

The ultrastructure of A, B and D pancreatic cells in normal and in diabetic ducks

Ultrastructurally and immunocytochemically identified A, B and D cells are highly concentrated in the splenic bulb of the duck pancreas. Ultrastructural features of normal A, B and D cells are similar in the duck and in other species so far studied. However, normal D cells present a striking characteristic, i.e. apical accumulation of dense bodies, which seem to derive from multivesicular bodies and are probably involved in a catabolic regulatory process. Subtotal pancreatectomy in the duck, leaving the splenic bulb and inducing transient diabetes, produces strong secretory stimulation of A and B cells, as indicated by the development of the rough endoplasmic reticulum and the Golgi apparatus and transient degranulation, more marked in B cells. Numerous B cells with degenerative aspects, observed after 12 days, seem to be exhausted following prolonged hyperstimulation: this could explain why diabetes reappears in some cases. In contrast, in D cells, functional inhibition after surgery is suggested by a dramatic increase in the number and size of the dense bodies, associated with a marked decrease in secretory vesicle storage. The morphological data correlate well with the previously reported evolution of plasma and pancreatic hormone concentration after surgery, and suggest that the normal inhibitory control of glucagon and insulin secretion by the local release of somatostatin might be reduced or suppressed during transient diabetes in subtotally depancreatized ducks.     

Ultrastructural changes in the secretion granules of the hypopharyngeal glands of the honeybee infected by Nosema apis and after treatment with fumagillin

Electron-dense secretion granules were numerous in the hypopharyngeal glands of healthy honeybees. In the hyopharyngeal glands of honeybees infected by Nosema apis, these granules appeared to have increased in size and lost their electron density, possessing a core area that consisted of numerous smaller granules, and a slightly electron dense fringe area, which in some cases possessed a crystalline structure. In the hypopharyngeal glands of infected honeybees after treatment with fumagillin, the secretion granules were also large and slightly electron dense, but the granular content was homogenous. Some of these granules were also partially crystallized. These ultrastructural changes in the secretion granules of diseased and fumagillin treated bees, is probably associated with a change in secretory activity of the glands.