Lamellar Endoplasmic Reticulum Research Articles

Defining the role of endoplasmic reticulum in the initiation of paligenosis

Reprogramming is crucial for cellular renewal in adult organs that lack dedicated stem cells for replacement of cell loss after injury. Because such cell plasticity is likely to be executed by a conserved cellular program, we have begun to identify the conserved cellular-molecular features of the process in which differentiated cells are recruited as progenitors. We have characterized three successive stages by which this occur with differentiated cells: 1) the cells downscale their organelle contents, 2) cells activate a progenitor-like gene network, and 3) cells reenter the cell cycle. We have given this conserved process the name paligenosis. Here, we investigate upstream triggers of paligenosis. Using a high-dose tamoxifen (HD-TAM) injury model to induce paligenosis in zymogenic chief cells of the murine stomach corpus, we observed ultrastructural changes in the endoplasmic reticulum (ER) during paligenosis initiation (e.g., swelling of ER lamellae, liberation of ribosomes from rough ER, and overall loss of ER). This led us to hypothesize that dynamic changes in ER were an upstream event in paligenosis. ER functioning is in part monitored by the Integrated Stress Response (ISR) with the paramount ER stress sensor being PERK, which then autophosphorylates and in turn phosphorylates the α subunit of the eukaryotic initiation factor 2 (eIF2α). Phospho-eIF2α halts global translation while upregulating a specific set of genes to restore homeostasis. Here, we go on to show that HD-TAM activates the ISR in paligenotic gastric chief cells, triggering global attenuation of protein synthesis. However, this “ER stress” ISR pathway must occur in a critical, temporally limited window early in paligenosis, as pharmaceutically-induced prolonged ER stress caused paligenosis to halt in early stages with cells failing to proliferate. Blocking the ISR also halted paligenosis in early stages. Ongoing experiments include the use of the glycosylation-inhibiting drug tunicamycin to determine if inducing the ISR via ER stress is sufficient to initiate paligenosis; and deleting PERK in chief cells during paligenosis using Perk flox/flox ; Mist1 CreERT2 mice. Our results provide initial insights on the dynamic functionality of the ER and its role and necessity during cellular regeneration. This work was supported by funding from the NIH through the National Institute of Diabetes and Digestive and Kidney Diseases and the National Cancer Institute. This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Sanguinarine biosynthesis is associated with the endoplasmic reticulum in cultured opium poppy cells after elicitor treatment.

Three key benzylisoquinoline alkaloid biosynthetic enzymes, (S)-N-methylcoclaurine-3'-hydroxylase (CYP80B1), berberine bridge enzyme (BBE), and codeinone reductase (COR), were localized in cultured opium poppy (Papaver somniferum) cells by sucrose density gradient fractionation and immunogold labeling. CYP80B1 catalyzes the second to last step in the formation of (S)-reticuline, the last common intermediate in sanguinarine and morphine biosynthesis. BBE converts (S)-reticuline to (S)-scoulerine as the first committed step in sanguinarine biosynthesis, and COR catalyzes the penultimate step in the branch pathway leading to morphine. Sanguinarine is an antimicrobial alkaloid that accumulates in the vacuoles of cultured opium poppy cells in response to elicitor treatment, whereas the narcotic analgesic morphine, which is abundant in opium poppy plants, is not produced in cultured cells. CYP80B1 and BBE were rapidly induced to high levels in response to elicitor treatment. By contrast, COR levels were constitutive in the cell cultures, but remained low and were not induced by addition of the elicitor. Western blots performed on protein homogenates from elicitor-treated cells fractionated on a sucrose density gradient showed the cosedimentation of CYP80B1, BBE, and sanguinarine with calreticulin, and COR with glutathione S-transferase. Calreticulin and glutathione S-transferase are markers for the endoplasmic reticulum (ER) and the cytosol, respectively. In response to elicitor treatment, large dilated vesicles rapidly developed from the lamellar ER of control cells and fused with the central vacuole. Immunogold localization supported the association of CYP80B1 and BBE with ER vesicles, and COR with the cytosol in elicitor-treated cells. Our results show that benzylisoquinoline biosynthesis and transport to the vacuole are associated with the ER, which undergoes major ultrastructural modification in response to the elicitor treatment of cultured opium poppy cells.

