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 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.     

The hypothalamo-hypophysial system in acipenseridae

Summary The percentage of peptidergic (A₁ and A₂) and adrenergic (B) neurosecretory terminals was studied in the neurohypophysis of sexually mature female sturgeons. Neurosecretory terminals of the A₂ type prevail in the neurohypophysis, whereas A₁ and B terminals are rare. The activity of these types of terminals was established (1) during upstream migration, (2) shortly after spawning, and (3) three to six weeks after spawning. Terminals of B type are the most active elements during all the periods studied. These elements become strongly activated in sturgeons during upstream migration, i.e., earlier than the peptidergic neurosecretory terminals. Peptidergic terminals, especially elements of type A₂, become synchronously and strongly activated in fish shortly after spawning. In the late postspawning period neurosecretory terminals of all three types become synchronously inactive, persisting in a quiescent state in comparison to the two previous periods. The appearance of neurosecretory material discharged into the intercellular clefts by exocytosis correlates on the whole with the activity level of the A₁ and A₂ terminals in each individual studied. A functional correlation exists between the activity of the peptidergic and adrenergic neurosecretory terminals in the neurohypophysis. The data obtained are discussed with reference to a concept regarding spawning in some fish species as a physiological stress (Polenov et al., 1976). A possible dual control (peptide and monoamine neurohormones) over the function of visceral organs and glandular cells of the intermediate lobe of the hypophysis is also suggested (Polenov, 1970, 1975, 1978; Polenov and Belenky, 1973).Dedicated to the memory of Professor Wolfgang Bargmann, a great scientist and a generous friend     

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.     

Development of innervation to the atrial myocardium of the rabbit

Summary The development of innervation to the atrial myocardium of rabbits from 20th day of gestation to 35 days postnatal was studied ultrastructurally by electron microscopy and by demonstration of catecholamines by histofluorescence. Special attention was directed to the first morphologic appearance of nerve fibers and terminals and the closeness of juxtaposition of terminals with myocardial cells. Adrenergic and cholinergic terminals were identified on the basis of their differential ability to take-up and store the false adrenergic neurotransmitter 5-hydroxydopamine. Adrenergic terminals were first encountered at 20 days of gestation whereas cholinergic terminals could not be positively identified until the 24th day of gestation. Throughout development adrenergic terminals were more numerous than cholinergic, about 71 % of the terminals encountered being adrenergic. Many terminals approach closely (20–30 nm) to the sarcolemma of the muscle cells of the atrium. In many instances adrenergic and cholinergic fibers travel together in the same nerve bundle and are closely apposed without intervening Schwann-cell cytoplasm. Such a relationship could allow peripheral interaction between these fibers in the myocardium.Supported in part by the Kentucky Heart Association, Human Development Studies Program of the University of Kentucky and DHEW Grant 1 RO1 HL 22226-01 HED from the National Heart, Lung and Blood Institute. The technical assistance of Merle Wekstein is appreciated     

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     

Distribution of fibres reacting with an α-endorphin antiserum in the neurohypophysis of Carassius auratus and Cyprinus carpio

Summary A strong positive immunoreaction with an -endorphin antiserum occurs in two distinct sites of the goldfish and carp neurohypophysis. Fluorescent nerve terminals are found in the laminar nerve processes located in the rostral pars distalis, but the immunocytological reaction is mainly localised on the nerve processes of the posterior neurohypophysis lying between the intermediate lobe cells. Almost all the digitations of the neurohypophysis are strongly fluorescent. The immunoreactive fibres probably originate from the hypothalamus, where perikarya displaying the same immunoreaction have been found in the pars lateralis of the nucleus lateralis tuberis and in some minor centres. The possibility that the immunoreactive substances revealed on the neurohypophyseal processes may originate in the intermediate lobe cells is also discussed. It has now to be established if this hypothalamo-hypophyseal system contains a substance with endorphic properties or only some immunologically related substance devoid of the corresponding physiological activities.     

Ocular adrenergic neurons of the flying fox,Pteropus giganteus brünn. (Megachiroptera)

Summary The distribution of adrenergic nerves in the eyes of the large fruit-bat (flying fox) Pteropus giganteus Brünn. was investigated with the histofluorographic method of Falck and Hillarp. The general pattern conforms to that seen in most mammalian eyes. The most notable observation was in the chamber angle, where the meshwork covering the outflow channels receives numerous adrenergic terminals, making possible a direct adrenergic nervous influence on the outflow of aqueous humour. As in other mammals, adrenergic terminals are noted in the cornea, in both iris muscles, and in association with the ciliary epithelium. In the normal retina, adrenergic neurons occur mainly at the border between the inner nuclear and inner plexiform layers. After the injection of -methylnoradrenaline, additional neurons become fluorescent. We do not know the true transmitter of these additional neurons.This work was supported by grants from the Swedish Medical Research Council (projects O4X-2321 and O4X712) and the Faculty of Medicine, University of Lund.     

