Areas Of Rough Endoplasmic Reticulum Research Articles

Ryanodine receptor localisation in the mammalian cochlea: An ultrastructural study

Calcium-induced calcium release (CICR) in the mammalian cochlea has been suggested to enhance neurotransmitter release from inner hair cells and facilitate the efferent response in outer hair cells. Light microscopic evidence exists for the presence of ryanodine receptors in the organ of Corti but there is so far no information about their ultrastructural localisation. We have therefore used post-embedding immunogold labeling with antibodies that predominantly recognise ryanodine receptor isoforms 1 (RyR1) and 2 (RyR2) to investigate their distribution in rat cochleae. In inner hair cells, the highest levels of labeling were observed over an area of rough endoplasmic reticulum that lies in the cytoplasmic region beneath the nucleus; in outer hair cells, the cytoplasmic region above the nucleus displayed most labeling. Labeling was also associated with the subsurface cisternae adjacent to the lateral membranes of both types of hair cell, with the efferent terminals on the outer hair cells and was observed in adjacent supporting cells. Labeling in outer hair cells was significantly higher than that in inner hair cells or in the supporting cells. Our results support the presence of RyR1 in the cochlea but do not rule out the presence of other isoforms. CICR may be involved in the control of calcium levels in the base of the inner hair cells and supporting cells, and in the cholinergic efferent response and motile behaviour of the outer hair cells.

Ultrastructural evidence of sexual dimorphism in supraoptic neurons: a morphometric study.

We have recently shown that in spite of the absence of receptors for gonadal steroids in the supraoptic nucleus (SON) of the rat hypothalamus, the volume of the nucleus and the size of its neurons are larger in males than in females, and that these differences between male and female rats are correlated with body weight and dependent on the vasopressinergic neurons. As supraoptic neurons and their organelles enlarge when they are engaged in active peptidergic secretion we have carried out a morphometric ultrastructural analysis to determine if cell structures involved in the synthesis and storage of neurosecretory material also display weight-dependent sex dimorphism. Groups of six male and six female rats aged 30, 60 and 180 days were used. Nucleoli, rough endoplasmic reticulum and neurosecretory granules were analysed and we estimated their volume or surface densities and the total volume of nucleoli and rough endoplasmic reticulum, and total surface area of rough endoplasmic reticulum. We found that, with the exception of neurosecretory granules, the densities of the organelles did not differ among the groups studied, but total values were higher in males. These differences were found to be weight-dependent. Since the organelles studied are regarded as reliable indicators of the neurosecretory activity of supraoptic neurons, our data fully support the view that the weight-dependent sexual dimorphism observed in this nucleus reflects greater synthetic activity of its vasopressinergic neurons associated with the need to maintain water balance in larger bodies.

Cellular changes in rat parathyroids provoked by progesterone and testosterone.

Male rats kept on a standard diet were treated either with progesterone or testosterone by a single intramuscular injection of preparations which are slowly absorbed and metabolized. The rats were anaesthetized 24 h after application of the hormones, perfused with glutaraldehyde, and the parathyroid glands prepared for electron microscopy. Morphometric analysis revealed that both progesterone and testosterone provoked (1) an increment in nuclear and cell volume and a concomitant increment in cell surface area, and (2) an increment in surface area of rough endoplasmic reticulum by 42% and 49%, and of the Golgi complex by 85% and 63%, respectively. Previously, we had found that oestradiol treatment led to a similar response in parathyroid cells. The conclusion is thus drawn that male and female sex hormones induce membrane synthesis resulting in an enhanced capacity for parathyroid hormone secretion since RER and Golgi complex are concerned with this secretion. It is considered probable that sex hormones have the ability fundamentally to modulate secretory activity in parathyroid cells.

Fine structure of hepatocytes in juvenile grey mullets: Liza saliens Risso, L. ramada Risso and L. aurata Risso (Teleostei, Mugilidae)

Livers of 45 fingerling grey mullets (Liza sp. 5-15 cm long) were subjected to light microscopy, transmission electron microscopy and enzyme cytochemical examination. The hepatic structure was found to be comparable in all three Liza species. As in other fish species, Liza liver parenchyma is arranged in cords (or dual-plated laminae, in sections parallel to the sinusoidal axis). Hepatocytes showed numerous microvilli in their perisinusoidal and canalicular areas. They contained a centrally located nucleus, fairly large areas of rough endoplasmic reticulum, consisting mainly of parallel cisternae, and pools of glycogen. A few small lipid droplets were evenly distributed in the cytoplasm. The Golgi fields, between the canaliculus and the nucleus, consist mainly of dictyosomes, lysosomes and autophagic vacuoles. Acid phosphatase activity was cytochemically localized to reaction products in secondary lysosomes and lipofuscin granules. A comparison between grey mullet liver and that of other teleosts was made.