Heat shock response of warm‐incubated barley aleurone layers

Heat shock suppresses secretory protein synthesis in GA(3)-stimulated barley (Hordeum vulgare cv. Himalaya) aleurone layers by selectively destabilizing their mRNAs and dissociating the stacked rough endoplasmic reticulum (ER) lamellae upon which they are translated. Heat shock also increases phosphatidylcholine (PC) synthesis, and these PC molecules have increased levels of fatty acid saturation. This appears to be adaptive, for aleurone layers maintained at heat shock temperatures for 18 h resynthesize secretory protein mRNAs, rebuild stacked ER lamellae, and resume secretory protein synthesis. In the present study aleurone layers were incubated at warmer than normal pre-heat shock temperatures to determine whether this would favor the formation of heat-resistant ER lamellae that could continue secretory protein synthesis during heat shock. Western blot and SDS-PAGE analyses showed that such treatment did not induce heat shock protein (HSP) synthesis, but it preserved significant secretory protein synthesis during heat shock. Northern hybridizations revealed that levels of mRNAs encoding secretory proteins were several-fold elevated as compared to 25°C preincubated controls, and transmission electron microscopic observations revealed stacked ER lamellae. Thin layer and gas chromatography showed that PC molecules in warm-incubated barley aleurone layers had more fatty acid saturation than did controls. These observations indicate that previous incubation temperature influences both the induction of HSP synthesis and the suppression of normal protein synthesis in the heat shock response. However, we found that it does not affect the temperature at which heat shock becomes lethal.

Slow heating of barley aleurone layers to heat‐shock temperature preserves heat‐shock‐sensitive cellular properties

In barley (Hordeum vulgare L. cv. Himalaya) aleurone layers, heat shock causes the selective suppression of α-amylase synthesis by destabilizing this secretory protein's mRNA. The lamellar stacks of the endoplasmic reticulum (ER), which serve as the site of α-amylase mRNA translation, are dissociated by heat shock, suggesting that heat-shock-induced changes in ER may be important in selectively targeting α-amylase mRNAs for destabilization. We have found that samples maintained at heat-shock temperature (40°C) for 18 h recover the ability to synthesize α-amylase and that the ER membranes in these samples contain membrane phospholipids with enhanced levels of fatty acid saturation. This present study investigated whether gradual warming to 40°C over 3-6 h (ramping) would preserve α-amylase synthesis by permitting ER membrane phospholipid retailoring during the gradual temperature increase. Analyses by sodium dodecyl-sulfate polyacrylamide gel electrophoresis revealed that α-amylase synthesis was markedly increased in ramped samples. Furthermore, northern hybridization analyses and transmission electron microscopy showed that these samples had increased α-amylase mRNA levels and stacks of ER lamellae, respectively. Gas chromatographic analyses of ER membrane phospholipids indicated that the fatty acids of ramped samples were more saturated than their heat-shocked counterparts. These data indicate that heat-induced increases in aleurone ER membrane phospholipid fatty acid saturation may be important in maintaining secretory protein expression at normally nonpermissive heat-shock temperatures.

Changes in calcium concentrations in subcellular compartments of rat submandibular gland acinar cells induced by cholinergic stimulation.

