Cytoplasm Of Perikarya Research Articles

Immunohistochemical localisation of two phosphatidylinositol 4-kinase isoforms, PI4K230 and PI4K92, in the central nervous system of rats.

The distribution and cellular localisation of the phosphatidylinositol 4-kinase isoforms, PI4K230 and PI4K92, that are believed to play important roles in the intracellular signalling mechanisms were studied in the rat brain (cortex, cerebellum, hippocampus and spinal cord) using immunocytochemistry with light and electron microscopy. PI4K230 was detected with a specific antibody purified by affinity chromatography from the egg yolk of chicken immunised with a 33-kDa fragment of bovine PI4K230, comprising amino acids 873-1175 of the native protein. PI4K92 was immunostained with a commercially available antibody raised in rabbit against amino acid residues 410-537 of human PI4K92. At the light microscopic level, the immunostaining of PI4K230 and PI4K92 showed a very similar distribution throughout the neurons and appeared as dense punctate labelling in the cytoplasm of perikarya and stem dendrites of various neurons. In addition to neurons, a strongly stained cell population was observed in the molecular layer of the cerebellar cortex that resembled Bergmann glia cells. Electron microscopy of neurons in the ventral horn of the spinal cord showed dense granular immunoprecipitates for both PI4K230 and PI4K92, mostly associated with the outer membrane of mitochondria and membranes of the rough endoplasmic reticulum. In addition, immunostaining of PI4K92 was also frequently found on the outer surface of cisterns and vesicles of Golgi complexes, whereas PI4K230 immunoreactivity was colocalised with some multivesicular bodies. Neither nuclear localisation nor a regular attachment to the cell membrane of these enzymes were observed. Our findings indicate that PI4K230 and PI4K92 are not involved directly in the ligand-stimulated turnover of phosphoinositides at the plasma membrane of neurons. However, they may provide regulatory phosphoinositides for intracellular vesicular traffic being associated with various organelles.

Immunohistochemical distribution of the prohormone convertase PC5-A in rat brain

Prohormone convertase 5 is an endoprotease of the kexin/subtilisin-like family, which has been postulated to play a role in the proteolytic maturation of a variety of pro-peptides in the mammalian brain. In order to gain insight into the functional role of prohormone convertase 5 in the central nervous system, the regional, cellular and subcellular distributions of the enzyme were investigated by immunohistochemistry in rat brain using an N-terminal-directed specific antibody shown previously to recognize both the mature and unprocessed forms of the enzyme. Throughout the brain, prohormone convertase 5 immunoreactivity was concentrated within nerve cell bodies and proximal dendrites. No prohormone convertase 5 immunoreactivity was associated with astrocytes, as confirmed by the absence of prohormone convertase 5 immunolabeling in cells immunopositive for the glial protein S-100α. Within neurons, prohormone convertase 5 immunoreactivity was concentrated within the Golgi apparatus, as revealed immunohistochemically within the same sections using antibodies against the medial cisternae protein MG-160. It was also present within small vesicular-like elements distributed throughout the cytoplasm of perikarya and dendrites, but not of axons, as confirmed by its lack of co-localization with the synaptic terminal marker Dynamin-1. These results suggest that prohormone convertase 5 is active within early compartments of the neuronal regulated secretory pathway and that it is unlikely to be released with its processed substrates. At the regional level, prohormone convertase 5-immunoreactive perikarya were distributed extensively throughout the forebrain. The most numerous and intensely labeled were detected in the olfactory bulb, cerebral cortex, globus pallidus, endopeduncular and subthalamic nuclei, septum, diagonal band of Broca, magnocellular and medial preoptic areas, supraoptic and arcuate nuclei of the hypothalamus, and anterodorsal, laterodorsal, paraventricular and reticular nuclei of the thalamus. Moderate to dense neuronal labeling was also evident in the olfactory tubercle, caudate–putamen, claustrum, bed nucleus of the stria terminalis, substantia innominata, hippocampus, amygdala, and remaining thalamic and hypothalamic nuclei. This widespread distribution suggests that prohormone convertase 5 is involved in the processing of a variety of neuropeptide and/or neurotrophin precursors in mammalian brain.