Short term effects of glucocorticoids upon hippocampal ultrastructure.

This study evaluated hippocampal cytoarchitecture in male Sprague-Dawley (225-250 g) rats treated with corticosterone (CORT) and in unhandled, unstressed rats. CORT-treated animals received 20 mg/kg CORT in sesame oil/day x 3 days; each dose produces levels in the upper physiologic range, similar to those produced by major stressors. Using quantitative electron microscopy, sections from regions Ca3b, Ca1, and dentate gyrus were examined. Surface density and surface area of Golgi and the number of Golgi-stacks increased following CORT-treatment (p less than 0.05, ANOVA). Paralleling these changes surface area of rough endoplasmic reticulum and number of cisternae also increased in neuronal perikarya (p less than 0.001, p less than 0.003 respectively). In contrast, surface densities of mitochondria, multivesicular bodies, and lysosomes were unaffected by CORT treatment. These data suggest that both pyramidal and granule cells of hippocampus are subject to short term effects of high physiological levels of CORT, and that these effects may involve stimulation of protein biosynthesis.

Effects of transplantation on mouse adrenal chromaffin cells.

The effects of long-term transplantation on the ultrastructure of adrenaline- and noradrenaline-storing cells from the adrenal medulla were determined using morphometric methods. Mouse adrenal medulla were freed from the adrenal cortex and grafted into the occipital cortex of the brain. Two types of chromaffin cells were identified by electron microscopy in grafts fixed with glutaraldehyde and osmium tetroxide. Noradrenaline-type cells were predominant and formed 70-80% of the surviving population of grafted chromaffin cells. A minority of the chromaffin cells contained medium-sized granules (140-210 nm in diameter) (medium granule cell; MGC) with finely granular moderately electron dense cores. Morphometric analysis of noradrenaline phenotype cells and MGC cells in transplants showed no significant differences compared with the noradrenaline-storing cells of normal adrenal glands. In contrast, noradrenaline-type cells and MGC cells in the grafts had areas of secretory vesicles which were significantly (P less than 0.01) larger and areas of rough endoplasmic reticulum which were significantly (P less than 0.01) smaller than those of the adrenaline-storing cells of normal adrenal glands. It was concluded that long-term transplantation caused no degenerative changes in the ultrastructure of mouse adrenal chromaffin cells.

Rat intermediate lobe in culture: a histological and biochemical characterization.

The histology, immunohistochemistry, peptide synthesis and secretion as well as proliferation rate of rat intermediate lobe (IL) were studied in primary cultures. The cultures contained two populations of cells: melanotrophs either organized in free floating lobules or in lobules which attached to the dishes and formed a monolayer. Both populations retained their in vivo morphology: polyhedral cells with smooth, ovoid nuclei and a large number of cytoplasmic secretory coated vesicles, a well developed Golgi apparatus, abundant mitochondria and extensive areas of rough endoplasmic reticulum. The melanotrophs stained with varying intensity for alpha-MSH and in situ hybridization showed the presence of pro-opiomelanocortin (POMC) mRNA. 35S-methionine incorporation combined with 2-D gel electrophoresis demonstrated POMC peptide synthesis and radioimmunoassay confirmed its secretion into the medium. 3H-thymidine uptake in the attached melanotrophs was considerably higher than that in the free-floating melanotrophs, demonstrating the dependency of proliferation rate on the cytoarchitecture of the explant. The retention of melanotroph morphology, biosynthetic and proliferative capacity in vitro affords a valid model system for studying POMC gene expression.