The effects of cholinergic and alpha-adrenergic stimulation on calcium concentrations in rat submandibular gland acinar cells were evaluated at the subcellular level by electron probe X-ray microanalysis, and the subcellular distribution of receptors for inositol 1,4,5-trisphosphate (IP(3)Rs) was investigated by electron microscopic immunohistochemistry. For measurement of calcium concentrations, experiments were carried out both in vivo and in vitro. In in vivo experiments, fragments of glands were removed and slam-frozen 3 min after intraperitoneal injection of an agonist. In in vitro experiments, the gland tissue was enzymatically dispersed, treated with an agonist in the presence or absence of extracellular Ca(2+), and slam-frozen. The frozen specimens were cut into ultrathin cryosections, which were then freeze dried. Spectra were collected from secretory granules, the perinuclear cytoplasm containing endoplasmic reticulum (ER), and the nucleus of the acinar cells. A clear decrease in calcium content in secretory granules was observed after cholinergic stimulation both in vivo and in vitro in the presence or absence of extracellular Ca(2+). In the presence of extracellular Ca(2+), cholinergic stimulation following alpha-adrenergic stimulation reduced the calcium content in secretory granules to the same extent as cholinergic stimulation alone. No significant changes in the calcium content of the perinuclear cytoplasm and the nucleus were noticed after cholinergic stimulation. alpha-adrenergic stimulation did not significantly affect the calcium concentrations in any of the three compartments studied. IP(3)Rs were localized to ER lamellae, but not to secretory granule membranes or to nuclear membranes. Our findings suggest that: (1) in rat submandibular acini, Ca(2+) can be released from secretory granules by exposure to cholinergic, but not alpha-adrenergic, stimuli, (2) there are, however, no IP(3)Rs present in the granular membrane and the trigger for the Ca(2+) release remains under study, and (3) the response to either type of stimulus does not involve changes in the Ca(2+) content of the nucleus or of the perinuclear cytoplasm/ER, despite the presence of IP(3)Rs in the latter structure. This may be the result of limitation in the technique used, which cannot measure changes in the proper compartment of the ER.

COMBINED IMMUNOFLUORESCENCE AND FIELD EMISSION SCANNING ELECTRON MICROSCOPE STUDY OF PLASMA MEMBRANE-ASSOCIATED ORGANELLES IN HIGHLY VACUOLATED SUSPENSOR CELLS OF WHITE SPRUCE SOMATIC EMBRYOS

Highly vacuolated suspensor cells of spruce somatic embryos were examined by immunofluorescence light microscopy using butyl-methyl-methacrylate (BMM) and polyethylene glycol (PEG) embedded sections, transmission electron microscopy (TEM) and field emission scanning electron microscopy (FESEM). The use of PEG embedded embryos provided a rapid method for light microscope detection of antigens before committing to FESEM analysis. BMM embedded specimens provided well preserved suspensor cells for immunofluorescence. FESEM permitted high resolution observation of large areas of the inner surface of the plasma membrane and associated cell organelles. Suspensor cells contained mostly transversely oriented cortical microtubules linked to the plasma membrane and adjacent microtubules by cross- bridges. Light and electron microscopy revealed numerous clathrin coated structures on the plasma membrane. These included flat patches of clathrin, coated pits and coated vesicles. Many coated vesicles were associated with microtubules. Both tubular and lamellar endoplasmic reticulum were observed on the plasma membrane by FESEM.

Heat shock proteins do not provide thermoprotection to normal cellular protein synthesis, α‐amylase mRNA and endoplasmic reticulum lamellae in barley aleurone layers