Septohippocampal neurons in the rat septal complex have substantial glial coverage and receive direct contacts from noradrenaline terminals

The ultrastructure of septohippocampal neurons in the septal complex and their relations with catecholamine, principally noradrenaline, terminals were examined in single thin sections. Projection neurons were identified by retrograde transport of wheat-germ agglutinated apo-horseradish peroxidase conjugated to colloidal gold particles (WAHG) following an injection into the hippocampal formation of anesthetized adult rats. After a 1 day survival, sections through the septal complex were labeled with antibodies to tyrosine hydroxylase (TH) or dopamine-β-hydroxylase (DBH). By light microscopy, numerous processes with TH- and DBH-immunoreactivity were near neurons containing retrogradely transported WAHG. By electron microscopy, most WAHG was associated with lysosomes, multivesicular and ‘sequestration’ bodies in the cytoplasm of perikarya and large dendrites. WAHG-labeled perikarya ( n = 114) had a large amount of astrocytic coverage (> 60% of surface) and a low amount of terminal coverage (< 25%). WAHG-labeled perikarya and dendrites were either directly contacted by TH- or DBH-labeled terminals or abutted glial processes apposed to TH- or DBH-labeled terminals. Immunoreactivity for TH and DBH was found primarily in axons and axon terminals. The morphology and synaptic associations of TH-labeled terminals was similar to that reported previously. DBH-labeled terminals ( n = 314; 0.5 ± 0.2 μm in diameter) contained numerous small clear vesicles and from 0–4 large, dense-core vesicles. DBH-containing terminals: (1) contacted perikarya and dendrites (58%), 10% of which contained WAHG; (2) were closely apposed to other terminals (7%); or (3) were separated by glial processes (35%). DBH-labeled terminals formed chiefly symmetric synapses on perikarya. However, most DBH-containing terminals formed both asymmetric and symmetric synapses on the shafts of small dendrites, suggesting both excitatory and inhibitory functions for noradrenaline terminals on septal neurons. The results demonstrate that septohippocampal neurons (1) are mostly engulfed by astrocytes and have very little terminal coverage; (2) are both directly contacted (synapses) and indirectly contacted (appositions to opposing astrocytes or axon terminals) by catecholamine, particularly noradrenaline, terminals.

A special type of ganglion cells in the submucous plexus of mouse small intestine.

The perikarya of sensory neurons have been reported frequently from electrophysiological studies of the plexus submucosus of many mammalian digestive tracts, but the fine structural characteristics of those cells or ganglia have rarely been demonstrated. Therefore, we conducted an electron microscopic study in search of possible sensory perikarya in the plexus of the mouse small intestine. Many ganglion cells scattered over the plexus submucosus possessed a few areas of fine granules frequently adjacent to dense bodies in their perikaryonal cytoplasm, as revealed first by Wilson et al. [23], who were unable to explain the significance, of this finding. On the other hand, one type of ganglion with a special three-dimensional structure was observed in the same plexus. This ganglion was composed of one central perikaryon, one innermost cellular semiperfect cover of the Schwann cell, a middle two-layered cellular coat, an outer neuropil and an outermost external lamina. The surface of the central perikaryon, which differed from those of the other ganglion cells, was almost entirely covered with one Schwann cell, most probably for isolation from various kinds of external electrical and/or chemical inputs. Dumbbell-shaped vesicles containing endings that were presumed to be sensory, were frequently localized in the outer neuropil of this ganglion. It is likely that this perikaryon belongs to the sensory ganglion cells and so acts as an important factor in the peristaltic reflex arc.

Light and Electron Microscopic Localization of the Neutral Metalloendopeptidase EC 3.4.24.16 in the Mesencephalon of the Rat.

The topographic and cellular distribution of the neurotensin-hydrolysing neutral metalloendopeptidase 24.16 (EC 3.4.24.16) was examined by light and electron microscopic immunohistochemistry in adult rat mesencephalon. Light microscopic immunoradioautography revealed a ubiquitous distribution of the enzyme throughout the midbrain with a relative enrichment of grey matter areas including the substantia nigra, ventral tegmental area, interfascicular nucleus, interpeduncular nucleus, rostral and caudal linear raphe nuclei, central grey and superficial grey of the superior colliculus. Peroxidase - antiperoxidase immunocytochemistry revealed two distinct cellular patterns of immunostaining: (1) weakly labelled neuronal perikarya more or less uniformly distributed throughout the grey matter, and (2) intensely immunoreactive glial cells heterogeneously distributed across the mesencephalon. Areas exhibiting dense concentrations of endopeptidase 24.16-containing glial cells corresponded to those displaying enhanced immunoreactivity in immunoradioautographs, suggesting that a major proportion of brain endopeptidase 24.16 is associated with glia. Electron microscopic examination of the substantia nigra and ventral tegmental area confirmed the association of the enzyme with a subpopulation of neurons and allowed identification of labelled glial cells as protoplasmic astrocytes. In neurons, endopeptidase 24.16 immunoreactivity was distributed heterogeneously within the cytoplasm of perikarya, dendrites and axons. Reaction product was also characteristically associated with restricted zones of the plasma membrane and underlying neuroplasm. In astrocytes, endopeptidase 24.16 immunostaining was densely and uniformly distributed throughout the cytoplasm of cell bodies and processes. Many of these processes were in direct contact with endopeptidase 24.16-immunopositive neuronal elements. The present results demonstrate that within the midbrain, endopeptidase 24.16 is both intracytoplasmic and membrane-associated in neurons and predominantly intracytoplasmic in glia. The presence of a large number of immunostained elements within areas of the midbrain known to display high levels of neurotensin and/or neurotensin receptors, together with the demonstrated catabolic activity of the enzyme on neurotensin in vitro, is consistent with a role of endopeptidase 24.16 in the functional inactivation of endogenous neurotensin in this region of the brain.