QUANTITATION OF CHONDROCYTE PERFORMANCE IN GROWTH PLATE CARTILAGE DURING LONGITUDINAL BONE GROWTH

The longitudinal growth of bone depends on the activities of individual chondrocytes of the growth plate. Each chondrocyte remains in a fixed location throughout its life, and there accomplishes all of its functions. Although a cell may perform several or all of its activities simultaneously, one of these will usually predominate during a particular phase of its life. The two most prominent stages are those of cellular proliferation and hypertrophy (including the mineralization of matrix) before the resorption of tissue during vascular invasion. By applying recently developed stereological procedures and improved methods for the fixation of cartilage, we compared cellular shape modulation, various ultrastructural parameters (surface areas or volumes of endoplasmic reticulum, Golgi membranes, and mitochondria), the production of matrix, and cellular turnover for proliferating and hypertrophic chondrocytes within the proximal tibial growth plate of the rat. By the late hypertrophic stage, fourfold and tenfold increases in the mean cellular height and volume, respectively, and a threefold increase in the mean volume of the matrix per cell were achieved. The high metabolic activity of hypertrophic cells was reflected by a twofold to fivefold increase in the mean cellular surface area of rough endoplasmic reticulum, the Golgi membranes, and the mean cellular mitochondrial volume. Rates of longitudinal growth were determined by fluorochrome labeling and incident-light fluorescence microscopy. Using these values and the stereological estimators describing cellular height, the rates of cellular turnover were calculated. The rapid progression of the vascular invasion front was found to eliminate, for each column of cells, one chondrocyte every three hours; that is, eight cells a day. The maintenance of a steady-state structure for growth-plate cartilage in rats in a steady state of growth thus necessitates efficient compensation for these losses, which is achieved by a high rate of cellular proliferation and rapid hypertrophy.

Quantitation of chondrocyte performance in growth-plate cartilage during longitudinal bone growth.

The longitudinal growth of bone depends on the activities of individual chondrocytes of the growth plate. Each chondrocyte remains in a fixed location throughout its life, and there accomplishes all of its functions. Although a cell may perform several or all of its activities simultaneously, one of these will usually predominate during a particular phase of its life. The two most prominent stages are those of cellular proliferation and hypertrophy (including the mineralization of matrix) before the resorption of tissue during vascular invasion. By applying recently developed stereological procedures and improved methods for the fixation of cartilage, we compared cellular shape modulation, various ultrastructural parameters (surface areas or volumes of endoplasmic reticulum, Golgi membranes, and mitochondria), the production of matrix, and cellular turnover for proliferating and hypertrophic chondrocytes within the proximal tibial growth plate of the rat. By the late hypertrophic stage, fourfold and tenfold increases in the mean cellular height and volume, respectively, and a threefold increase in the mean volume of the matrix per cell were achieved. The high metabolic activity of hypertrophic cells was reflected by a twofold to fivefold increase in the mean cellular surface area of rough endoplasmic reticulum, the Golgi membranes, and the mean cellular mitochondrial volume. Rates of longitudinal growth were determined by fluorochrome labeling and incident-light fluorescence microscopy. Using these values and the stereological estimators describing cellular height, the rates of cellular turnover were calculated. The rapid progression of the vascular invasion front was found to eliminate, for each column of cells, one chondrocyte every three hours; that is, eight cells a day. The maintenance of a steady-state structure for growth-plate cartilage in rats in a steady state of growth thus necessitates efficient compensation for these losses, which is achieved by a high rate of cellular proliferation and rapid hypertrophy.

Ultrastructure of the effects of calcitonin on the development of mouse tooth germs in vitro

Mandibular first molars, from 17-day-old embryos, were cultivated in control medium or medium containing 0.1, 0.01 or 0.001 unit/ml of calcitonin (CT) for periods up to 10 days. In untreated tooth germs, cells of the dental papilla differentiated into pre-odontoblasts up to 4 days and predentine was seen on day 6. Cells treated with 0.1 unit/ml of CT differentiated into pre-odontoblasts up to 4 days, but no predentine was formed even after 10 days in culture. With 0.01 unit/ml, cells differentiated into odontoblasts, and had already secreted predentine a few days earlier than the untreated group. With 0.001 unit/ml, the developing germs were similar to the control expiants during the entire 10-day cultivation period. The proportional area of rough endoplasmic reticulum to cytoplasm of the odontoblasts was low at 0.1 unit/ml of CT and high at 0.01 unit/ml compared to the untreated expiants.

