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Membrane retrieval by vacuoles after exocytosis in the neural lobe of Brattleboro rats.
The possible role of microvesicles and vacuoles in the recapture of membrane after pituitary hormone release by exocytosis has been studied in homozygous Brattleboro rats. These mutant animals are unable to synthesize vasopressin and exhibit a steady state hypersecretion of oxytocin from the neural lobe as a result of the osmotic imbalance caused by their diabetes insipidus. This can be converted to a second steady state which approximates to the rate of secretion found in normal Long Evans rats by the administration of exogenous vasopressin daily for 30 days. In the Brattleboro rat, presumptive oxytocinergic nerve endings contain typical 160-170 nm diameter neurosecretory granules; other magnocellular nerve endings contain a population of smaller (approximately 100 nm diameter) dense-cored granules. The number of dense-cored granules was reduced in both types of nerve ending in the hypersecreting Brattleboro rats, but increased as a result of vasopressin treatment to levels which, for the classical neurosecretory granules, approximated that found in Long Evans rats. The microvesicle population of the nerve endings was essentially similar in quantitative terms in all the three groups (i.e. hypersecreting Brattleboro rats; vasopressin-treated Brattleboro rats and Long Evans controls). The number of vacuoles, on the other hand, was increased in nerve endings in the hypersecreting animals but reduced to levels found in Long Evans rats in the Brattleboro animals treated with vasopressin. Furthermore, the size of the vacuoles was comparable to the size of the dense-cored granules contained in the nerve endings. These changes in the vacuole population are exactly those that would be predicted for an organelle responsible for recapture of the granule membrane. We therefore conclude that membrane retrieval after exocytosis of neurosecretory granules in the neural lobe is achieved by vacuoles and that these organelles probably retrieve the membrane of the granule intact.
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Scanning and transmission electron microscopy of intraventricular dendrite terminals of hypothalamic cerebrospinal fluid contacting neurons in Triturus vulgaris.
A scanning (SEM) and transmission electron microscopic (TEM) study of the ventricular wall of the hypothalamus of Triturus vulgaris was performed with special regard to the intraventricular dendrite terminals of the cerebrospinal fluid (CSF) contacting neurons of the preoptic area (magnocellular and parvocellular preoptic nuclei), the infundibular lobe (anterior periventricular nucleus, infundibular nucleus), and the paraventricular organ. In the preoptic area and infundibular lobe, the terminals were knob-like or club-shaped, of various sizes (diameter about 0,5 to 3,0 micrometer) and located immediately above the ependyma. Ultrastructurally, they may contain dense-core vesicles of varying sizes. The CSF contacting dendrite endings of the paraventricular organ built up a supraependymal labyrinthic layer which could be divided into a rostral crest-like part and a caudal flat and broad division. In both parts, three main types of terminals of various size and shape could be distinguished: a) ramifying, b) elongated, and c) bulb-like dendrite endings which also differed by their TEM structure. The bulk-like terminals, first of all the small ones, originated from the distal part of the nucleus of the organ (nucleus organi paraventricularis) while the other two types took their origin from its intra- and subependymal part. In all areas investigated, each intraventricular dendrite ending gave rise to a solitary cilium (type 9 X 2 + 0). It differed from the ependymal kinocilia by both SEM and TEM characteristics. In the paraventricular organ, the neuronal cilia were hidden inside, or below the supraependymal layer of terminals. There were intraventricular axons which formed synapses on CSF contacting dendrite endings of both parts of the paraventricular organ. Free intraventricular neurons, further ependymal areas heavily or scarcely ciliated, were described. The CSF contacting dendrite terminals were predominantly present near ventricular recesses and in regions where the ependyma was scarcely ciliated.
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Abnormalities of central axons in a dysmyelinative rat mutant.
The absence of normal myelin from the CNS of the dysmyelinative rat mutant, md, is associated with axonal abnormalities including organelle-poor and organelle-rich spheroids (OPS and ORS, respectively), wrinkling of the axolemma, persistence of glycogen aggregates and vacuoles in cerebellar mossy fiber terminals, and coalescence of synaptic vesicles in terminal boutons of the nucleus interpositus. OPS have a special predilection for medullary pyramid and the axons of Purkinje cells and further differ from ORS in their possession of nematosomes and in their lack of neurofilaments, microtubules, and degenerating mitochondria. Purkinje cells of md fail to increase in size after 30 days postnatal age and, unlike these neurons in normal neonatal rats, may have massed or dispersed granules of cytoplasmic glycogen which persist for at least 86 days postnatally. Morphometric study of axons of medullary pyramid and cervical corticospinal tract at 19-43 days of age shows a shift in frequency to axons of smaller size in md, as compared to age-matched controls, except that approximately 1% of md axons are larger than any encountered in controls. Finally, the pyramidal axons of md at 43 days of age have a significantly larger area of axoplasm occupied by mitochondria than obtains for the control condition. We conclude that the described abnormalities are secondary to the lack of a myelin investment and/or the loss of oligodendrocytes.
