The hypothalamo-hypophysial system in acipenseridae

Summary The distribution of adrenergic and peptidergic (Gomori-positive) structures of the hypophysial neuro-intermediate complex in Acipenseridae has been studied by means of light, fluorescence, and electron microscopy. Adrenergic fibres (B-fibres) and their terminals have been detected in the neurohypophysis of these fishes. The terminal swellings of B-fibres as well as the terminals of the neurosecretory peptidergic fibres (A₁ and A₂) make contact with the basement membrane of the connective tissue layer separating the neurohypophysis from the intermediate lobe. Capillaries are situated within this layer and, therefore, the main part of the fibre terminals is in contact with the pericapillary space. The release of catecholamines from the adrenergic terminals into the capillaries connected with the general circulation is supposed. The diffusion of catecholamines through the connective tissue layer into the parenchyma of the intermediate lobe is also suggested. Hence, the glandular activity of the intermediate lobe seems to be under the dual control of adrenergic and peptidergic elements of the hypothalamus.The authors wish to express their deep appreciation to G. M. Persov, Dr. Sc. Biol., Head of the Laboratory of Experimental Ichthyology, Petershof Biological Institute of the University of Leningrad, for the material supplied for fluorescence microscopy, and to Mr. G. V. Sabinin for photographic services.     

The hypothalamo-hypophysial system in acipenseridae

Summary The neurohypophysis of sexually mature male and female Acipenser güldenstädti Brandt and Acipenser stellatus Pallas was studied light and electron microscopically. The recessus hypophysei lined with ependymal cells of two main types, narrow and wide, are in the center of the neurohypophysial roots. Processes from both cell types run radially to the basement membrane of the connective tissue layers abutting on the hypophysial intermediate lobe. Protrusions penetrating deep into the recessus hypophyseus are found in the apical parts of the wide cells. Pituicytes are rare in the neurohypophysis. The ultrastructure of both ependymal cell types and of the pituicytes is described. Nonmyelinated Gomori-positive (peptidergic) neurosecretory A₁ and A₂ type fibres and their terminals containing elementary neurosecretory granules (1400–1800 Å and 1000–1400 Å respectively) are the main structural elements of the neurohypophysis. Some dark and single myelinated neurosecretory fibres have been found. The adrenergic fibres (type B) were described earlier (Polenov et al., 1972a). The structural peculiarities of the neurohypophysis are discussed in functional and comparative-morphological terms.     

Ecologic-histophysiological analysis of the neurohypophysis-metaadenohypophysis complex of the sazan female before and after spawning]

Much neurosecretory material (4.4 +/- 0.1 units) is seen in the neurohypophysis of sazan females before spawning. A1, A2, and B type terminals contain numerous neurosecretory granules, in particular, elementary granules. Synaptic vesicles are not numerous. The amount of neurosecretory material decreases during and shortly after spawning down to 3.6 +/- 0.3 units. Neurosecretory terminals have less elementary granules. The number of granulated, disintegrating, residual granules and synaptic vesicles somewhat increases. It is hypothesized that mainly peptide neurohormones are discharged from A1 and A2 terminals in the common circulation, and that both peptide neurohormones and monoamines reach glandular cells of the metaadenohypophysis in sazan females during and immediately spawning. The role played by neurohormones and monoamines in controlling the function of both the metaadenohypophysis glandular cells and visceral organs involved in the general adaptive reactions of the fish organism is discussed.     

The hypothalamo-hypophysial system of the wild carp,Cyprinus carpio L.