The effect of lipofuscin on cellular function

The neurons of the supraoptic nucleus of male C57BL/Icrfa 1 mice at 6 or 28 months of age were examined from normally hydrated, osmotically loaded and osmotically loaded-rehydrated animals. Using quantitative morphological techniques, a reduction in the concentration of lipofuscin in the neurons was observed in osmotically loaded mice at both ages, and these levels were restored to control values during rehydration. In addition, there was a significant difference in the pattern of response of lipofuscin levels between the two age groups during the experiment. The concentration of hormone containing neurosecretory granules in the neurons of the supraoptic nucleus did not differ significantly between the two age groups during the experiment. However, the surface area of rough endoplasmic reticulum per unit volume of the supraoptic nucleus cell did differ significantly between the two age groups over the course of the experiment. It is concluded that increasing concentrations of lipofuscin do not affect the ability of the cell to control the concentration of neurosecretory granules or rough endoplasmic reticulum. The simplistic view that lipofuscin accumulates with age to the detriment of cell function must be revised.

The effects of constant light and constant darkness on the pineal organ of the goldfish, Carassius auratus.

Goldfish (Carassius auratus) were subjected to both long- and short-term (1, 3, 6, and 170 days) constant light or constant darkness to determine the effects of environmental lighting on the morphology of photoreceptor cells in the pineal organ. Morphometric analysis of electron micrographs from each experimental group suggested that the effects of constant darkness are morphologically expressed only after long-term (6 months) exposure. These included an increase in the volume of photoreceptor and phagocytic cells, area of rough endoplasmic reticulum, and volume of mitochondria per photoreceptor cell, and a decrease in the size of individual Golgi bodies. On the other hand, the response to constant light occurred generally within the first 6 days. Under these conditions, there was a reduction in outer-segment volume, but an increase in both the volume of Golgi per photoreceptor cell and size of individual Golgi bodies. Synaptic ribbons in these specimens were longer at every time sampled. The results of this study provide evidence that photoreceptor cell organelles involved in synthesis and secretion, as well as photosensory functions, are influenced by environmental lighting.

The influence of age on the response of the supraoptic nucleus of the hypothalamo-neurohypophyseal system to physiological stress II. Quantitative morphology

A quantitative study of various morphological parameters of neuroendocrine cells from the hypothalamo-neurohypophyseal system has been carried out. The cells of the supraoptic nucleus (SON) of male C57BL/ICrfa t mice at 6 and 28 months of age were examined from both normally hydrated and osmotically loaded animals. The only significant age-related difference between the principal components of the SON ( i.e. SON cells, glia, capillary and neuropil) from normally hydrated mice was an increase in the relative volume fraction of the capillary lumen in the SON of old animals. No significant differences could be detected in the above-mentioned components of the SON from young and old salt-stressed mice. There were differences relating to age and to salt stress in the volume fractions of several sub-cellular components of SON cells. Under the influence of osmotic stress there was an increase in the relative volume of the SON cell occupied by cytoplasm when compared to the controls; the response was similar at both ages tested. Similarly for the nucleus, the relative volume of the SON cell occupied at both ages in control and salt-stressed animals was of the same order. The relative volume of SON cells occupied by mitochondria did not alter at either age during salt stress; however, the number of mitochondria per unit area of SON cell cytoplasm fell significantly in the old salt-stressed mice. The volumes of the Golgi and the hormone-containing granules (neurosecretory granules) of these cells remained the same with both age and salt stress. The surface area of the rough endoplasmic reticulum, however, was similar in SON cells from old and young controls, but there was significantly less rough endoplasmic reticulum in cells from the old when compared with the young salt-stressed animals. There was a significant increase in the volume of the soncell occupied by lipofuscinnin old animals, and the relative volume of age pigment was significantly reduced in the cells of salt-stressed animals at both ages. The morphological correlates of functional activity in the SON cells from young, osmotically loaded mice are similar to those seen in other laboratory rodents under the same conditions. In the old animal, the overall response to osmotic stress by the SON as an organ, and also by the individual cells ( i.e. cytoplasm and nucleus), are similar to those in young animals. However, there are significant differences in detail, particularly with respect to mitochondrial numbers, surface area of rough endoplasmic reticulum and the relative volumes of lipofuscin and “lipid bodies” in these cells. The morphological correlates of hormone production (neurosecretory granules) do not show age-related changes. The findings are discussed with respect to the efficiency of function of the aged neuroendocrine cell under osmotic stress conditions.

