Presence Of Dense-core Vesicles Research Articles

Ultrastructural Heterogeneity of Gangliogliomas

Gangliogliomas are rare brain tumors, composed of neuronal and glial cells mixed in a different proportion. The basic histopathological pattern of gangliogliomas is well recognized but the variable microscopic appearance still can pose a challenge to the neuropathologist. The authors reanalyzed their series of gangliogliomas in the files of two departments of neuropathology. All analyzed tumors fulfilled the WHO histological criteria of ganglioglioma. Seven tumors were examined by electron microscopy. The following ultrastructural features were graded: presence of dense-cored vesicles, synaptic vesicles, synapses and intermediate filaments, abundant basal membranes, dystrophic neurites, autophagic vacuoles, and multivesicular bodies. Most of the neoplastic neurons were large, polyglonal or oval with well-developed subcellular organelles, round nuclei, and prominent nucleoli. In most cases there were abundant dense core vesicles, observed in both the tumor cell bodies as well as in their processes. Synapses were typically observed. Intermediate filaments were abundant in all tumors. The most intriguing ultrastructural finding was abundant presence of autophagic vacuoles. In 4 cases, multivesicular bodies were observed. All of the tumors with multivesicular bodies also contained abundant autophagic vacuoles.

Caudal neurosecretory system of the zebrafish: ultrastructural organization and immunocytochemical detection of urotensins

The caudal neurosecretory system is described here for the first time in the zebrafish, one of the most important models used to study biological processes. Light- and electron-microscopical approaches have been employed to describe the structural organization of Dahlgren cells and the urophysis, together with the immunohistochemical localization of urotensin I and II (UI and UII) peptides. Two latero-ventral bands of neuronal perikarya in the caudal spinal cord project axons to the urophysis. The largest secretory neurons (approximately 20 microm) are located rostrally. UII-immunoreactive perikarya are much more numerous than those immunoreactive for UI. A few neurons are immunopositive for both peptides. Axons contain 75-nm to 180-nm dense-core vesicles comprising two populations distributed in two axonal types (A and B). Large dense vesicles predominate in type A axons and smaller ones in type B. Immunogold double-labelling has revealed that some fibres contain both UI and UII, sometimes even within the same neurosecretory granule. UII is apparently the major peptide present and predominates in type A axons, with UI predominating in type B. A surprising finding, not previously reported in other fish, is the presence of dense-core vesicles, similar to those in neurons, in astrocytes including their end-feet around capillaries. Secretory type vesicles are also evident in ependymocytes and cerebrospinal-fluid-contacting neurons in the terminal spinal cord. Thus, in addition to the urophysis, this region may possess further secretory systems whose products and associated targets remain to be established. These results provide the basis for further experimental, genetic and developmental studies of the urophysial system in the zebrafish.

Presynaptic localization of the carboxy-terminus epitopes of the μ opioid receptor splice variants MOR-1C and MOR-1D in the superficial laminae of the rat spinal cord

Opioids inhibit nociceptive transmission at the level of the spinal cord, possibly through inhibition of neurotransmitter release by presynaptic μ opioid receptors (MORs) thus preventing the activation of ascending pathways and the perception of pain. Most nociceptive primary afferents are unmyelinated fibers containing peptides such as substance P and/or calcitonin gene-related peptide. However, few terminals contain both substance P and MOR. Recently, we identified new carboxy-terminal MOR splice variants that are localized in the superficial laminae of the dorsal horn. We now report the precise cellular distribution of two of these MOR-1 variants, MOR-1C (exon 7/8/9 epitope) and MOR-1D (exon 8/9 epitope), at the ultrastructural level. In the superficial laminae of the dorsal horn, the majority of the labeling of MOR-1C and MOR-1D was found in unmyelinated axons. This distribution contrasts with that of MOR-1 (exon 4 epitope), in which labeling is equally found in dendrites and soma, as well as in axons. The presence of dense core vesicles in many of the MOR-1C-like immunoreactive terminals implies that this splice variant might be involved in presynaptic inhibition of transmitter release from peptide-containing afferents to the dorsal horn. Consistent with this finding, confocal microscopy analyses showed that many MOR-1C profiles in laminae I–II also contained calcitonin gene-related peptide, whereas fewer MOR-1 profiles contained either substance P or calcitonin gene-related peptide in this same region. From these findings we suggest that there are differential distributions of MOR-1 splice variants as well as distinct peptide colocalizations in the dorsal horn.