Parvalbumin-immunoreactive material in the earthworm, Lumbricus terrestris

Parvalbumin-like material was localized using an immunoeytochcmieal method, in neurons of the central nervous system and in cells intermingled in the skin of the earthworm. Lumbricus terreslris L. Parvalbumin-immunoreactive material was found in the cytoplasm of perikarya and neutrites, not in the nucleoplasm. In contrast to vertebrates, Lumbricus musculature did not contain parvalbumin-immunoreactive material.

A transmission electron microscopic study of rabbit liver sinusoids with special remarks on an experimentally induced canalicular system and the "pored domes" in the endothelial cells.

Rabbits were starved for a week and then allowed to take the standard diet; the animals were sacrificed after 20 or 24 hr. One group of the animals were treated in winter, and the other, in summer. Control animals were fed solid rabbit food ad libitum. In the control rabbits, short rudimentary canaliculi and occasional small-sized pored domes were found in the thicker portions of the endothelial extension. In the animals experimentally treated in winter, well-developed meandering canaliculi with vacuolar expansions and constrictions appeared in the thicker portions and perikaryonal cytoplasm. In addition prominent pored domes were produced. The canaliculi often formed sponge-like networks with openings both to the sinusoid and the Disse's space. The development of the canalicular system and pored domes correlated to an increased blood flow through the sinusoids; this seemed to be induced upon sudden feedings after lengthy starvation. The rabbits subjected to the experimental treatment in summer exhibited no distinct development of the canaliculi and pored domes. This result was ascribed to the inhibited functions of the rabbit liver during this season.

Immunoelectron microscopical location of the acid cysteine proteinase inhibitor in the lymphatic tissue of the tonsils

Previous studies have shown the acid cysteine proteinase inhibitor (ACPI) to be a common characteristic of human squamous epithelial cells and dendritic reticulum cells (DRC) in lymphoid secondary follicles. In the present study, we used the PAP-preembedding technique for the immunoelectron microscopic identification of ACPI in DRC in palatine tonsillar tissue. ACPI appeared to be located in the cytoplasm of perikaryon and of dendritic processes of DRC on the membranes of the endoplasmic reticulum and the outer membrane of mitochondria. Further more, the tonofilaments and the desmosomal membranes exhibited a strong ACPI reactivity.

A transmission electron microscopic study on sinusoidal cells of guinea pig liver, with special reference to the occurrence of a canalicular system and "pored domes" in the endothelium.

Hepatic sinusoidal cells in the guinea pig were examined by transmission electron microscopy (TEM). A meandering canalicular system was detected in the sinusoidal endothelial cell both in thicker portions of cytoplasmic extensions and in small areas of the perikaryon. It consisted of meandering canaliculi with vacuolar expansions and constrictions, which penetrated the endothelial cytoplasm, forming as a whole a network. The canaliculi possessed more than two openings which usually communicated with the sinusoid, but occasionally poured themselves into the Disse's space. This network of canaliculi seems to permit infiltration of blood plasma. The "pored domes" recorded by Fujita and his collaborators on the glomerular endothelium of the rat and rabbit kidney were also revealed on the perikaryonal cytoplasm of the sinusoidal endothelium of guinea pig liver. Osmium-blackened lipid droplets were found in the sinusoidal endothelium, which suggested the release of lipid into the sinusoid. Short-term administrations of excessive vitamin A exerted no influence on the endothelial lipid droplets. The guinea pig is a rodent species which stores a very small amount of lipid droplets in its fat-storing cells and the so-called empty fat-storing cells were frequently detected. A single cilium was often found in the fat-storing cells in the guinea pig as in other species.