Summary The posterior neurohypophysis (PNH)-pars intermedia complex of the wild and pond carp, Cyprinus carpio L., has been studied by light, fluorescence and electron microscopy. Gomori-positive neurosecretory fibres are abundant in the main trunk of the neurohypophysis as well as its roots penetrating the pars intermedia. Terminals of these fibres are in contact with capillaries of the general circulation and with glandular cells of the pars intermedia. Monoaminergic fibres with a weak green fluorescence, somewhat increasing after injection of nialamide into the pond carp, have largely the same distribution. Three types of neurosecretory fibres and their terminals have been recognized in the PNH-pars intermedia complex. Types-A₁ and -A₂ fibres, containing granules of 140–180 nm and 100–160 nm in diameter respectively, are peptidergic Gomori-positive. Type-A₂ fibres predominate in the PNH. The least frequent monoaminergic type-B fibres have granules of 60–100 nm in diameter. Numerous peptidergic and few monoaminergic neurosecretory terminals make contact with the capillaries located within the roots of the PNH as well as at the border between them and the pars intermedia. Both peptidergic and monoaminergic terminals make direct synaptoid contacts with the gland cells or end close to connective tissue septa, basal lamina or pituicytes. The PAS-positive gland cells and to a lesser degree the leadhaematoxylin-positive gland cells show these relationships with neurosecretory terminals. The question concerning the mode of interaction between peptidergic and monoaminergic structures in the dual control of the gland cells of the pars intermedia of teleosts is discussed.     

The hypothalamo-hypophysial system of the frog Rana temporaria

Summary The median eminence (ME) of the adult frog, Rana temporaria, was studied by means of electron microscopy including quantitative electron-microscopic autoradiography. In frogs captured in May and June numerous peptidergic neurosecretory fibres extending via the internal zone to the pars nervosa display large swellings containing few granules, mitochondria, neurotubules and cisternae of the smooth endoplasmic reticulum. In addition, few secretory globules up to 1.5 m in diameter occur in these varicosities. In animals collected during the autumn period many of these neurosecretory swellings filled with neurosecretory granules and polymorphic inclusions resemble Herring bodies. Three types of granule-containing neurosecretory fibres were observed in the external zone (EZ) of the ME of adult R. temporaria. Peptidergic A₁- and A₂-type fibres are characterized by granules 150–220 nm and 100–160 nm in diameter, respectively. Monoaminergic fibres of type B with granules approximately 100 nm in diameter represent 50% of all neurosecretory elements in the EZ of the frog ME; 12% of the total number of granule-bearing axons in the EZ actively taking up radiolabelled 5-hydroxytryptophan are thought to be serotoninergic terminals. Neurosecretory terminals of all types and glial vascular endfeet establish direct contacts with the perivascular space of the primary portal capillaries. Some neurosecretory terminals are separated from the lumen of the third ventricle by a thin cytoplasmic lamella of tanycytes. The possible physiological significance of this structural pattern is discussed.     

Ultrastructure of the peptidergic and monoaminergic neurons in the hypothalamic neurosecretory system of Anuran Batracians

Summary In the toad Bufo arenarum Hensel the following regions of the hypothalamic — neurohypophyseal system were studied under the electronmicroscope: preoptic and paraventricular nuclei, median eminence and infundibular process of the neurohypophysis.Neuronal perikarya of the preoptic nucleus are loaded with typical neurosecretory granules of peptidergic nature having a mean diameter of 1660 Å. While most neurons of the winter toad are in a storage stage a few show signs of a more active synthetic activity. A distinctive feature of preoptic neurons is the presence of large lipid droplets. The paraventricular nucleus contains small neurons containing granulated vesicles with a mean diameter of 800-1000 Å. In the region extending between these two nuclei and the median eminence axons containing either neurosecretory elementary granules or granulated vesicles are observed.The inner zone of the median eminence is occupied by axons of the preoptic neurohypophyseal tract; two types of axons, according to the size and density of the neurosecretory granules, may be recognized. The outer zone of the median eminence contains mainly axons and nerve terminals containing granulated vesicles of probable monoaminergic nature and only a few with granules of peptidergic type.The neurohypophysis contains two kinds of axons: one with more dense granules of 1800 Å and the other with granules of lesser electron density and 2100 Å. At the ending proper small clear vesicles of synaptic type are found.A progressive increase in volume of the peptidergic granules along the axon is demonstrated. This is of the order of 218% from the preoptic perikarya down to the infundibular process. The physiological significance of the two neurosecretory systems — i.e. the monoaminergic and the peptidergic — and the probable nature of the two types of peptidergic axons is discussed.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).The authors want to express their gratitude to Mrs. Defilippi-Novoa and Mr. Alberto Sáenz for their skillful assistance.     