A Regulated Secretory Pathway in Cultured Hippocampal Astrocytes

Glial cells have been reported to express molecules originally discovered in neuronal and neuroendocrine cells, such as neuropeptides, neuropeptide processing enzymes, and ionic channels. To verify whether astrocytes may have regulated secretory vesicles, the primary cultures prepared from hippocampi of embryonic and neonatal rats were used to investigate the subcellular localization and secretory pathway followed by secretogranin II, a well known marker for dense-core granules. By indirect immunofluorescence, SgII was detected in a large number of cultured hippocampal astrocytes. Immunoreactivity for the granin was detected in the Golgi complex and in a population of dense-core vesicles stored in the cells. Subcellular fractionation experiments revealed that SgII was stored in a vesicle population with a density identical to that of the dense-core secretory granules present in rat pheochromocytoma cells. In line with these data, biochemical results indicated that 40-50% of secretogranin II synthesized during 18-h labeling was retained intracellularly over a 4-h chase period and released after treatment with different secretagogues. The most effective stimulus appeared to be phorbol ester in combination with ionomycin in the presence of extracellular Ca(2+), a treatment that was found to produce a large and sustained increase in intracellular calcium [Ca(2+)](i) transients. Our findings indicate that a regulated secretory pathway characterized by (i) the expression and stimulated exocytosis of a typical marker for regulated secretory granules, (ii) the presence of dense-core vesicles, and (iii) the ability to undergo [Ca(2+)](i) increase upon specific stimuli is present in cultured hippocampal astrocytes.

Fine structure of the free-living parakeet pineal in relation to the breeding cycle.

Seasonal changes in the ultrastructure of the free-living Rose-Ringed Parakeet Psittacula krameri pineal were examined in relation to the sub-tropical environment and seasonal reproduction. Dark and light pinealocytes of the presumptive neuroendocrine cell line predominated, while supporting cells, ependymal cells, myelinated and non-myelinated nerve fibers with nerve endings, and regressed photoreceptor elements were also observed. Unlike in pineals of many animals, particularly mammals, the presence of dense-core vesicles (DCVs) with varying core density, and absence of clear vesicles and vacuoles with flocullent material, indicate the involvement of DCVs in the synthesis and secretion of pineals principle/s. In November (pre-breeding) when the day length registered a drop to LD 10:14, pinealocytes showed significantly decreased and smaller DCVs and mitochondria, nuclei with heterochromatin, and greater distribution of glycogen and lipid droplets, all indicating low pineal metabolic activity. During the shortest day regime from December to March, when the birds peaked breeding, the number and size of DCVs and mitochondria increased, and Golgi body-endoplasmic reticulum-lysosome complex (GERL) was very well defined. Images of DCVs suggested possible secretion of pineal principle/s by dissolution, and exocytosis. Coincidence of these features with peak gonadotrophic (circulating LH) and spermatogenic and testicular endocrine activity described previously suggested an active turnover of pineal products during this short day length regime when parakeets breed. In contrast, during the post-breeding season (April onwards), when the day-length increased to LD 13:11 and hypophyseal-gonadal function was down, nuclei and RER continued to show active profile, the Golgi body and associated complex were moderately seen, and the DCVs and mitochondria were significantly smaller and lesser. It is therefore probable that the pineal is an important relay to translate cues related to less drastic sub-tropical environmental change into DCV-linked neurohormones that in turn may be involved in modulating seasonal breeding in parakeets.

