Morphometric and electron microscopic studies of the secretory granules of neurosecretory cells of the supraoptic and paraventricular nuclei of white rats and mice]

The morphometrical and electron microscopic analysis of secretory granules in the perikaryons of neurosecretory cells of the supraoptic and paraventricular nuclei in male rats and mice has shown than in the cells of these nuclei in both species of animals there occur secretory granules of the same kind and size. Therefore this method fails to determine which of them contain oxytocin and which of them contain vasorpressin. The neurosecretory granules located in the Golgi apparatus zone are of a less size and have more osmiophilic cnetral material than the granules localized on the periphery which mainly have granular central material and are of a greater size. The distinctions in the size and type of secretory granules are associated with certain stages of their "maturation". Granular particles appear to be "swallen", more active forms of storing neurohormones. The presence of larger granular particles in the supraoptic nucleus of mice allows to suggest greater reactivity of this nucleus than in rats which is likely to be associated with a higher ability of mice, as compared with rats, to adaptate to disturbances in water-salt metabolism.     

Neurosecretion

Summary Ultrastructural specializations characteristic of sites of release of neurosecretory material from axons were examined in several species of blattarian insects. Discharge of such material may take place within or outside of neurohemal organs and is not restricted to fiber terminals. Structurally distinctive areas serving this function occur intermittently and may be more or less transient. Many of these specialized zones face the extracellular stroma that forms sheaths and partitions of neurohemal organs (corpora cardiaca, perisympathetic organs), others contact various cellular elements (nerve fibers with or without neurosecretory granules, glial cells, non-neural endocrine cells).Irrespective of the milieu, these sites of release are characterized by small electron lucent vesicles clustered near the internal surface of the plasma membrane, and by variously shaped accumulations of electron dense material on either side of this membrane. These ultrastructural features are strikingly similar to those of the presynaptic component of conventional interneuronal junctions. However, the functional implications of this morphological resemblance seem to be limited. In neurosecretory systems, physiological phenomena comparable to chemical transmission are out of the question in the absence of postsynaptic cells.In peptidergic neurons of the insect species used in the present study, as in those of various mammals examined by other investigators, the small vesicles observed seem to be the result of fragmentation of neurosecretory granules prior to the discharge of their contents. The presence of variable intermediate stages speaks against a cholingergic role of these synapticlike vesicles at least some of which seem to contain neurosecretory material instead of a neurotransmitter. Furthermore, the variously shaped intra- and extracellular dense material in synaptoid areas seems to represent a neurosecretory product in transit and is therefore not equivalent to dense material customarily found within or on either side of the regular synaptic cleft.Sites of release not directly affiliated with the stroma often share a common narrow gap between adjoining neurosecretory fibers and face each other in mirror image fashion. It is here where the distinction from regular synapses is sometimes more difficult to make because the structural elements of one side of the paired complex may mimic postsynaptic dense material. A further source of difficulty in the interpretation of special contact areas of this sort is the existence of unusual junctions between two classes of neurosecretory neurons (B and A fibers) in which pre- and postsynaptic details are discernible. These, and synaptoid junctions with non-neural endocrine effector cells, seem to serve for the dispatch of local neurosecretory signals that resemble, but are nevertheless apart from, conventional neurohumoral communication. The special neurosesecretory products involved here do not qualify as neurohormones.Synaptoid neurosecretory contact areas with pre- and postsynaptic features should be classified as a group distinct from another group in which the postsynaptic component is absent.Supported by grants AM-3984, NB-00840, and NB-05219 from the U.S.P.H.S.I am greatly indebted to Mrs. Sarah Wurzelmann for her excellent technical assistance.     

On degeneration of peptidergic neurosecretory fibres in the albino rat

Three types of degenerating peptidergic neurosecretory fibres have been found in the posterior pituitary of chronically dehydrated albino rats. "Dark" neurosecretory fibres and their swellings contain neurosecretory granules, neurotubules, shrunken mitochondria and diffusely distributed fine dense material. Some swellings are filled with synaptic vesicles and/or conglomerations of dense membranes. The transitional forms exist between these fibres and extracellular accumulations of electron dense material. Synaptic vesicles, single neurosecretory granules, lipid-like droplets and lamellar bodies occur in the latter. Some neurosecretory fibres and swellings have numerous polymorphous inclusions arising due to degradation of secretory inclusions and organelles, mitochondria and neurotubules in particular. "Dark" neurosecretory elements and those with numerous polymorphous inclusions are enveloped by pituicyte cytoplasm. Sometimes the plasma membranes both of the pituicytes and neurosecretory fibres are destroyed or transformed into a multi-membrane complex. It is assumed that pituicytes may phagocytize degenerating neurosecretory elements. N urosecretory fibres with a locally dissolved neuroplasm and/or large lucent vacuoles seem to be due to axonal degeneration by the "light" type. These neurosecretory elements, the largest of them in particular, may transform into large cavities bordered by a membrane and containing flake-like material and single-membrane vacuoles. Degeneration of neurosecretory elements seems to occur mainly due to hyperfunction of the hypothalamo-hypophysial neurosecretory system.     