Electron microscopical study on the development of the nerve supply of the pituitary pars intermedia of the mouse

Summary The development of the nerve supply of the pituitary pars intermedia (PI) of C3H mice was studied by electron microscopy. Nerve fibres and terminal structures, most probably adrenergic, first appear in the newborn. The adult innervation pattern is achieved by the end of the first postnatal week.In the adult animal two types of nerve terminals were distinguished; type A (peptidergic or neurosecretory) and type B (adrenergic). The peptidergic fibres were scarce and exhibited no synapse-like contacts. It is suggested that they are of secondary importance in a direct nervous hypothalamic control of PI function. Type B terminals were found throughout the PI. They formed synapse-like contacts with the glandular cells, indicating that the primary innervation is exerted by adrenergic neurons.An autonomous differentiation of the glandular cells and in the adult a combined direct nervous and neurohumoral control of PI function is suggested.This investigation was supported by grant No B 2180-026 from the Swedish Natural Science Research Council. The skilful technical assistance of Mrs Ulla Wennerberg is gratefully acknowledged     

Ultrastructural localisation of ATP-gated P2X2 receptor immunoreactivity in the rat hypothalamo-neurohypophysial system

The distribution of the P2X₂ subtype of the purine receptor associated with the extracellular signalling activities of ATP was studied in the rat hypothalamo-neurohypophysial system at the electron microscope level. Receptors were labelled with ExtrAvidin-horseradish peroxidase preembedding immunocytochemistry using a polyclonal antibody against a fragment of an intracellular domain of the receptor. Immunoreactivity to P2X₂ receptors was localised in: (i) paraventricular and supraoptic nuclei—in subpopulations of endocrine neurones, neurosecretory and non-neurosecretory axons and dendrites; and (ii) the neurohypophysis—in pituicytes and subpopulation of neurosecretory axons. In both the hypothalamic nuclei examined, labelled asymmetric axo-dendritic synapses were commonly observed. These synapses involved either P2X₂-labelled axon terminals (synaptic buttons) and unlabelled dendrites or labelled dendrites and unlabelled axon terminals. Axo-somatic synapses established by P2X₂-positive axons on P2X₂-positive endocrine cell bodies as well as on P2X₂-negative somata were also observed. The functional significance of these findings is discussed.     

Fine structure of pars neuro-intermedia of the loach,Misgurnus fossilis L.

Summary Two types of glandular cells are present in the pars intermedia of the loach, Misgurnus fossilis. Basophils are characterized by the presence in their cytoplasm of numerous secretory granules containing electron-dense and homogeneous material and by scarce endoplasmic reticulum. Weak acidophils contain in their cytoplasm abundant endoplasmic reticulum and numerous granules of different electron densities.The distal part of the neurohypophysis is composed of several types of neurosecretory axons, strongly branched pituicytes, numerous capillaries, and connective tissue elements. The axon terminals form the neuroglandular, neurovascular and neurointerstitial contacts. Some axon terminals are closely apposed to the basement membrane separating neurohypophysis from meta-adenohypophysis. At points of absence of continuity of this membrane, some neurosecretory axons become directly contiguous with cytoplasmic membranes of the intermedia cells.The investigation was partly supported by a research grant from the Zoological Committee of the Polish Academy of Sciences.     

The Ca2+ channel β2 subunit is selectively targeted to the axon terminals of supraoptic neurons

The assembly of high voltage-activated Ca²⁺ channels with different β subunits influences channel properties and possibly subcellular targeting. We studied β subunit expression in the somata and axon terminals of the magnocellular neurosecretory cells, which are located in the supraoptic nucleus (SON) and neurohypophysis, respectively. Antibodies directed against the 4 Ca_Vβ subunits (Ca_Vβ₁-Ca_Vβ₄) were used for immunoblots and for immunostaining of slices of these two tissues. We found that all 4 β subunits are expressed in both locations, but that Ca_Vβ₂ had the highest relative expression in the neurohypophysis. These data suggest that the Ca_Vβ₂ subunit is selectively targeted to axon terminals and may play a role in targeting and/or regulating the properties of Ca²⁺ channels.     