Heat shock in barley (Hordeum vulgare L. cv. Himalaya) aleurone layers induces the synthesis of heat shock proteins (hsps) and suppresses the synthesis and secretion of α‐amylase, the principal secretory protein. This is accompanied by the destabilization of α‐amylase mRNA and a concomitant dissociation of ER lamellae. In the absence of heat shock α‐amylase mRNA is extremely stable (Belanger et al. 1986. Proc. Natl. Acad. Sci. USA 83: 1354–1358). In most organisms there is a direct correlation between the synthesis of hsps and thermotolerance. The ability of hsps to provide thermoprotection to secretory protein synthesis, α‐amylase mRNA and ER lamellae was analyzed. Sodium dodecyl sulfate‐polyacrylamide gel electrophoresis (SDS‐PAGE) of pulse‐chased, [35S]‐methionine‐labeled proteins revealed that the half‐life of hsps in barley aleurone cells recovering from heat shock was approximately 12 h. Within approximately 6 h, there was a recovery of α‐amylase mRNA and a reformation of ER lamellae. Heat shock protein synthesis was induced by either heat shock (40°C) or arsenite, the cells were allowed to recover for 8 h, then were re‐exposed to heat shock. Results from SDS‐PAGE showed that, despite the presence of hsps, α‐amylase synthesis was suppressed. Northern blot hybridizations showed that α‐amylase mRNA levels were reduced in heat‐shocked tissues. Transmission electron microscopy demonstrated that ER lamellar structures were dissociated. The synthesis of hsps did not enable barley aleurone cells to sustain the synthesis of any proteins at lethal temperature. In contrast, similar conditions established thermotolerance and provided thermoprotection to protein synthesis in germinating barley embryos. Our findings suggest that the aleurone layer does not become thermotolerant following the induction of hsp synthesis.

Heat shock proteins do not provide thermoprotection to normal cellular protein synthesis, alpha-amylase mRNA and endoplasmic reticulum lamellae in barley aleurone layers

Heat shock proteins do not provide thermoprotection to normal cellular protein synthesis, alpha-amylase mRNA and endoplasmic reticulum lamellae in barley aleurone layers

Actomyosin-based motility of endoplasmic reticulum and chloroplasts in Vallisneria mesophyll cells

Intracellular localization and motile behaviour of the endoplasmic reticulum (ER), plastids and mitochondria were studied in living mesophyll cells of Vallisneria using the vital fluorochrome 3,3'-dihexyloxacarbocyanine iodide (DIOC6(3)). In quiescent cells, the ER was composed of a three-dimensional network of tubular and lamellar elements. Chloroplasts were distributed evenly throughout the cell periphery and appeared embedded within the ER network. The ER network was relatively stationary, with the exception of rare motile episodes occurring as movement of tubular ER strands and adjacent areas of the polygonal network in localized areas of the cell. During experimental induction of streaming, most of the lamellar ER elements transformed into tubules and together with the chloroplasts they began to translocate to the anticlinal walls to establish the circular streaming around the circumference of the cell. Microwave-accelerated fixation followed by immunofluorescence revealed an hitherto unknown phase of actin reorganization occurring within the cells and most interestingly at the surface of the chloroplasts during streaming induction. Myosin was localized in an ER-like pattern in quiescent as well as in streaming cells, with bright fluorescent label localized on mitochondria and proplastids. In addition, myosin label appeared on the surface of the chloroplasts, preferentially in streaming mesophyll cells. Motile activities were impeded by the actin-depolymerizing drug cytochalasin D (CD), the thioreagent N-ethylmaleimide (NEM), and thapsigargin, an inhibitor of the ER-Ca(2+)-ATPase. These inhibitors also interfered with the integrity of actin filaments, the intracellular distribution of myosin and calcium-homeostasis, respectively. These effects suggested an obligate association of at least one type of myosin with the membranes of ER and smaller organelles and are consistent with the appearance of another type of myosin on the chloroplast surface upon streaming induction.

Cytological characterization of isolated sperm cells of perennial ryegrass (Lolium perenne L.)