Glutamate-like immunoreactivity in retinal terminals of the mouse suprachiasmatic nucleus.

With a view to identifying the neurotransmitter content of retinal terminals within the mouse suprachiasmatic nucleus, a highly specific antiserum to glutaraldehyde-coupled glutamate was used in a postembedding immunogold procedure at the ultrastructural level. Retinal terminals were identified by cholera toxin--horseradish peroxidase transported anterogradely from the retina and reacted with tetramethyl benzidine/tungstate/H2O2, or by their characteristically pale and distended mitochondria with irregular cristae. Controls included model ultrathin sections containing high concentrations of various amino acids. Alternate serial sections were labelled with anti-glutamate and anti-gamma-aminobutyric acid (GABA). Data were analysed by computer-assisted image analysis. Density of glutamate labelling (gold particles per micron2) on whole retinal terminals was > 3 times higher than that on postsynaptic dendrites, and > 5 times higher than that on miscellaneous non-retinal non-glutamatergic terminals in the suprachiasmatic nucleus. The overall density of gold particles over retinal terminals was approximately 3 times higher than that over GABAergic terminals, in which glutamate-like immunoreactivity was mainly mitochondrial. Labelling of vesicles in retinal terminals was almost 5 times greater than the apparent labelling of vesicles in GABAergic terminals, underscoring the location of transmitter glutamate within synaptic vesicles in retinal terminals. In the retino-recipient region of the suprachiasmatic nucleus there was also a small population of non-retinal glutamatergic terminals. Their overall immunoreactivity was similar to or exceeded that of retinal terminals, but morphological features clearly distinguished between these two types of glutamate-containing terminals. The present results indicate that the vast majority of retinal terminals may use glutamate as a transmitter, in keeping with electrophysiological and neuropharmacological data from other sources. The possibility of cotransmitters within retinal terminals, suggested by the presence of dense-core vesicles among the glutamate-containing synaptic vesicles, has still to be addressed.

Ultrastructural study on cell differentiation of the rabbit carotid body.

The development of the carotid body in both fetuses from 13 to 30 days of gestation and newborn rabbits was studied by light and electron microscopy. The carotid body anlage first appeared as a cellular aggregation close to the ventral wall of the third branchial artery on the 14th day of gestation. Type-I cells were recognized as such by the presence of dense-cored vesicles in the cytoplasm on the 16th day of gestation, whereas cells destined to develop into Type-II cells became clearly distinguishable by their typical relationship to the Type-I cells on the 20th day of gestation. Afferent and efferent synapses as well as reciprocal-like synapses between Type-I cells and nerve endings were also observed in perinatal fetuses. Although cell differentiation was almost finished by birth, Type-I cells and their innervation did not seem to be fully matured. Thus, it was concluded that ultrastructurally, the rabbit carotid body during the fetal and perinatal periods was relatively inactive.

Neuroendocrine markers in central nervous system neuronal tumors (gangliocytoma and ganglioglioma).

We studied five cases of central nervous system neuronal tumor, one gangliocytoma and four gangliogliomas, both ultrastructurally and immunohistochemically, using antibodies to neuroendocrine markers including tyrosine hydroxylase (TH), serotonin (5HT), somatostatin (SOM), met-enkephalin (MEK), leu-enkephalin (LEK), substance P (SP), gastrin, vasopressin, oxytocin, vasoactive intestinal polypeptide, adrenocorticotropic hormone and calcitonin. In all cases, the presence of dense-core vesicles (60-250 nm) in the neuronal elements was the characteristic ultrastructural finding. Synapses were observed in two cases. Immunohistochemically, variable numbers of neuronal cells showed positive staining for SOM in five cases, TH, MEK and LEK in three cases, and 5HT and SP in one case each. The others were negative. Positive immunoreactivity for multiple markers was shown in all cases. SOM, TH, 5HT and SP were present in the small- to medium-sized cells, while MEK and LEK were almost exclusively confined to the large cells. Our study clearly indicated that these tumors contained neuronal cells which were not homogeneous with regard to neuroendocrine markers.