Histochemical studies on the neurosecretory system of the common garden lizard Calotes versicolor (Daudin) under experimental condition

Summary Histochemical localizations of various substances in the hypothalamic neuro-secretory cells as well as neurohypophysis of the normal and stressed (scalded) garden lizards, Calotes versicolor suggest that the neurosecretory materials, secreted by the hypothalamic neurones may be a mucoprotein-lipoprotein complex which is finally stored in the neuro-hypophysis. Only the proteinaceous fraction of this complex appears to be released by the nerve terminals in the neurohypophysis under the stressed condition. Such suggestion implies a chemical transformation of neurosecretory substances after leaving the neurosecretory cell body.     

The hypothalamo-hypophysial system of the lamprey,Lampetra fluviatilis L.

The distribution of monoaminergic structures was studied in the proximal neurosecretory contact region and neurohypophysis of the lamprey by light and electron microscopic radioautography. Only weak radioautographic reactions were found in the proximal neurosecretory contact region 1 h after injection of 3H-dopamine. High-resolution radioautography revealed some labeled neurosecretory terminals mainly in contact with the basement membrane of the connective tissue layer separating the proximal neurosecretory contact region from the hypophysial pars distalis. The number of silver grains as well as the number of neurosecretory terminals marked by the presence of labeled dopamine was much higher in the neurohypophysis of the same species. In the latter, labeled neurosecretory terminals were found in contact with the connective tissue layer containing blood vessels of the general circulation. Some neurosecretory terminals make synaptoid contacts with tanycyte perikarya and their basal processes. According to their ultrastructure and the size of their granules, the labeled neurosecretory terminals are identical with the B type terminals described in both neurohemal regions (transmission electron microscopy). No labeled neurosecretory terminals were observed in the proximal neurosecretory contact region and the neurohypophysis of lampreys treated with the serotonin precursor, 3H-5-hydroxytryptophan.     

The hypothalamo-hypophysial system in acipenseridae

Summary The peptidergic neurosecretory cells (NSC) of the nucleus praeopticus (NP) of male and female sturgeons, A. güldenstädti Brandt, were studied light microscopically at different stages of their life cycle and under experimental conditions. Four main NSC types reflecting different phases of secretory cycle and life course of the cell have been tentatively distinguished.The maximum percentage of the high and moderate active NSC are found in juvenile animals in the sea (stages I and I–II of gonadal maturity, sgm), in upstream migrating fish in spring before spawning, in down-stream migrating fish 1–1.5 months after spawning, and in experimental fish kept for 8.5 hours in a 32 sodium chloride solution. The least active NSC accumulating neurosecretory material (NSM) are characteristic of juvenile fish (sgm II) in the sea, sturgeons maintained in a sodium chloride solution for 3.5 or 6 hours and fish which remained in a net, thrown to the sea, for some hours before fixation. The lowest percentage of these cells is observed in autumn migrants, in females soon after spawning (sgm VI) and in fish kept for 8.5 hours in a sodium chloride solution. Cells rich in basophilic substance and poor in NSM occur in juvenile and in down-stream migrating fish. Cells reflecting the state of exhaustion after hyperactivity and ageing cells are seen in adults, especially in sgm VI fish, and in autumn migrants. Pyknomorphous NSC are constantly present in all fish; they are most numerous in sturgeons found in a net. A diagram demonstrating the life course and the secretory cycle of the NSC is presented and the role of the hypothalamo-hypophysial neurosecretory system (HHNS) under stress conditions is discussed.The authors are grateful to Dr. V.Z. Trusov, M.Sc. (Biol.), for supplying a portion of the sea sturgeons and to P.E. Garlov, M.Sc. (Biol.), for his valuable help during the experiments     

Calcium channels that are required for secretion from intact nerve terminals of vertebrates are sensitive to omega-conotoxin and relatively insensitive to dihydropyridines. Optical studies with and without voltage-sensitive dyes