The cytoplasmic content and the distribution of intramembrane particles (IMPs) of the plasma membrane of isolated sperm cells of perennial ryegrass (Lolium perenne L.) have been characterized using flow cytometry, transmission electron microscopy, confocal scanning laser microscopy and freeze-fracture studies. The isolated haploid sperm cells contain a variety of cell organelles with the exception of microtubules. Proplastids and plastids with starch were observed, although only rarely. Vacuoles containing remnants of organelles and stacked lamellae of endoplasmic reticulum with cytoplasmic inclusions were observed frequently, indicating that autophagy takes place. The number of mitochondria varies from 11 to 26 with an average of 17. Generally, the nucleus has a lobed shape and displays various interphasic stages of chromatin condensation. The analysis of the number of mitochondria and the nuclear state did not show evidence of sperm cell dimorphism. The cytological variability observed, could be explained by differences in developmental stages already present in vivo at the moment of isolation. No correlation between the number of mitochondria and the nuclear cross-sectioned area and/or the condensation state of the chromatin could be found. The density of intramembrane particles of the plasma membrane on the exoplasmic fracture face is more than twice that on the protoplasmic fracture face. That is the opposite of what was found for sporophytic cells of perennial ryegrass. These results are discussed in relation to the potential use of these cells for biotechnology and developmental studies.

Heat shock in mechanically wounded carrot root disks causes destabilization of stable secretory protein mRNA and dissociation of endoplasmic reticulum lamellae

In many organisms, the synthesis of heat shock proteins during heat shock is concomitant with the cessation of at least a portion of normal cellular protein synthesis. Heat shocked barley aleurone layers selectively stop the synthesis and secretion of secretory proteins. Exposure to 40°C causes a disruption of endoplasmic reticulum (ER) lamellae, which we have hypothesized leads to the destabilization of otherwise stable mRNA previously associated with ER‐bound polyribosomes. We report here that this was also observed in wounded carrot (Daucus carota L.) root parenchyma tissue which synthesizes and secretes cell wall proteins when mechanically wounded. Nondenaturing cationic polyacrylamide gel electrophoresis of radiolabeled proteins indicated that heat shock caused the cessation of the synthesis and secretion of extensin, a hydroxyproline‐rich cell wall glycoprotein. Northern blot analyses indicated that the mRNA levels for both extensin and another cell wall protein (p33) were rapidly diminished during heat shock. Under nonheat shock conditions extensin mRNA had a half‐life of greater than 4 h, but this appeared to be reduced to less than 30 min during heat shock. There was also a concomitant dissociation of ER lamellae in wounded, heat shocked carrot root tissue, as observed by transmission electron microscopy. These observations indicate that this response may be universal among plant secretory tissues.

Cortical ultrastructure of freeze-substituted protonemata of the mossFunaria hygrometrica

The ultrastructural organization of the cortical cytoplasm has been examined in caulonemata, branches and buds of the mossFunaria hygrometrica, which were prepared by rapid freeze-fixation and freeze-substitution (FS). The same structural components occur in the cortex of all three cell types: microtubules (MTs), endoplasmic reticulum (ER), coated and uncoated vesicles, coated pits, and dictyosomes. However, the configuration and density of the cortical ER varies between the three. Caulonemata have an open, polygonal network of ER associated with long MTs oriented mostly parallel to the length of the cell. Lamellar ER, covered with polysomes, is interspersed in the network. Branches have a more tightly arranged ER network, at places occurring in a thick layer, and occasional polysome-decorated lamellae. MTs, which extend to the tip of the branch, are oriented mainly parallel to the cell's long axis and are associated with the cortical ER. Buds have the tightest ER network, which is frequently arranged in a thick layer. Tubules in the polygonal ER of buds are densely covered with ribosomes, whereas tubules in the ER network of caulonemata and branches range from nearly smooth to moderately rough. Closely-spaced ER lamellae, with many polysomes, occur in some buds. The MTs of buds extend into the apical dome and are associated with the cortical ER, but are more randomly oriented than in caulonemata or branches. Close appositions between the ER and PM are observed in all three cells, but are more frequent in buds.