Cytologie du lobe anterieur de l'hypophyse du chien

Summary Detailed histochemical studies have been made on the distribution of various enzymes such as phosphatases, cholinesterases, glycolytic enzymes and respiratory enzymes in various components of the hypothalamus with special reference to the supraoptic and paraventricular nuclei of the Squirrel Monkey. Cytological studies have also been made by the McManus, Einarson, Gomori and Bargmann methods.A few neurons of these nuclei showed scanty Gomori-positive material in the cytoplasm for the Gomori and Bargmann methods. Nissl granules were located in the peripheral cytoplasm of most neurons. No glycogen granules were observed in these neurons. For these reasons, the Squirrel Monkey, like the rat, may not be a suitable species for the study of neurosecretory phenomena.The axons of these neurons were negative for the specific cholinesterase test, though the perikaryon and some parts of the processes gave a moderately positive reaction. These neurons may be non-cholinergic and the cholinergic fibers from an unknown nucleus may end in synapses on their cell bodies. Blood vessels and glial cells in the neurosecretory nuclei showed non-specific cholinesterase activity. This enzyme may hydrolyze the acetylcholine which has escaped splitting by specific cholinesterase. Alkaline phosphatase and acid phosphatase in these neurons may be involved in the metabolism concerned with the production of neurosecretory material. The neurons may be physicochemical receptors and may get enough energy and raw material to synthesize the neurosecretory material from the rich blood supply. Neurons of the supraoptic and paraventricular nuclei as well as other hypothalamic neurons, like neurons of other regions of the brain, are well equipped with the enzymes of the glycolytic pathways and the tricarboxylic acid cycle. Since the glial cells of these nuclei have amylophosphorylase activity and glycolytic pathways, they may work as energy donators to the neurons of the neurosecretory nuclei. T. R. Shanthaveerappa in previous publications.     

Some histochemical reactions on the neurosecretory cells of Rivulogammarus syriacus chevreux (Crustacea, Amphipoda).

The histochemistry of neurosecretory material in neurosecretory cells of Rivulo-gammarus syriacus has been investigated. Histochemically, these cells contain different neurosecretory substances. The material in A and C cells consists of mucopolysaccharides and lipids, that in B, B' and D cells there are a protein containing cystine S-S group, mucopolysaccharides and lipids, and that in E cell contains a weak protein with cystine S-S group, a substance showing beta metachromasia and lipids. The lipids are found in all the cells.     

Neuroendocrine control of oocyte development in Poecilocera picta (Insecta: Orthoptera)

The neuroendocrine control of oocyte development in Poecilocera picta Fab. has been described. The secretory activity of the A type of neurosecretory cells has been correlated with ovarian development. In P. picta during the first four days after emergence the neurosecretory material is seen passing down the axons but the cells are largely devoid of neurosecretory material. When the oocytes are developed the A type cells are with stored neurosecretory material.
In P. picta the mature males do not appear to accelerate the process of maturation in females. The females which are reared without males or with castrated males also mature at the same time as the females which are reared with males. The corpus allatum also enlarges and decreases at the same period. The number of resorptive bodies is much more in the females which are reared with castrated males or without males. There appears to be some correlation in the secretion of the neurosecretory material, copulation, and the appearance of resorptive bodies. In P. picta the role of the mature male is only in copulation which very likely allows the cell to synthesize and secrete and release a large amount of neurosecretory material whose discharge in the haemolymph enables a successful development of the oocyte. Corpus allatum appears to be controlled by a precursor from the brain.
Cautery of the cerebral neurosecretory cells, allatectomy and sectioning of the nerves have been done to find out the role of neurosecretory material.     