Gangliocytoma of the cerebellum

A 31-year-old man presenting with a cerebellopontine angle syndrome underwent subtotal resection of a gangliocytoma of the left middle cerebellar peduncle. The tumor was positive for 68-kD neurofilament protein and vasoactive intestinal peptide. No glial cells were identified on immunohistochemical or ultrastructural examination. Electron microscopic study of neoplastic ganglionic cells revealed the presence of dense core vesicles, synaptic junctions, Hirano bodies, and marked dystrophic changes in the neuritic processes. The nosology of this lesion is discussed. Similarities between the morphological changes shown by mature neoplastic ganglion cells and comparable structural alterations seen in the physiological senescence of nonneoplastic neurons are stressed. (Neurosurgery 22:576-581, 1988)

味神経切断, 吻合後の茸状乳頭ならびに味蕾の変化に関する電子顕微鏡的研究

The fungiform taste buds and some mechanosensory endings in rabbits, which are supplied with chorda tympani and lingual nerves, were examined by means of light and electron microscopy. Three to seven taste buds were present in the dorsal epithelium of a fungiform papilla. The taste bud was composed of the type I, the type II, the type III cells and the basal cells. The type III cell was characterized by the presence of dense-core vesicles and synaptic vesicles, and showed afferent synaptic contact with nerve endings. Thus, it was suggested that the type III cell is involved in the chemoreceptive function. The presence of numerous free nerve endings in the extragemmal epithelium and mechanosensory endings, and of encapsulated corpuscles and Ruffini-like endings in the connective tissue of the fungiform papillae suggested that the fungiform papillae of the rabbit were highly adapted to both taste and mechanical sensation. The fungiform papillae were examined by light and electron microscopy from 1 week to 6 months after denervation followed by microsurgical reinnervation of both chorda tympani and lingual nerves. Further, these results were compared with those of experiments which were carried out in the same manner except for the microsurgical reinnervation. The first remarkable finding was the appearance of an ulcer in the dorsal surface of the tongue of the denervated side after the first week following denervation. The ulcer remained visible for about 10 weeks after the operation, and a severe defect of the tongue of the denervated side was observed occasionally. Taste buds and mechanosensory endings in fungiform papillae disappeared completely after the first week following denervation. The dorsal epithelium of the fungiform papillae showed filiform-like structures because of the hyperparakeratotic changes. However, 15 weeks after denervation followed by microsurgical reinnervation, the fungiform papillae regained their normal appearance. Encapsulated corpuscles and Ruffini-like endings regenerated in the connective tissue of reinnervated fungiform papillae in this experimental period, although these nerve endings were small in size and in number. 5 months after microsurgical reinnervation, regeneration of the taste buds was observed in the dorsal epithelium of the fungiform papillae. Ultrastructure of regenerated taste buds was essentially identical with that of normal taste buds, although the numbers of regenerated taste buds and their constituents were smaller than those of normal fungiform taste buds. In groups where no microsurgical reinnervation was carried out, regeneration of mechanosensory endings in the connective tissue was observed 5 months after denervation. Regeneration of taste buds, however, could not be observed during the experimental periods, even 6 months after denervation. The present study showed morphologically that the rapid and reliable regeneration of taste buds and mechanosensory endings in denervated fungiform papillae by chorda tympani-lingual nerves could be expected only when the proximal and distal cut ends of these nerves were repaired microsurgically.

The innervation of the external urethral sphincter; an ultrastructural study in male human subjects.