Vasoactive intestinal polypeptide (VIP)- and neuropeptide Y (NPY)-containing nerve fibres in the penile cavernous tissue of green monkeys (Cercopithecus aethiops)

Summary The cavernous body of green monkeys contains many unmyelinated and few myelinated axons. The unmyelinated axons form terminals in the adventitia of the arteries, between trabecular muscle cells, in the interstitium, and close to endothelium cells of the sinuses. All terminals displayed predominantly small clear vesicles and very few large granular vesicles; small granular vesicles were not seen. However, in rabbit penises, terminals with many large granular vesicles are prominent. Immunohistochemistry (PAP technique) showed a dense network of VIP- and NPY-reactive fibres around the arteries and around trabecular muscles. The density of nerve fibres was particularly high around the subendothelial cushions of the helicine arteries. Double staining for NPY and VIP revealed that both peptides were colocalized. Immunocytochemistry (preembedding PAP technique) showed VIP- and NPY-reactivity in terminals with small clear vesicles; the reaction product was bound to the cytoplasmic face of different membrane types. Although the intracellular localization of the reaction product is probably due to artefactual displacement during preparation, the uniformity of the terminals questions the view that large and small granular vesicles in all species characterize peptidergic and noradrenergic terminals, respectively. The essential findings can be summarized as (1) a high degree of uniformity of nerve terminals, (2) colocalization of VIP and NPY, (3) heavy innervation of the subendothelial cushions of the helicine arteries, and (4) possible innervation of endothelial cells.     

Ultrastructure of dynamic and static skeletofusimotor endings in a cat muscle spindle

Summary Endings of four skeletofusimotor axons in a spindle of the cat tenuissimus muscle were examined in semithin (1-m thick) and ultrathin transverse serial sections. Two (dynamic) axons terminated on the nuclear bag₁ intrafusal muscle fiber and on extrafusal fibers of the dark type. Two (static) axons terminated on the nuclear chain intrafusal fibers and extrafusal fibers of the intermediate type. The degree of indentation of axon terminals into the muscle surface, thickness of the sole plate and extent of folding of subjunctional membranes differed among intrafusal and extrafusal terminations of the same axon. Endings of axons on the bag₁ and chain fibers were also morphologically dissimilar. Motor axons may not determine ending morphology. Rather the form and structure of a bag₁ or chain ending may be determined by the type of intrafusal fiber on which the ending lies and the ending's distance from the primary sensory axon.     

Binding of Amyloid Beta-Protein to Intracellular Brain Proteins in Rat and Human

Amyloid beta-protein (A), in its soluble form, is known to bind several circulatory proteins such as apolipoprotein (apo) E, apo J and transthyretin. However, the binding of A to intracellular proteins has not been studied. We have developed an overlay assay to study A binding to intracellular brain proteins. The supernatants from both rat and human brains were found to contain several proteins that bind to A 1–40 and A 1–42. No major difference was observed in the A binding-proteins from brain supernatants of patients with Alzheimer's disease and normal age-matched controls. Binding studies using shorter amyloid beta-peptides and competitive overlay assays showed that the binding site of A to brain proteins resides between 12–28 amino acid sequence of A. The presence of several intracellular A-binding (AB) proteins suggests that these proteins may either protect A from its fibrillization or alternatively promote A polymerization. Identification of these proteins and their binding affinities for A are needed to assess their potential role in the pathogenesis of Alzheimer's disease.     

Is hemoglobin A2 important?

There have been claims that the minor hemoglobin A₂ in some primates is functionally and adaptively less vital than hemoglobin A (Boyer et al., 1971). These claims are criticized on several grounds, and evidence suggesting that selection acts on hemoglobin A₂ is reviewed. An estimate is made of the proportion of mutations in the , , , and genes that could be expected to be selectively neutral. Evidence is presented that the gene is better buffered from the effects of mutation than is the gene and that is better buffered than , although quantitatively the latter difference is very small.     