Extrinsic absorption changes exhibited by potentiometric dyes have established the ionic basis of the action potential in synchronously activated populations of nerve terminals in the intact neurohypophyses of amphibia and mammals (Salzberg et al., 1983; Obaid et al., 1983, 1985b). Also, large and rapid changes in light scattering, measured as transparency, have been shown to follow membrane depolarization and to be intimately associated with the release of neuropeptides from the nerve terminals of the mouse neurohypophysis (Salzberg et al., 1985; Gainer et al., 1986). We report some experiments that help to define the pharmacological profile of the calcium channels present in intact neurosecretory terminals of vertebrates. For these, we used the peptide toxin omega-conotoxin GVIA (1-5 microM) and the dihydropyridine compounds Bay-K 8644 and nifedipine (2-5 microM), together with the after-hyperpolarization of the nerve terminal action potential. This undershoot depends upon the activation of a calcium-mediated potassium channel, as suggested by its sensitivity to [Ca++]o and charybdotoxin. omega-conotoxin GVIA substantially reduced the after-hyperpolarization in neurosecretory terminals of Xenopus, while neither of the dihydropyridine compounds had any effect under conditions that mimic natural stimulation. The effects of these calcium channel modifiers on the action potential recorded optically from the terminals of the Xenopus neurohypophysis were faithfully reflected in the behavior of the light-scattering changes observed in the neurohypophysis of the CD-1 mouse. omega-conotoxin GVIA (5 microM) reduced the size of the intrinsic optical signal associated with secretion by 50%, while the dihydropyridines had little effect. These observations suggest that the type of calcium channel that dominates the secretory behavior of intact vertebrate nerve terminals is at least partially blocked by omega-conotoxin GVIA and is insensitive, under normal conditions, to dihydropyridines.     

The hypothalamo-hypophysial system in Acipenseridae

Summary The proximal neurosecretory contact region (PCR) of Acipenseridae, a homologue of the tetrapod median eminence, has been studied by light, fluorescence and electron microscopy. It occupies the rostral and chiefly the ventral surfaces of the hypothalamic tuber cinereum. PAF-positive fibres occur in the zone of the preoptico-hypophysial tract but their terminal enlargements are concentrated mainly in the external zone. They make contact with the primary portal capillaries situated in the pia mater. Monoaminergic fibres and terminals with an intense green fluorescence are localized in the same regions. The fibres of some bipolar monoaminergic neurons of the PCR make contact both with the third ventricle and the primary portal capillaries.Three types of granule-containing neurosecretory fibres and terminals have been recognized in the PCR. Fibres of types A₁ (d=120–300 nm) and A₂ (d=100–170 nm) are peptidergic PAF-positive, although some fibres, including some of type A₁; belong possibly to PAF-negative type. Monoaminergic type B fibres have granules 80–100 nm in diameter. Neurosecretory terminals and vascular endfeet of tanycytes make contact with the 70 nm thick outer basement membrane of the primary portal capillaries. Several laminae of thin horizontally oriented tanycyte processes form a boundary between the external zone and the preoptico-hypophysial tract. Few neuroglial cells with pale cytoplasm, numerous lysosomes and lipofuscin granules are seen in this region. It is hypothesized that, as in other vertebrates, both peptide hypophysiotropic neurohormones and monoamines are discharged from the PCR into the portal circulation and affect the activity of the glandular cells of the pars distalis.Dedicated to Prof. Dr. Drs. h.c. W. Bargmann on the occasion of his 70th birthday     

Immunogold labelling of serotonin-like and FMRFamide-like immunoreactive material in neurohaemal areas on abdominal nerves of Rhodnius prolixus

The ultrastructure of neurohaemal areas on abdominal nerves of the blood-sucking bug Rhodnius prolixus was investigated. Four types of axon terminals were found, distinguished by the morphology of their neurosecretory granules. By use of post-embedding immunogold labelling, granules in Type I axon terminals were shown to contain serotonin-like immunoreactive material, and granules in Type II axon terminals were shown to contain FMRFamide-like immunoreactive material. There was no colocalization of these materials. It is suggested that Type III terminals contain peptidergic diuretic hormone, which has previously been reported to be present in electron-dense neurosecretory granules in this neurohaemal area. The identity of material in Type IV terminals is unknown.     