Innervation of the external urethral sphincter (EUS) was studied in male human subjects. In the region of EUS at the distal end of prostatic urethra, a large axon bundle surrounded by perineurium was evident in the intramural connective tissue gap. Because of the presence of dense core vesicles, the small nonmyelinated axon profiles in the bundle were considered to be adrenergic. After ramifications to smooth musculature, the axons were traced to the EUS. In the EUS, axon bundles containing many non-myelinated axons were recognized as a sole autonomic nerve among the striated muscle cells. A single or at most two or three axons were surrounded by a Schwann cell, and some possessed dense core vesicles which suggested an adrenergic function. These autonomic adrenergic nerve ends formed surface junctions with the striated muscle of EUS. The clinical relevance of these data are discussed.

Monoamine-containing basal cells in the taste buds of the newt Triturus pyrrhogaster

Monoamine-containing cells were examined by fluorescence histochemistry and electron microscopy. Two or three serotonin-like fluorescent cells were located just above the basal lamina and failed to reach the free surface of the taste bud. Ultrastructurally this cell type was characterized by the presence of dense-cored vesicles and finger-like cytoplasmic processes. Many characteristics of Merkel cells were present.

The ultrastructural localization of enkephalin and substance P immunoreactivities in the nucleus tractus solitarii of the cat.

In the medial and commissural subdivisions of the nucleus tractus solitarii enkephalin and substance P immunoreactivities were localized within synaptic terminals, unmyelinated axons, and neuronal cell bodies. Both enkephalin and substance P immunoreactivities were contained within synaptic terminals which had a mixture of small clear vesicles and dense core vesicles. The presence of dense core vesicles within both the enkephalin- and substance P-immunoreactive terminals was a consistent feature, although they were not associated with the actual synaptic junction. While enkephalin- and substance P-immunoreactive terminals shared a similar morphology, their respective distributions along the dendritic tree were quite distinct. Enkephalin-immunoreactive terminals contacted mainly the cell body and proximal portions of the dendritic tree. In contrast, substance P-immunoreactive terminals synapsed predominantly with spines and shafts of small to medium-sized dendrites. Few substance P-immunoreactive terminals contacted proximal dendrites and they were never presynaptic to the neuronal cell body. This apparent segregation of synaptic terminals on neurons suggests that enkephalin synapses have a more pronounced effect than substance P terminals.

Uptake of 3H-glycine in the outer plexiform layer of the retina of the toad, Bufo marinus.

The uptake of 3H-glycine in the retina of the toad, Bufo marinus, was investigated by light and electron microscopical autoradiography. Uptake of 3H-glycine was very prominent in large cell bodies in the inner nuclear layer as well as in discrete clusters in both the outer plexiform layer (OPL) and the inner plexiform layer. This pattern in similar to that described for 3H-glycine-accumulating putative interplexiform cells in goldfish, frog, and Xenopus retinas. Electron microscopical autoradiography of the OPL revealed large, grain-containing varicosities which had electron-lucent cytoplasm and contained both small, agranular and large, dense-core vesicles. The varicosities made extensive en passant and spine synapses in the OPL. Definitive identification of their postsynaptic targets was not achieved. However, autoradiographic analysis with 3H-GABA uptake as well as electrophysiological evidence suggests that axons but not cell bodies or dendrites of 3H-GABA-accumulating horizontal cells (H1 cells) are postsynaptic targets of the varicosities. The presence of dense-core vesicles in the varicosities suggested co-occurrence of glycine and a biogenic amine or neuropeptide. The indirect immunofluorescence technique was used to determine whether any such substances were present in the OPL of the toad retina. However, no specific labeling was found in the OPL for any of 19 substances tested. The extensive synaptic output provided by glycine-accumulating varicosities in the toad OPL may indicate an important role of glycine in the synaptic function of the distal toad retina. We suggest that these varicosities derive from a presumably glycinergic interplexiform cell.

Dense core vesicles during photoreceptor development.

The presence of dense core vesicles in the terminal expansions of photoreceptors in development is described in the chick embryo retina, from the 16th to the 18th day of incubation.

Ultrastructure of the pineal gland of the fox.