Membrane dynamics in the parotid acinar cell during regranulation: A stereological study following isoprenaline‐induced secretion

Recently weaned male rabbits were injected either with 150 micrograms/kg isoprenaline in saline containing 0.01 M ascorbic acid or simply with the drug vehicle. Groups of drug-injected animals were killed at various time after injection. Parotid gland tissue samples from all animals were fixed, embedded and thin sectioned, and micrographs were prepared at standard magnification. Estimations of membrane areas of each membrane type in parotid acinar cells were made. It was found that in animals killed 2 hours after induced secretion apical area was larger than in controls. In animals killed at successively later times the apical area was progressively less. No elevation of any internal smooth membrane areas was ascertained at any sampling time, though the areas of rough endoplasmic reticulum in 2-12 hour samples were larger. It is suggested that excess apical membrane, though probably removed by interiorization, is afterwards disassembled in side the cell to create fresh macromolecular building units (protein molecules), perhaps after passing through the Golgi apparatus. This cryptic pool of building units can provide about 900 micrometers2 of secretion granule membrane per cell, the supply apparently being exhausted in the first eight hours after degranulation, whilst granule numbers are being increased. Thereafter, apparently, limited granule fusion occurs, so that ultimately the cellular complement of secretion granule membrane comes to enclose a greater volume of secretory product, though the average granule number per cell is small.

An ultrastructural study of the red nucleus in the rat.

The red nuclei of 14 adult male rats of the Wistar strain were prepared for electron microscopic study following perfusion with a mixture of aldehydes, Neurons of four size categories were identified in 1 mu Epon sections and their ultrastructural characteristics were studied in adjacent thin sections. Giant (greater than 40 mu) and large (26-40 mu) neurons are distinguished primarily by size and possess similar ultrastructural features: extensive areas of rough endoplasmic reticulum (RER), a prominent perinuclear Golgi complex, numerous mitochondria and pigment granules and a large, ovoid nucleus which occasionally contains intranuclear rodlets. Medium size neurons (20-25 mu) have less extensive, poorly organized RER and randomly distributed Golgi complexes. The nuclear envelopes of these cells frequently show multiple invaginations and continuity with the RER cisternae. In small neurons (less than 20 mu) the RER occurs as single or anastomosing strands whi le golgi complexes and pigment granules are few. In both medium size and small neurons, aggregates of condensed chromatin are adherent to the inner nuclear membrane. Three main types of synaptic terminals may be distinguished in the red nucleus: (1) small terminals with flattened vesicles and symmetrical densities (F terminals), (2) small terminals with rounded vesicles and asymmetrical densities (RS terminals), and (3) large (10-15 mu) asymmetrical, rounded vesicle terminals which form multiple contacts along their length (RL terminals). The small neurons receive both F and RS terminals on their dendrites and infrequently on their cell somas. The large and giant neurons receive F, RS and RL terminals on their somas and proximal dendrites and F and RS terminals on their distal dendrites. The somas and dendrites of medium size neurons receive both F and RS terminals but RL terminals do not lie in relation to them. Spine contacts are common throughout the nucleus and occur on both somas and dendrites.

Subcellular morphometric and biochemical analysis of developing rat hepatocytes.

Livers of rats between the 16th gestational and 100th postnatal day of age were subjected to quantitative biochemical and electron microscope, morphometric analyses. The amount of total mitochondrial protein per gram of liver remained at 34% of the adult level throughout the last 4 days of gestation but this was the period of rapid rise in the levels of cytochrome c oxidase, aspartate aminotransferase, and glutamate dehydrogenase in mitochondria; the nuclear fraction also acquired some glutamate dehydrogenase but lost most of it during postnatal development. During early postnatal life the amount of mitochondrial protein rose in parallel with the levels of cytochrome c oxidase and glutamate dehydrogenase but the upsurges of glutaminase and, later, of ornithine aminotransferase were accompanied by relatively little change in total mitochondrial protein. The surface area of rough endoplasmic reticulum per unit volume of hepatocyte cytoplasm (S(v) (RER)) did not change significantly throughout the period of development studied. From the 16th day of gestation to term the surface area of smooth ER (S(v) (SER)), the volume occupied by mitochondria (V(v) (MT)) and their number (N(v) (MT)) remained at 30, 66, and 45% of their adult values, respectively. V(v) (MT) and N(v) (MT) attained their maximal levels by the 2nd postnatal day and S(v) (SER) between days 2 and 12. Mitochondria of adult liver are thus smaller and contain more protein per unit volume than do those of fetal liver. After the 12th postnatal day, hepatocytes treble their size; they acquire more cytoplasm with additional enzymes but without further change in organelle concentration. The data reveal several distinct phases in the differentiation of hepatocytes. Each phase can be characterized by the extent to which the quantity and composition of various subcellular compartments evolve.