Induction of differentiation increases Met5-enkephalin and Leu5- enkephalin content in NG108-15 hybrid cells: an immunocytochemical and biochemical analysis

Regulation of cellular content of the endogenous opioid peptides Met5-enkephalin and Leu5-enkephalin was investigated in neuroblastoma X glioma hybrid cells NG108-15 grown in both serum-supplemented and serum-free defined media. Untreated cells and cells induced to differentiate were stained using anti-Met5-enkephalin and anti-Leu5-enkephalin with the peroxidase-antiperoxidase immunocytochemical technique at the light microscopic level. In untreated NG108-15 cells grown in serum-supplemented medium, intense enkephalin-like immunoreactivity was localized in cell bodies and short processes of a select population of cells. The volume fraction of stained untreated cells remained constant throughout the time period investigated. When cells were induced to differentiate with N6,O2'-dibutyryl adenosine 3':5'-cyclic monophosphate (dBcAMP) or 8-bromo cyclic adenosine monophosphate (1.0 mM) treatment for 5 days, staining was found throughout the cytoplasm of perikarya and the extensive processes which were expressed, and the volume fraction of stained cells increased over 2-fold. Receptor-mediated stimulation of adenylate cyclase by prostaglandin E1 (10 microM) for 5 days produced results similar to those with dBcAMP. Pure cultures of differentiated cells with intense staining were obtained by further treatment of cultures, grown in the presence or absence of dBcAMP, with arabinosylcytosine (araC). Untreated, dBcAMP-treated and araC-treated NG108-15 cells grown in defined medium expressed staining patterns and volume fractions of stained cells similar to those grown in serum-supplemented medium; sodium butyrate (1.0 mM), however, increased the volume fraction of stained cells grown in defined medium over 3-fold, whereas it had little effect on staining of cells grown with serum. The presence of both Met5- and Leu5-enkephalin-like activities in NG108-15 cells was confirmed in acid extracts of cells by radioreceptor assay after separation by reverse phase high pressure liquid chromatography. Induction of differentiation in NG108-15 cells by dBcAMP treatment increased the cellular concentration of both enkephalins to over 2 times the levels found in untreated cells. The biochemical analysis for Met5-enkephalin- and Leu5-enkephalin-like activity compared well with the immunocytochemical data indicating that the enkephalin content is correlated with the state of differentiation of NG108-15 cells.

On the occurrence of a myeloid body in pinealocytes of the white-footed mouse, Peromyscus leucopus. An electron-microscopic study.

The fine-structural features of pinealocytes of the white-footed mouse, Peromyscus leucopus, were examined. A single population of pinealocytes was observed in both superficial and deep components of the gland. Cells in both locations are characterized by the presence of an indented nucleus exhibiting a prominent nucleolus. The usual organelles in the perikaryon are the Golgi complexes, mitochondria, endoplasmic reticulum, polysomes, microtubules and dense-core vesicles. In addition the perikaryonal cytoplasm often contains a single myeloid body. These bodies are usually lenticular in shape, are formed of an array of flattened membranous cisternae, and are not bounded by a limiting membrane. This organelle usually lies in the vicinity of the nucleus and is infrequently associated with lipid bodies. Complex forms were also observed. The cisternae are continuous with elements of the endoplasmic reticulum at points along their expanded rims. The outer surface of the cisternal membrane exhibits a granularity or fuzziness. The cisternae may be fenestrated. Pinealocyte processes with an abundance of clear vesicles, and some dense-core vesicles and vesicle-crowned rodlets are present in the parenchyma.

An electron microscopic study of autonomic nerve cells in the cloacal region of the lamprey, Lampetra japonica.

Two types of autonomic nerve cell in the cloacal region of lamprey, Lampetra japonica have been studied by electron microscopy. Large ganglion cells (LGC) were unipolar and individually invested with a satellite cell sheath. The LGC-satellite cell complex measured 24 microns X 38 microns on average. Granular endoplasmic reticulum and cored vesicles (80-140 nm in diameter) were scattered in the perikaryon. Two kinds of peculiar cytoplasmic filament were seen in LGC: one type was about 20 nm in diameter with periodic dense material on the surface and the other had a diameter of about 8 nm and showed an undulating profile. Nerve endings containing abundant small clear vesicles (30-50 nm in diameter) and a few larger cored vesicles (50-100 nm in diameter) were found in synaptic contact with LGC. Small ganglion cells (SGC) were also unipolar and covered incompletely by a satellite cell sheath. The SGC-satellite cell complex measured 6 and 12 microns on average. The SGC was packed with organelles and the perikaryon appeared more electron dense than that of LGC. SGC perikaryonal cytoplasm contained dispersed granular endoplasmic reticulum and numerous large cored vesicles (55-220 nm in diameter). Nerve endings containing numerous large cored vesicles (70-170 nm in diameter) and variable numbers of small clear vesicles (30-50 nm in diameter) were seen on the surface of SGC.