The hypothalamo-hypophysial system of hypophysectomized rats

Summary The median eminence (ME) of hypophysectomized rats was studied by means of light and electron microscopy. Paraldehyde-fuchsin (PAF)-positive material is seen in the external zone (EZ) of the ME 2–5 days after the operation. Its amount gradually increases especially in the caudal part of the ME during the following few days. Some PAF-positive fibers make contact with the subependymally located blood capillaries. In the most caudal region of the recessus infundibuli they penetrate into the third ventricle. PAF-positive material decreases markedly from the ME of rats two months after hypophysectomy and exposure to a 1% salt load. Fibers of types A₁, A₂ and B containing granules of 120–220 nm, 100–150 nm and 80–100 nm in diameter, respectively, are seen in the EZ of the ME in hypophysectomized rats, although almost exclusively A₂- and B-type structures make contact with the primary portal capillaries in intact animals. All types of neurosecretory fibers establish contact with the subependymal nonfenestrated blood capillaries and penetrate the recessus infundibuli. Some neurosecretory terminals of different types make direct contact with the glandular cells of the pars tuberalis or are separated from them by a thin basal lamina. It is assumed that mainly neurosecretory fibers of types A₂ and B are permanently connected with the primary portal capillaries in the EZ of the ME in intact mammals, while the overwhelming majority of fibers of A₁-type shows ingrowth during the course of postoperative reparation. The possible physiological significance of the described changes is discussed.     

Morphological aspects of the hypothalamic-hypophyseal system

Summary The neural connections of various parts of the neurohypophysis of mammals (cats, dogs, rats) were investigated with the aid of the Golgi method, the aldehyde fuchsin technique (in combination with perfusion of the vascular system with India ink gelatine) and electron microscopy. The abundant supply of the palisade zone of the median eminence with thin axon terminals of the tubero-infundibular system is described. The independence of these terminals from the axons of the magnocellular neurosecretory system of the hypothalamus, which terminate in the caudal parts of the neurohypophysis, is evident. The clearcut differences between morphology and localization of the terminals of the parvi- and magnocellular neurosecretory systems at various levels of the neurohypophysis are described. The morphological aspects of the release of neural mediators in regions of these terminals are discussed.     

FINE STRUCTURE OF THE NEUROHYPOPHYSIS OF THE OPOSSUM (DIDELPHIS VIRGINIANA)

The neurohypophysis of the opossum (Didelphis virginiana) was studied by electron microscopy in order to amplify Bodian''s classic light microscopic observations in which he demonstrated a definite lobular pattern. The lobule of the opossum neurohypophysis is divided into three regions: a hilar, a palisade, and a septal zone. The hilar portion contains bundles of nerve fibers, the extensions of the hypothalamo-hypophyseal tract containing neurofilaments but few neurosecretory granules. In the opossum, pituicytes have a densely fibrillar cytoplasm. Herring bodies are prominent in the hilar region. They are large bodies packed with neurosecretory granules that have been described as end bulb formations of axons. From the hilar region, axons fan out into a palisade zone where the nerve terminals packed with neurosecretory granules, mitochondria, and microvesicles abut upon basement membranes. The neurosecretory granules are similar to those present in the neurohypophysis of other mammals, except for an occasional huge granule of distinctive type. Material morphologically and histochemically resembling glycogen occurs as scattered particles and as aggregates within nerve fibers. The septal zone, containing collagen, fibroblasts, and numerous small capillaries, is separated from the adjacent glandular tissue by a basement membrane.     

Participation of vertebrate forebrain activating systems in organization of the wakefulness-sleep cycle

The article considers mechanisms of diencephalic-telencephalic interactions in regulation of the wakefulness-sleep cycle in various classes of vertebrates. In such interactions a special role is played by the dopaminergic systems that perform neurosecretory function at the level of diencephalon and neurotransmitter function at the level of telencephalon. Concepts of A.I. Karamyan and A.L. Polenov about the stage pattern of development of CNS and neurosecretory systems are presented, as well as the interconnection of dopaminergic and glutamatergic neurotransmitter systems in the mammalian neostriatum in the wakefulness-sleep cycle is considered. Comparison of dynamics of expression of the dopamine metabotropic receptors and of the glutamate ionotropic receptors in neostriatum showed unidirectional changes of D₁ and AMPA on the background of the 6-h sleep deprivation as well as of D₂ and NMDA on the background of postdeprivative sleep. The corticofugal direction of glutamate impulsation and its relatively fast actions allow admitting its triggering action on generation of the sleep-inducing processes in the underlying brain parts.