GABA A -receptors: Structural requirements and sites of gene expression in mammalian brain

GABA_A-receptors, the major synaptic targets for the neutotransmitter GABA, are gated chloride channels. By their allosteric drug-induced modulation they serve as molecular control elements through which the levels of anxiety, vigilance, muscle tension and epileptiform activity can be regulated. Despite their functional prominence, the structural requirements of fully functional GABA_A-receptors are still elusive. Expression of cDNAs coding for the ₁- and ₁-subunits of rat brain yielded GABA-gated chloride channels which were modulated by barbiturates but displayed only agonistic responses to ligands of the benzodiazepine receptor. GABA_A-receptors with fully functional benzodiazepine receptor sites were formed when the ₁- and ₁-subunits were coexpressed with the ₂-subunit of rat brain. These receptors, however, failed to show cooperativity of GABA in gating the channel. In order to determine the subunit repertoire available for receptor assembly in different neuronal populations in vivo, the sites of subunit gene expression were (₁, ₂, ₃, ₅, ₆, ₁, ₂, ₃, ₂) mapped by in situ hybridization histochemistry in brain sections. The mRNAs of the ₁-, ₁- and ₂-subunits were co-localized e.g. in mitral cells of olfactory bulb, pyramidal cells of hippocampus as well as granule cells of dentate gyrus and cerebellum. The lack of colocalization in various other brain areas points to an extensive receptor heterogeneity. The presence of multiple GABA_A-receptors in brain may contribute to synaptic plasticity, differential responsiveness of neurons to GABA and to variations in drug profiles.Special issue dedicated to Dr. Erminio Costa     

Mechanism of sex determination inRumex hastatulus Baldw.

Summary The results presented indicate that the sex determination mechanism in the Texas race ofR. hastatulus 2n = 10 (XX + 8A); 2n = 10 (XX + 8A)] is intermediate between theX/Y andX/A systems. In this race, sex is determined to some extent by theX/A balance, but theY chromosome also affects sex expression, maleness or intersexuality being correlated with different ratios ofX andY chromosomes.The results obtained for the Texas race are fully compatible with data presented by Smith (1963) for the North Carolina race [ 2n = 8 (XX + 6A); 2n = 9 (XX ₁ Y ₂ + 6A)]. It may be concluded that evolution of the karyotype in this species is not accompanied by changes in the mechanism of sex determination.     

Variability in gold bead density in cells

Summary Variability in gold bead distribution between individual cells was demonstrated in both pituitary melanotrophic cells immunocytochemically reacted for ACTH and in neurohypophysia terminals reacted for oxytocin-neurophysin. Gold beads were confined to the secretory granules compartment of both tissues. Density of gold beads in melanotrophic cells reacted for ACTH varied from 100–480 gold beads/m². A much narrower range of gold beads distribution (460–900 gold beads/m²) was observed in axons of the neurohypophysis reacted with anti-oxytocin-neurophysin. These results indicate that the labeling density varies from cell to cell (as well as axon terminals) within morphologically homogeneous population. Thus, it may reflect certain physiological differences between cells. A suggestion is being made that mean gold bead density coefficient of variation should be calculated by comparison between individual cells.     

Comparative Modulation by 3α,5α and 3β,5β Neurosteroids of GABA Binding Sites During Avian Central Nervous System Development

Neurosteroids are endogenous Central Nervous System (CNS) compounds which act mainly by allosteric modulation of the GABA_A receptor complex. The presence of a 3-hydroxyl group and a 5-hydrogen atom have been found to be essential structural requirements for biological activity in mammals. In the present work we report the enhancing activity on [³H]GABA binding to its receptor sites in chick optic lobe produced by progesterone metabolites 3-hydroxy,5-pregnan-20-one (3,5-P) and 3-hydroxy,5-pregnan-20-one (3,5-P). Both steroids were found able to enhance [³H]GABA binding along ontogeny, displaying a similar profile at early developmental stages, while in adulthood 3,5-P had greater potency (EC₅₀ 0.22 M) and enhancing effect (E_max: 122%). In adult synaptic membranes, the two compounds displayed a complex interaction with the GABA_A receptor, disclosed by a Schild plot with slope below one and an incomplete displacement of 3,5-P by its 3,5 isomer. Such complexity could be related to the steroidogenic profile in avian CNS, with 5-reduced progesterone metabolites present since early development, while 3,5-P is found only in adulthood. Bearing in mind differences between avian and mammalian steroidogenic profiles and the relevance of 5-steroids in early avian development, we propose that 3,5-P, instead of the classical potent 3,5-steroids, may be the endogenous modulator of GABAergic activity in developing avian brain.     

Establishment of a gene tagging system in Arabidopsis thaliana based on the maize transposable element Ac

Summary An Ac-derived, two-component transposable element system has been developed and analyzed with respect to its use in Arabidopsis thaliana. This system consists of an immobilized Ac element (Ac clipped wing, Ac_cl) as the source of transactivating transposase and a nonautonomous Ds element, Ds_A, which is inserted into a chimaeric neomycinphosphotransferase gene used as excision marker. After separate introduction of Ac_c1 and Ds_A into Arabidopsis thaliana, progeny analysis of crosses between five different Ac_cl lines and seven different Ds_A lines shows that: (1) different Ac_cl lines differ greatly in their capacity to transactivate Ds_A; (2) different Ds_A lines do not differ significantly with respect to Ds_A transactivation by one Ac_cl line; (3) reintegration of excised Ds_A elements, both at (genetically) linked and unlinked sites, occurs in about 50% of the excision events; and (4) plants with a high rate of somatic excisions can be used as source of new Ds_A transpositions, allowing the creation of a large number of independent Ds_A insertions.     