Ultrastructure of pinealocytes of the cotton rat, Sigmodon hispidus

Fine structural features of pinealocytes of cotton rats (Sigmodon hispidus) were examined. Golgi complexes, mitochondria, endoplasmic reticulum and polysomes are usual organelles seen in the perikaryonal cytoplasm of pinealocytes. Many non-granulated vesicles (40 to 80 nm in diameter) and a few granulated vesicles (about 100 nm in diameter) are associated with the Golgi cisternae. Occasionally, the cisternae contain granular materials. The perikaryonal cytoplasm of pinealocytes is characterized by the presence of inclusion bodies. These bodies are usually round in shape, not bounded by a limiting membrane and composed of fine granular or filamentous materials of high electron-opacity, which are similar in appearance to the substance seen in the nucleolonema. Pinealocyte processes, filled with abundant non-granulated vesicles and some granulated vesicles, are mainly found within the parenchyma and occasionally in perivascular spaces.

Electron microscope study on the hepatic sinusoidal wall and fat-storing cells in the bat.

The three cell types known to form the hepatic sinusoidal wall were electron microscopically observed in three kinds of bats captured in winter and summer. 1. The cytoplasmic extensions of sinusoidal endothelial cells consisted of continuous thicker parts ("cytoplasmic processes") and discontinuous thinner parts ("sieve plates"). The alternate disposition of the two parts was rather irregular, and the sizes, shapes and spacings of the fenestrae were variable. In the thinner parts with numerous small fenestrae, larger gaps were also mingled. The endothelium was simple-layered and devoid of basal lamina. Interendothelial junctions were found mainly between closely apposed margins of the "cytoplasmic processes" and agreed in structure with the "junctional complex" of WISSE (1970). 2. Kupffer cells, morphologically distinct from the endothelial cells, bulged strongly into the sinusoidal lumen. Provided with many microvillous pseudopods, they were stellate in appearance. They were fixed to the endothelial lining by small junctional areas which occurred between the Kupffer cell body and the "cytoplasmic processes" of the endothelium. 3. Fat-storing cells were located in the Disse's space. They generally contained only smaller amounts of lipid in a few droplets. So-called empty fat-storing cells were numerous, especially in winter bats. The perikaryonal cytoplasm revealed a large Golgi complex and well-developed granular endoplasmic reticulum. The three mesenchymal cell types of the sinusoidal wall possessed the centriole in common within the Golgi complex, but only the fat-storing cell was provided with the single cilium. Fat-storing cells extended cytoplasmic processes ramifying beneath the endothelial lining occasionally surrounding the sinusoids almost completely, and which seemed to reinforce the endothelial lining and to bring about the constriction of the sinusoid. In hypervitaminotic bats that daily received 6,000 I.U. vitamin A for three days, remarkable increase in size and number of lipid droplets was observed in slightly hypertrophic fat-storing cells, and the empty cells disappeared simulating an increased number of fat-storing cells. Suggestion was made of an antidotal function of fat-storing cells against excess vitamin A which might be considered a toxic agent in the broad sense. 4. The Disse's space of bat liver contained plasma cells, lymphocytes and macrophages, the latter too often being under migration through the endothelial fenestrae.

Quaking mouse: ultrastructural evidence for arrest of myelinogenesis

The central nervous systems of 20 Quaking and littermate control mice from 15 to 180 days of age were studied. Some younger control animals, not carrying the Quaking gene, were also examined. The most striking feature of the myelin sheath in Quaking mice was its apparent arrest in growth and maturation, e.g., thin myelin sheaths, lack of compaction with pockets of oligodendroglia cytoplasm and uneven growth of lateral loops with poorly developed nodes of Ranvier. In essence, this picture was observed in Quaking mice irrespective of age, although in mature animals (4 and 6 months of age) a few fibers acquired a myelin sheath of almost normal thickness. Demyelination, in the strict sense of complete destruction of ultrastructurally normal myelin, was not observed in the mutant mice. However, the perikaryonal cytoplasm of the oligodendroglial cells, as well as the oligodendroglial processes forming the myelin sheath, often contained whorls of myelin-like material. Similar accumulations of myelin ‘debris’ were occasionally observed in young normal mice, presumably as the result of the normal remodeling process. Some of the myelin figure accumulation in mutant mice may be the result of this phenomenon. It is also possible that the genetic lesion in Quaking mice results in a defect in assembly of normal mature myelin, because there is a deficit in some essential component. Therefore biosynthesized lipids cannot be used in myelin formation and accumulate in the oligodendroglial cells as myelin ‘debris’.