Membranes and calcium sequestration during spermiogenesis in the cotton seed bug (Dysdercus intermedius: Heteroptera)

By use of osmium ferricyanide (OsFeCN) staining the fate of cytoplasmic membranes was followed during spermiogenesis in the cotton seed bug (Dysdercus intermedius). During early spermiogenesis interzonal lamellae of endoplasmic reticulum become aggregated as a stack of membranes traversing the entire cell body from the nucleus to the cytoplasmic bridge connecting neighbouring spermatids. Cisternae of endoplasmic reticulum ensheath the acroblast from which vesicles of different sizes are pinched off into the cytoplasm. The oxalate method was used to show that acroblast and associated vesicles are calcium-sequestering sites in spermatids. Membrane profiles with dense calcium oxalate precipitate derived from the acroblast form an uninterrupted membranous sheath at the apical side of the nucleus where the proacrosome will be attached. With further development of the spermatids, the vesicles derived from the acroblast also participate in forming a calciumsequestering sheath enveloping the axoneme and the mitochondrial “nebenkern” derivatives.

Physical and Biochemical Changes in the Microsomal Membranes of Tomato Fruit Associated with Acclimation to Chilling

Summary Mature green tomato fruit ( Lycopersicon esculentum L. cv. Caruso) were acclimated to cold temperatures and their physiological properties compared to chilled and ripening fruit. Spin labelling studies on isolated microsomal membranes showed shifts to higher breakpoint temperatures from the initial 11–12 °C in the spin label motion parameter vs. temperature plots for the chilled and ripened fruit membranes; breakpoint temperature for the acclimated fruit shifted upwards initially and decreased to 8 °C after 15 days of acclimation. No changes were observed in the average fluidity of the membranes. Energy of activation (Ea) for ATP hydrolysis at low temperatures (5–17.5 °C) increased for the three treatments; however, after 15 days the Ea for the acclimating membranes was lower than for the chilled and ripening membranes. Over the duration of storage and for fruit from all treatments, changes in the expression of membrane associated proteins were observed. No changes were observed in the relative proportions of membrane phospholipids or fatty acids. Ultrastructural studies indicated microvesiculation of the endoplasmic reticulum (ER) and disruption of the ER lamellae due to chilling. Acclimating cells showed «budding off» of larger membrane vesicles, which may constitute a «retailoring» process responsible for increased chilling tolerance.

In vivo effect of dopamine antagonists on melanocyte-stimulating hormone cells of the goldfish ( Carassius auratus L.) pituitary

Young goldfish were injected with three dopaminergic antagonists, pimozide, sulpiride, and domperidone, for 5 (low dose) and 7 days (higher dose). Cytological and immunocytochemical techniques using anti-α-melanocyte-stimulating hormone (MSH) serum were applied to the pituitary. MSH cells in the three treated groups showed a decrease in immunoreactive cytoplasmic granules, a significant nuclear hypertrophy, and, after 7 days, a cellular enlargment. The nucleolus and the lamellar endoplasmic reticulum were more developed and some mitotic figures occurred. Erythrophores and occasional melanophores were in a stage of maximal dispersion. These changes were not apparent in the solvent-injected controls. The responses to the three blockers of dopaminergic receptors were similar. These data suggest that MSH release seems to be under a dopaminergic inhibitory control in the goldfish. The other cell type of the pars intermedia (PAS-positive and calcium-sensitive in the goldfish) was not clearly affected by the three drugs.

A critical evaluation of the use of filipin-permeabilized rat hepatocytes to study functions of the endoplasmic reticulum in situ.