In vitro peptidergic neurons from the adult locust pars intercerebralis: Morphological and immunocytological studies

Primary cell cultures were prepared from a major neurosecretory center of the adult locust brain, the pars intercerebralis, in order to characterize neurosecretory cells growingin vitro. Individual pars intercerebralis could be removed free of surrounding tissue and dissociated by mechanical treatment. Mature neurosecretory neurons of different sizes regenerate new neurites during the initial three daysin vitro in serum-free medium. They show a tendency to sprout one primary neurite from which fine processes develop. By means of electron microscopy, we observed the integrity of the cellular organelles, indicating that cultured neurons are healthy, and we were able to distinguish three types of neurosecretory neurons on the basis of the ultrastructural aspects of the neurosecretory material. These three types have the same ultrastructural characteristics asin situ neuroparsin, ovary maturing parsin and locust insulin related peptide neurons. Immunogold labelling at the electron microscopic level, using the two available specific antibodies, anti-neuroparsin and anti-ovary maturing parsin, confirms the morphological characterization of neuroparsin and ovary maturing parsin cells. These results show for the first time that cultured locust neurosecretory neurons behave like thosein vivo, in terms of their ultrastructure and immunocytochemistry. Moreover, the presence of recently-formed neurosecretory material both in the Golgi zone of the perikaryon and in the neuronal processes indicates that cultured neurons have functional capacity since they are able to synthesizede novo and to transport the neurosecretory material along the neurite. Thus our well-characterized culture system provides a suitable invitro model to investigate the secretory mechanism of locust neurosecretory neurons.     

The structure and cytochemistry of the neurosecretory cells in the intertidal gastropod Thais bufo (lamarck)

Three types of neurosecretory cells ('A', 'B' and 'C' cells) have been distinguished in the central ganglion of Thais bufo. A few homogenous groups are met with and the rest are all heterogenous groups. The histochemical observations reveal that the neurosecretory material is rich in carbohydrates, disulphides, protein bound amino groups, glycoprotein and lipids. Thus the neurosecretory material seems to be a lipoprotein--glycoprotein complex.     

Ultrastructure of the hypophysiotropic region of the hypothalamus in early ontogenesis in rats]

The results of electron microscopic studies have shown that the 16--18 days old rat embryos already have in the hypophysiotropic area some structures necessary for the realization of neuroendocrine regulations. In the arcuate nuclei, the neurosecretory cells differentiate which are capable to synthesize specific neurosecretory granules of 800--1,000 A in diameter. In the median eminence, the primary portal capillaries develop with which tanicytes and a few axon terminals make contact. One can see in the tanicytes the signs of active transport and accumulation of electron dense polymorphic material. All these phenomena are strengthen during the subsequent development. Hence, several days are before birth the neurosecretory and glial elements of the embryos show the signs of functional activity which strengthen during ontogenesis and are expressed most distinctly in the adult animals.     

Neurosecretory cells in the optic lobe of two aquatic coleopteran species,Dineutes indicus aube and Cybister rugulosus redt

In each optic lobe and optic peduncle of two aquatic beetles viz. Dineutes indicus and Cybister rugulosus the neurosecretory cells are observed with the help of various histochemical techniques. These cells are arranged to form a discrete group. A group in the optic lobe of both species contains about 25 to 30 neurosecretory cells. On the basis of staining properties the neurosecretory cells are classified into A and B types. These cells stain with chrome haematoxylin-phloxine and paraldehyde fuchsin, but do not stain with azan. Histochemically, the neurosecretory material is positive for proteins and shows a negative reaction for 1,2-glycols. The cells show variations in RNA contents in correlation with the state of secretory activity. Axons of the neurosecretory cell group of the optic lobe are observed directed to the optic peduncle. The axonal tract from neurosecretory cells in the optic peduncle runs towards the lateral margin of the brain.     

Histology, histochemistry, and ultrastructure of neurosecretory cells in the optic lobe of the cockroach, Periplaneta americana

In the region of the distal optic chiasma of each optic lobe of Periplaneta americana, there is a group of about 120 monopolar neurosecretory cells. These cells do not stain with paraldehyde fuchsin but remain acidophilic after oxidation. They stain red or sometimes indigo with the azan technique. Histochemically, the neurosecretory material is positive for protein and the amino acids tryptophan and arginine but negative for 1, 2-glycols and strongly acidic groups. At the ultrastructural level, the cytoplasm of the cells contain many elementary neurosecretory granules 100 to 170 nm in dia. The cells also contain well-developed Golgi bodies and endoplasmic retieulum. The axons from these cells run toward the interior of the optic lobe. In this region, axons containing dense granules (mean diameter 70 nm) and synaptic vesicles synapse onto the axons from the neurosecretory cells. The neurosecretory axons then cross over to the anterior side of the optic lobe and run towards the brain. The function of these neurosecretory cells is unknown, but they may be involved with photoperiodically controlled activity rhythms.     