The ultrastructure of the pineal gland of the fox was examined and compared with that of other mammals. The pineal gland of the fox is composed of two different populations of pinealocytes (I and II). The pinealocytes I were distributed homogeneously throughout the parenchyma, while pinealocytes II were located generally near blood vessels. A Golgi apparatus, granular endoplasmic reticulum, mitochondria, lysosomes, centrioles, and cilia were present in both cell populations. A characteristic feature of pinealocytes I was the presence of dense-core vesicles, presumably of Golgi origin; whereas glycogen deposits and pigment granules were common features of pinealocytes II. In addition to the pinealocytes, the parenchyma contained fibrous astrocytes. The capillaries of the pineal gland of the fox consisted of a nonfenestrated endothelium. Numerous never fibers, presumably adrenergic, were observed throughout the parenchyma.

Presence of dense-core vesicles in pinealocytes of the cat.

Dense-core vesicles, 90–200 nm in diameter, were found in pinealocytes of the cat. They were present both in perikarya and cell processes.

The diagnosis and surgical management of olfactory neuroblastoma: the role of craniofacial resection.

The diagnosis and clinical management of olfactory neuroblastoma are discussed. Computerized tomography is a valuable means of assessing the pre-operative extent of disease and the efficacy of subsequent major surgery. The tissue diagnosis is facilitated by electron microscopy, demonstrating the presence of dense core vesicles and cytoplasmic filaments. Radiotherapy followed by craniofacial resection is the treatment of choice for olfactory neuroblastomas that are locally extensive at presentation.

Monoamine precursor uptake in ganglia of a mollusc Anodonta cygnea L. (Bivalvia) An autoradiographic and ultrastructural study.

In vitro-uptake and localization of 3H-5 hydroxytryptophan and 3H-dihydroxyphenylalanine have been investigated by means of light microscopic autoradiography in the central nervous system of the freshwater mussel, Anodonta cygnea L. Accumulation of the labelled monoamine precursors could be observed both over neuronal perikarya and neuropil in the ganglia. 3H-5-hydroxytryptophan and 3H-dihydroxyphenylalanine were taken up by neurons characterized by a small size (10-20 micrometer), a polygonal form and an intense staining with toluidine blue. Labelled axon branches situated either in the neuropil or between nerve cells in the cortex exhibited only a moderate activity, frequently of a diffuse character. After pretreatment fro 24 h with colchicine, silver grains were also seen over a neuronal perikarya and some axon profiles. This shows that local uptake of the monoamine precursors and, presumably, the synthesis of the monoamines takes place both at perikaryal and axonal/terminal level. The nerve cells accumulating the monoamine precursors contain a nucleus with rich chromatin content and a deeply invaginate nuclear envelope. Further ultrastructural characteristics of these neurons are their well-developed rough endoplasmic reticulum system, a great number of mitochondria and glycogen granules, and the presence of dense-cored vesicles of different types. All those features are responsible for the electron-dense appearance of the cytoplasm.

The cerebral cortex of the reeler mouse embryo. An electron microscopic analysis.

The cerebral cortex of reeler (rl/rl) and control (rl/+) embryos has been examined at embryonic day 14 (EO = day of the plug). At this early stage, the cells of the cortical plate in reeler are characteristically maloriented when compared to the controls but, despite this malorientation, they assume a normal differentiation; in particular the regions of contact between the cells or the cortical plate are not different from the controls. The marginal zone and the subcortical plate of reeler are less well delimited than in normal fetuses, yet each of these layers contains the same contingent of cells and fibers as the normal embryos. In particular an early afferent fiber system is visualized in reeler and shows the same characteristics as in normal embryos: weak osmiophily, presence of dense core vesicles and of multilocular dense vacuoles, formation of asymmetric synapses in the marginal zone. The nature of these fibers is unknown but it is suggested that they could correspond to the aminergic system described in rats at the same developmental stage. The observations reported, though compatible with the hypothesis that the reeler gene acts upon a cell recognition-adhesion phenomenon between CP cells, do not provide a positive evidence for it and do not exclude other explanations.