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Species differences in vacuolation of the choroid plexus induced by the piperidine-ring drug disobutamide in the rat, dog, and monkey.
A subchronic oral toxicity study of disobutamide, a piperidine ring compound with antiarrhythmic activity, was conducted at doses of 30, 100, and 250 mg/kg in rats, 45 mg/kg in dogs, and 90 mg/kg in monkeys. Numerous vacuoles were observed in various organs such as the liver, kidneys, heart, lungs, spleen, thymus, stomach, and choroid plexus in these animals. The epithelium of the choroid plexus (CP), however, showed severe vacuolation in rats and monkeys but not in dogs. The vacuoles corresponded to enlarged and myelin-figured lysosomes observed by electron microscopy, revealing morphological characteristics which have been reported as drug-induced phospholipidosis. In a further study, the drug penetration to cerebrospinal fluid (CSF) and the drug concentration in CP were examined in these animals. Daily po doses of 250, 45, and 90 mg/kg were, respectively, administered to rats, dogs, and monkeys to maintain approximate equivalency in peak blood concentrations across species, over a course of 35 days. The concentration of the drug in the CP was higher in rats and monkeys than in dogs, and the CSF/serum ratio of the drug concentration was extremely high in rats. The uptake of the drug by the CP in vitro was high in rats, monkeys, and dogs, in this order. In dogs, both direct contact of the drug with the CP during incubation and intraventricular administration induced vacuolation in the epithelium. From these results it was concluded that differences of the drug's penetration into the CSF and its uptake by the choroid plexus epithelium are responsible for the species differences of CP vacuolation in the animals.
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Ultrastructure of Golgi-impregnated and gold-toned spiny and aspiny neurons in the monkey neostriatum.
Golgi-impregnated, gold-toned spiny and aspiny neurons in the monkey neostriatum were deimpregnated and examined at the electron microscope level. Spiny type I neurons have relatively large nuclei with few indentations and aggregates of chromatin under the nuclear membrane which in some regions give the appearance of a dark rim. The small quantity of surrounding cytoplasm is poor in organelles. Aspiny type I neurons have eccentric, highly indented nuclei. The relatively large proportion of cytoplasm is rich in organelles especially Golgi apparatus and rough endoplasmic reticulum which often appears in stacks. Synapses with symmetric membrane densities are common on the somata of spiny type I neurons. Those on the proximal and distal dendritic shafts are few in number and asymmetric, and those on spines more frequent and primarily asymmetric. Aspiny type I neurons have few synapses on their cell bodies. Proximal and distal dendrites, however, are contacted by numerous profiles which contain small round vesicles and make both symmetric and asymmetric synapses. The same axon terminals also synapse with dendritic spines of spiny neurons, indicating that an input, most likely of afferent origin, is shared by both cell types. Other less frequently occurring profiles forming symmetric membrane densities also contact the dendrites of aspiny and spiny neurons. The axon hillocks and initial segments of both neuronal types receive a synaptic input, which is more common on spiny cells. Results offer unequivocal evidence for the differences in the ultrastructure of these two most common categories of medium-size neostriatal neurons, which may help in their proper identification in standard material, as well as information on the types and distributions of synaptic inputs onto these neurons. Moreover, the findings clarify some controversies in the literature probably originating from observations on a mixed population of cells of medium size.
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Pools of serotonin in the pineal gland of the mouse: the mammalian pinealocyte as a component of the diffuse neuroendocrine system.
In the pineal gland of the mouse the distribution of serotonin (5-HT) and its eventual relationships to a protein secretion were examined by means of fluorescence histochemical (Falck-Hillarp) and ultracytochemical (chromaffin and argentaffin) methods. (1) Yellow formaldehyde-induced fluorophores, characteristic of high concentrations of 5-HT, were found in pinealocytes, interstitial cells and sympathetic adrenergic nerve endings. The 5-HT content was studied according to the circadian variations and different drug treatments. (2) By use of ultracytochemical methods in untreated control mice precipitates indicative of 5-HT were found over dense-core vesicles (DCV; mean diameter: 100 nm) of the pinealocytes and the vesicular compartment of the sympathetic nerve fibers. After reserpine and p-CPA treatments, the reactivity of the DCV disappeared, but the protein secretion accumulated within the DCV was still present. After nialamide treatment the precipitates in the DCV increased and, similar to control mice, masked the protein secretion. Taking into account the specificity and sensitivity of these three complementary methods, previous biochemical data in mammals and ultracytochemical data in submammalian vertebrates, synthesis, storage, catabolism and release of 5-HT in the mouse and hamster pineals are discussed at the cellular level. Different pools of 5-HT are present: agranular and granular in pinealocytes and sympathetic nerve endings; agranular in interstitial cells. In the mouse, only a small portion of the total 5-HT content appears to bae secondarily taken up by the DCV. Within the DCV, 5-HT is possibly bound to a protein secretion of unknown significance (peptidergic neurohormone?). In the pinealocytes, which are sensitive to a large variety of inputs, processes of indole and protein secretion are found. These cells apparently are the recepto-secretory elements of the mammalian pineal gland. They can thus be classified as a member of the diffuse neuroendocrine system of the paraneuron group.