Contextualizing Images in Dreams and Daydreams

A contextualizing image (CI) is a powerful central image of a dream which appears to contextualize (provide a picture-context for) the dreamer's emotion. For instance, dreamers who have experienced any serious traumatic event sometimes dream, I was overwhelmed by a tidal wave. This appears to picture their feeling of terror and/or vulnerability.A scoring system for CIs is examined here and is applied to dreams and daydreams supplied by 40 students. Two raters scoring dreams on a blind basis showed good inter-rater reliability. Recent dreams were shown to have more as well as more intense CIs than recent daydreams; likewise, dreams that stand out had more intense CIs than daydreams that stand out. Students with thin boundaries had more and more intense CIs than students with thick boundaries in their recent dreams and nightmare, but not so clearly in dreams and nightmares that stand out. The emotions judged as contextualized by the powerful images tended towards fear/terror and helplessness/vulnerability in dreams (especially in dreams that stand out) whereas emotions contextualized by images in daydreams showed a wide range with no clusters.     

On the axonal migration of catecholamines in constricted sciatic nerve of the rat

Summary The radioautographic technique has been used to study the axonal migration of catecholamines in sympathetic fibres of the sciatic nerve of rats after ligature. A first series of experiments aimed at ascertaining the capacity of the proximal portion of adrenergic fibres to take up and store exogenous tritiated catecholamines (³H-DOPA; ³H-DA and ³H-NA) 3 to 22 hours after ligation. The results are qualitatively similar in rats pretreated and non-pretreated with IMAO, but the intensity of the radioautographic reaction is lower in the latter cases. Most of the labeled axons are filled mainly with vesicular and tubular profiles of endoplasmic reticular origin, large dense bodies (probably lysosomes) and/or mitochondria. The silver grains are generally superimposed on the vesicular and/or the tubular profiles, but in some cases on the large dense bodies, suggesting that exogenous catecholamines can be stored in lysosomes. The question whether SGV specifically store catecholamines also in the modified adrenergic fibres has been investigated in KMnO₄ fixed material. These results show that beside a large number of fibres in which there is a strict correlation between labeling and SGV, some fibres containing SGV do not retain the ³H-NA. Conversely some fibres which contain mainly agranular vesicles display radioautographic reaction. Therefore, in case of ligated fibres, SGV cannot be considered the specific organelles for storage of catecholamines.The axonal migration of labeled catecholamines has been studied in animals pretreated with IMAO. A moderate, but selective, labeling is present in the proximal portion of sciatic fibres of rats in which administration of labeled catecholamine preceeded of 2 hours the ligature and this was performed 22 hours before fixation.From these combined types of experiments, it is concluded that despite the presence of all the structures necessary for the storage of a high amount of catecholamines in the modified adrenergic fibres, only a small fraction of catecholamines accumulated above the ligature has been transported by axonal migration. Therefore, the axonal migration of catecholamines appears as an epiphenomenon related to the distal migration of enzymatic and storage proteins from the perikaryon.Part of this work was presented to the 84th Annual Session of American Association of Anatomists (Sotelo and Taxi, 1971).Some of the electron microscopic observations were made at the Unité 106 I.N.S.E.R.M.     

Occurrence and distribution of neuropeptide-Y-immunoreactive nerves in the respiratory tract and middle ear

Summary Nerve fibres displaying neuropeptide-Y (NPY) immunoreactivity are abundantly distributed in the respiratory tract of cats, guinea-pigs, rats and mice. Fine beaded NPY fibres were seen in whole-mount spreads of the middle-ear mucosa. In the nasal mucosa and in the wall of the Eustachian tube NPY fibres were numerous around arteries and arterioles but sparse in the vicinity of veins; single fibres were found close to the acini of seromucous glands. In the tracheobronchial wall NPY fibres occurred in the proximity of blood vessels, in the subepithelial layer and in the smooth muscle. Surgical and chemical (6-hydroxydopamine treatment) sympathectomy resulted in disappearance of adrenergic and NPY-containing nerve fibres in the nasal mucosa. Sequential staining with antibodies against dopamine--hydroxylase (DBH) and NPY revealed that DBH and NPY occur in the same perivascular nerve fibres in the nasal mucosa. The distribution of NPY fibres in the respiratory tract suggests multiple functions of NPY, such as regulation of local blood flow, glandular secretion and smooth muscle activity.