Cytophysiological study of neurosecretory and pheromonal influences on sexual development in Ophryotrocha puerilis (Polychaeta,Dorvilleidae)

Summary

Prominent secretory nerve endings are found at the posterior margin of the supraesophageal ganglion in the protandric polychaete, Ophryotrocha puerilis. Solitary juveniles developing as primary males, and then as females, accumulate neurosecretory material in the nerve endings which thereby swell and become filled with granules. Females maintained in mass culture have similar terminals, whereas in secondary males (males which had been females before), these axon terminals are very small and contain no material. When such males are isolated, they accumulate neurosecretory material within the nerve endings and become females. When formerly isolated females are put together, their stores of neurosecretory material are rapidly discharged. Subsequently they lay egg masses and switch to the male state. These effects are mediated by a pheromone released during social contact of formerly isolated females. The complexity of the relationship between neurosecretory activity and sexual state is indicated by the situation in animals maintained in pairs, when both male and female partners have swollen nerve endings packed with secretory material.     

The effect of castration upon the ultrastructure of the rat hypothalamus

Summary The neurosecretory hypothalamic nuclei and the inner zone of the median eminence of castrated rats were studied under the electron microscope. After one month of castration all the neurosecretory neurons of both nuclei show signs of hyperactivity characterized by dilated cisternae of the endoplasmic reticulum containing a macromolecular filamentous material and an increase in the number of ribosomes. After six months of castration, some neurosecretory neurons show an increased number of neurotubules and larger lysosomes than in the controls. Other neurons show a very significant hypertrophy of the endoplasmic reticulum, with large amounts of intracisternal filamentous material. These cells have few neurosecretory granules and in the adjacent synapses the number of granulated vesicles is increased. In the supraoptic nucleus there are two kinds of neurosecretory axons: the clear ones, which are similar to those that appear in control animals and the dark ones, which have smaller elementary granules. In the inner zone of the median eminence the axons show an increase in the number of neurosecretory granules with respect to the controls. After supplementary administration of sexual hormones, all the modifications produced by castration disappear. The ultrastructural changes observed in the neurosecretory nuclei after castration are discussed in relation to those previously described in the neurohypophysis under the same experimental conditions. A feedback regulatory action of sex hormones on hypothalamic neurosecretory neurons is postulated.Supported by grants from the Consejo Nacional de Investigaciones Científicas y Técnicas and by the Air Force Office of Scientific Research (AF-AFOSR 963-67).We are deeply indebted to Mrs. Defilippi-Novoa and Mr. Alberto Saenz for their skillful assistence.     

The electrical activity of mechanoreceptive and neurosecretory neurons in the stick insectCarausim morosus

Summary Action potentials have been recorded from the neurosecretory cells which lie on the link nerve inCarausius morosus. The neurosecretory cells are spontaneously active in completely isolated preparations, firing with a regular but low frequency (<1 imp/s) or in small bursts (12 imp/s). The action potentials recorded extracellularly from the neurosecretory fibres are characteristically of long duration (2 to 10 ms), whereas those of motor or sensory fibres are of shorter duration (0.6 to 0.8 ms). The neurosecretory action potentials are also characterised by their slow conduction velocity (0.15 to 0.25 m/s) compared to those from motor and sensory fibres (0.54 to 0.7 m/s). The action potentials are propagated from the region of the cell body towards the terminals and have been recorded passing along all the major nerves in the periphery.Recordings from three of the non-neurosecretory cells which lie on peripheral nerves show that they respond to stretching of the nerves upon which they lie or of nerves which branch in the immediate vicinity. The action potentials are propagated away from the cell body towards the central nervous system. The neurons are termed peripheral nerve stretch receptors.We are grateful to the Science Research Council for financial support.     

Histological studies of the neurosecretory and retrocerebral complex of the water beetle,Hydrophilus olivaceus Fabr. (Insecta,Coleoptera)

The neurosecretory cells (NSC) in the breeding phases show gigantic sized droplets. NSC show two peaks of activity in females at midnight and at 9 A.M., while a single peak occurs in males between midnight and 3 A.M. Light has a triggering effect on the neurosecretory release. The corpus cardiacum is the main neurohaemal organ. It has a nerve core formed by the nervi corpus cardiacum I and II, which is surrounded by the glandular region. Intrinsic secretory chromophilic cells and chromophobic cells occur randomly. Aorta is probably not a storage-release centre. The corpora allata are lobulated structures, showing cyclical activity which is correlated with egg-maturation. No neurosecretory material occurs in allatum.     