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[Ultrastructure of the neurohypophysis of the rat in postnatal ontogenesis].
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Progranulin enhances the engraftment of transplanted human iPS cell-derived cerebral neurons.
Cerebral organoids (COs) in cell replacement therapy offer a viable approach to reconstructing neural circuits for individuals suffering from stroke or traumatic brain injuries. Successful transplantation relies on effective engraftment and neurite extension from the grafts. Earlier research has validated the effectiveness of delaying the transplantation procedure by 1 week. Here, we hypothesized that brain tissues 1 week following a traumatic brain injury possess a more favorable environment for cell transplantation when compared to immediately after injury. We performed a transcriptomic comparison to differentiate gene expression between these 2 temporal states. In controlled in vitro conditions, recombinant human progranulin (rhPGRN) bolstered the survival rate of dissociated neurons sourced from human induced pluripotent stem cell-derived COs (hiPSC-COs) under conditions of enhanced oxidative stress. This increase in viability was attributable to a reduction in apoptosis via Akt phosphorylation. In addition, rhPGRN pretreatment before in vivo transplantation experiments augmented the engraftment efficiency of hiPSC-COs considerably and facilitated neurite elongation along the host brain's corticospinal tracts. Subsequent histological assessments at 3 months post-transplantation revealed an elevated presence of graft-derived subcerebral projection neurons-crucial elements for reconstituting neural circuits-in the rhPGRN-treated group. These outcomes highlight the potential of PGRN as a neurotrophic factor suitable for incorporation into hiPSC-CO-based cell therapies.
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The ultrastructural morphology of the squirrel monkey area postrema.
Examination of the squirrel monkey (Saimiri sciureus) area postrema (AP) revealed this circumventricular organ to be primarily composed of two types of glial cells and a single type of neuronal element. No pattern of neuronal arrangement could be discerned, however, this cell type was frequently observed in close relation to the perivascular spaces. The neuronal elements, although slightly larger than the glial cells, were characteristically less electron dense. The neurons routinely displayed an infolded nuclear membrane, a single nucleolus and the normal complement of subcellular organelles. Synaptic terminals were numerous, and both axo-somatic and axo-dendritic varieties were observed with the latter being more numerous. Both clear-cored and dense-cored vesicles could be observed in the same ending. Unmyelinated neuronal processes were the predominant type within the interior of the AP, although myelinated processes were also regularly present. Non-neuronal elements with the AP resembled CNS astrocytes and were as numerous as the neuronal elements. This cell type appeared to envelope completely the vasculature and separated the parenchyma from the perivascular spaces. The ventricular surface of the AP was covered by modified ependyma which lacked kinocilia but frequently demonstrated microvillar projections. Opposed ependymal cell membranes showed interdigitations, and zonula adherens-type cell junctions connected the ependymal cells near the ventricular lumen. Two types of bulbous projections were observed in the ventricular lumen close to the ependymal surface. The most characteristic feature of the AP, however, was its vascularity. Perivascular spaces surrounding fenestrated capillaries contained fibroblasts and collagen. The vascular endothelium routinely demonstrated pinocytotic activity, and the basal lamina was prominent.
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Cytology and time of origin of interstitial neurons in the white matter in infant and adult human and monkey telencephalon.
The fine structure, synaptic relationships, distribution and time of origin of interstitial neurons situated within the white matter subjacent to the visual, somatosensory and motor cortices were studied in the human and monkey telencephalon. The analysis was carried out on Nissl-stained serial sections, rapid Golgi impregnations, by acetylcholinesterase (AChE) histochemistry, electron microscopy and [3H]thymidine ([3H]TdR) autoradiography. Interstitial neurons have a similar distribution, morphology and histochemistry in both human and monkey telencephalon. Their highest density and the most extensive distribution is found in the neonatal period in both species. The number of interstitial neurons decreases during infancy, but numerous cells remain in the adult. Two types of interstitial neuron can be recognized in Golgi preparations: polymorphic cells, usually situated close to the cortex and fusiform cells, located predominantly in the depths of the white matter. The polymorphic cell type is prevalent during neonatal and infant stages, while fusiform cells are relatively more numerous in the adult. Interstitial cells have ultrastructural features and organelles typical of neurons of the central nervous system with well-defined axosomatic and axodendritic synapses of both symmetrical and asymmetrical types. About 20% fo the interstitial cells show strong specific AChE activity. Autoradiographic analysis of postnatal monkeys exposed to [3H]TdR at various embryonic (E) and early postnatal days indicates that interstitial neurons which lie beneath the visual and somatosensory-motor cortices are generated between E38 and E48. Contrary to the prevailing notion that interstitial neurons are the latest generated cells arreste during migration across the maturing white matter, they prove to be produced at the end of the first third of the 165-day gestation in the rhesus monkey concomitantly with the generation of neurons destined for the deep neocortical layers. These findings raise the possibility that interstitial cells represent a vestige of the transient embryonic subplate layer.