We have used transmission (TEM) and scanning electron microscopy (SEM) and leakage of lactate dehydrogenase (LDH; EC 1.1.1.27) to evaluate two published procedures which use filipin to render isolated rat hepatocytes permeable to ionic substrates. Cells treated by the procedure of Jorgenson and Nordlie retained less than 10 per cent of their LDH. TEM revealed severe damage to the internal structure of these cells, which included swelling, disintegration and extensive vesicularization of the endoplasmic reticulum (ER). Hepatocytes treated with filipin by the procedure of Gankema et al. retained 65-75 per cent of their LDH and displayed incomplete but highly variable permeability to Trypan blue. SEM revealed the loss of microvilli, other signs of swelling, and the presence of large lesions in the plasma membrane. TEM revealed signs of cell swelling, but the nuclei and the mitochondria were only moderately altered. The rough ER was not swollen, but significant fragmentation was evident and characteristic stacks of lamellar ER were never seen. We conclude that useful information about the functions of the ER in situ cannot be obtained from studies of filipin-treated cells. Our results indicate that retention of LDH is not a sufficient criterion of preservation of cell morphology and that staining with Trypan blue may significantly underestimate the permeability of cells to small ionic metabolites.

Measurement of enzymic lipid peroxidation by intact, lamellar endoplasmic reticulum using electropermeabilized rat hepatocytes

Conference Article| April 01 1987 Measurement of enzymic lipid peroxidation by intact, lamellar endoplasmic reticulum using electropermeabilized rat hepatocytes RICHARD S. G. STANLEY; RICHARD S. G. STANLEY 1Department of Biochemistry, Royal Free Hospital, School of Medicine, Rowland Hill Street, London NW3 2PF, U.K. Search for other works by this author on: This Site PubMed Google Scholar RITA O'REILLY; RITA O'REILLY 1Department of Biochemistry, Royal Free Hospital, School of Medicine, Rowland Hill Street, London NW3 2PF, U.K. Search for other works by this author on: This Site PubMed Google Scholar CATHERINE RICE-EVANS; CATHERINE RICE-EVANS 1Department of Biochemistry, Royal Free Hospital, School of Medicine, Rowland Hill Street, London NW3 2PF, U.K. Search for other works by this author on: This Site PubMed Google Scholar TERENCE HALLINAN TERENCE HALLINAN 1Department of Biochemistry, Royal Free Hospital, School of Medicine, Rowland Hill Street, London NW3 2PF, U.K. Search for other works by this author on: This Site PubMed Google Scholar Biochem Soc Trans (1987) 15 (2): 226–227. https://doi.org/10.1042/bst0150226 Article history Received: September 24 1986 Views Icon Views Article contents Figures & tables Video Audio Supplementary Data Peer Review Share Icon Share Facebook Twitter LinkedIn MailTo Cite Icon Cite Get Permissions Citation RICHARD S. G. STANLEY, RITA O'REILLY, CATHERINE RICE-EVANS, TERENCE HALLINAN; Measurement of enzymic lipid peroxidation by intact, lamellar endoplasmic reticulum using electropermeabilized rat hepatocytes. Biochem Soc Trans 1 April 1987; 15 (2): 226–227. doi: https://doi.org/10.1042/bst0150226 Download citation file: Ris (Zotero) Reference Manager EasyBib Bookends Mendeley Papers EndNote RefWorks BibTex toolbar search Search Dropdown Menu toolbar search search input Search input auto suggest filter your search All ContentAll JournalsBiochemical Society Transactions Search Advanced Search Keywords: ER, endoplasmic reticulum, TBA, thiobarbituric acid, TBARP, TBA-reactive products, GT, UDP-glucuronyl-transferase This content is only available as a PDF. © 1987 Biochemical Society1987 Article PDF first page preview Close Modal You do not currently have access to this content.

Cytological and ultrastructural changes in the pituitary pars distalis of the goldfish kept in calcium-free environments.