Etude histologique,histochimique et ultrastructurale de la pars intercerebralis chez Locusta migratoria L. (Orthoptere)

Summary A histological, histochemical and ultrastrucutral study of the pars intercerebralis (PI) has been made in Locusta migratoria. The acellular neural lamella is made up of an elastic tissue and collagen fibrils. The cells of the perilemma contain numerous lysosome structures and lipid granules.Three different types of neurosecretory cells (NSC A, B and C) have been distinguished in the PI associated with giant neurons.The cells termed A and B seem not to have an activity cycle during the two last larval instars. At the moment of sexual maturity the NSC A show an important accumulation of neurosecretory material and their number increases at the expense of the NSC B. The NSC A, which are characterized by a highly developped endoplasmic reticulum, contain numerous secretory granules which appear to be individualized in the Golgi complex in three different ways. The NSC B, with a reduced endoplasmic reticulum and an almost quiescent Golgi complex, contain abundant lysosome structures and more seldom some neurosecretory granules. In fact, the study of the fine structure shows different intermediate types, linking in a continuous way typical A cells and typical B cells. NSC A and NSC B might correspond to two opposed stages of secretory activity of one single cell type: the A cell representing the activity stage and the B cell the quiescent stage.NSC C show an accumulation of their neurosecretory products in relation to metamorphosis and sexual maturity. Ultrastructural evidence confirms their neurosecretory activity.A mode of regulating neurosecretion in NSC A and B by internal catabolism of the secretion and formation of lysosome like structures is discussed in the present paper.The giant neurons, which are surrounded by a glial envelope (trophospongium), contain several dense granules originated from Golgi complex.     

Requirements for the identification of dense-core granules

Dense-core granules (DCGs), cytoplasmic organelles competent for regulated exocytosis, show considerable heterogeneity depending upon the specificity of their expressing cells--primarily neurons and neurosecretory cells. DCGs have been mainly identified by detecting their cargo molecules, often members of the granin family, and using conventional electron microscopy and immunocytochemistry. However, by a critical analysis of the various stages of DCG "life" within neurosecretory cells, we have highlighted several specific molecular and functional properties that are common to all these organelles. We propose that these properties be considered as strict requirements for the identification of DCGs.     

Aminergic neuron systems of lobsters: morphology and electrophysiology of octopamine-containing neurosecretory cells

In the American lobster (Homarus americanus) the biogenic amines serotonin and octopamine appear to play important and opposite roles in the regulation of aggressive behavior, in the establishment and/or maintenance of dominant and subordinate behavioral states and in the modulation of the associated postural stances and escape responses. The octopamine-containing neurosecretory neurons in the thoracic regions of the lobster ventral nerve cord fall into two morphological subgroups, the root octopamine cells, a classical neurohemal group with release regions along second thoracic roots, and the claw octopamine cells, a group that selectively innervates the claws. Cells of both subgroups have additional sets of endings within neuropil regions of ganglia of the ventral nerve cord. Octopamine neurosecretory neurons generally are silent, but when spontaneously active or when activated, they show large overshooting action potentials with prominent after-hyperpolarizations. Autoinhibition after high-frequency firing, which is also seen in other crustacean neurosecretory cells, is readily apparent in these cells. The cells show no spontaneous synaptic activity, but appear to be excited by a unitary source. Stimulation of lateral or medial giant axons, which excite serotonergic cells yielded no response in octopaminergic neurosecretory cells and no evidence for direct interactions between pairs of octopamine neurons, or between the octopaminergic and the serotonergic sets of neurosecretory neurons was found.     

Neurosecretory system of the ventral ganglia in the dragonfly,Orthetrum chrysis (selys) (Odonata: Libellulidae)

Summary The neurosecretory cells of the ventral ganglia in the adult dragonfly, Orthetrum chrysis, are classified into A, B, C1 and C2 cells. The neurosecretory material in the ventral ganglia is composed of PAS-positive material with 1-, 2-glycol groups and some proteins. The proteins rich in cystine or cysteine occur predominantly in the A cells, moderately in C cells and negligibly in B cells. Proteins containing arginine occur in A and B cells only, and those containing basic amino acids occur in C2 cells. The neurosecretory pathways and the neurohemal organs are also described.