The cytology and ultrastructure of the pars distalis, mainly that of prolactin (PRL) cells, were investigated in goldfish adapted to fresh water (FW) or deionized water (DW) for 20 and 40 days, or gradually adapted to 1/3 artificial sea water (ASW) or 1/3 Ca-free sea water. When compared to PRL cells of goldfish kept in FW, those of goldfish adapted to DW did not show signs of increased activity. The lack of exocytotic activity and the low development of various organelles suggested that cell activity was slightly reduced. In 1/3 ASW, PRL cells were smaller and less active. In 1/3 Ca-free ASW, PRL cells appeared slightly stimulated compared with those of fish in 1/3 ASW. The Golgi area was more developed and a few lamellae of endoplasmic reticulum were observed in some cell islets. However, there was no significant difference between PRL cells of goldfish kept in 1/3 Ca-free ASW and in FW. In 1/3 ASW, which is isosmotic to the blood, thyrotrophs (TSH cells) corticotrophs (ACTH cells) and somatotrophs (STH cells) were not clearly affected. In DW, these cells and their nuclei were significantly enlarged. Their stimulation was also evident in 1/3 Ca-free ASW; values for cellular and nuclear areas were maximal in this environment and significantly higher than those of fish in FW and 1/3 ASW. These data suggest that in addition to the PAS-positive cells of the pars intermedia, highly stimulated in Ca-free environments, other cell types of the pars distalis may be involved in osmoregulation, and that the role of PRL cells is not primordial in the goldfish.

Fetal rat somatotropes in vitro: effects of insulin, cortisol, and growth hormone-releasing factor on their differentiation: a light and electron microscopic study.

Somatotropes first appear in the fetal rat pituitary just before term. These cells have never been detected in cultured fetal pituitaries. A modified culture medium has, however, enabled their differentiation in vitro. Hypophysial primordia were explanted on days 13-18 of gestation and cultured in different media until the equivalent of term. Immunoreactive somatotropes could be detected, by light and electron microscopy, in cultured primordia explanted on day 14 of gestation or later. The size and numbers of immunoreactive cells depended on culture medium composition. The control medium, containing insulin, cortisol, T3, and glucagon, proved favorable to somatotrope differentiation and proliferation. Increased insulin concentration reduced somatotrope numbers. In the presence of only insulin and cortisol (or corticosterone) somatotropes were more numerous than in the control. Culture medium enriched with insulin alone, with insulin and T3, or with insulin and glucagon, was not suitable for development of this cell type. Addition of GH-releasing factor ( GHRF ) to the medium during the first culture day did not accelerate the first appearance of the somatotropes but did significantly increase their size. GHRF addition 1/2 h before the end of culture did not modify their morphology. The ultrastructure of somatotropes in vitro is very similar to that observed in vivo on day 21 of gestation. The cells were characterized by their lamellar endoplasmic reticulum and immunoreactive secretory granules (300-400 nm maximal section diameter). Fetal somatotropes can, therefore, be successfully caused to differentiate in vitro. Their appearance depends on insulin and glucocorticoid concentration. T3 and/or glucagon may be inhibitory. GHRF may increase storage in somatotropes. These factors may also regulate the development of somatotropes in vivo.

Stereological study of endoplasmic reticulum, Golgi complex and secretory granules in the B-cells of normal and alloxan-treated mice.

Different B-cell organelles (lamellar and vesicular endoplasmic reticulum, Golgi complex, whole secretory granules and secretory granule cores) were studied stereologically in pancreatic islets from control mice and mice killed 10 or 60 min following alloxan injection. Ten min following alloxan a significant decrease was observed in the volume, surface and numerical densities of whole secretory granules and their cores, and a significant increase was found in the volume and surface densities of vesicular endoplasmic reticulum. At the 60 min observation time, a significant decrease was seen in the volume density of lamellar endoplasmic reticulum and Golgi complex, and in the volume, surface and numerical densities of whole secretory granules and their cores, and a significant increase was observed in the volume and surface densities of vesicular endoplasmic reticulum, and in the mean values for volume and surface of whole secretory granules and their cores. The stereological data indicate swelling of endoplasmic reticulum, decreased Golgi area, and decreased number and total volume and surface of secretory granules during the first hour after alloxan administration to mice. The observations may be consistent with inhibited insulin synthesis.