Effect of glutaraldehyde fixation on the localization of various oxidative and hydrolytic enzymes in the brain of rhesus monkey, Macaca mulatta

Synopsis Histochemical investigations have been made on the localization of certain oxidative and hydrolytic enzymes in the different areas of rhesus monkey brain using unfixed, freshfrozen tissue and 3% glutaraldehyde-fixed material. After glutaraldehyde fixation, the oxidative enzymes lose most of their activity normally demonstrable in the fresh-frozen section. The hydrolytic enzymes are somewhat resistant to fixation but also lose about half of the enzyme activity observed after no fixing procedure. The glycogen is better preserved in the glutaraldehyde-fixed material compared to fresh-frozen or even formaldehyde-fixed tissue. The significance of these observations is discussed in relation to glutaraldehyde as a fixative of choice in electron histochemistry.List of abbreviations used in the Figures ALH area lateralis hypothalami - APH area posterior hypothalami - AS aquaeductus Sylvii - ATN anterior thalamic nuclei - BC brachium conjunctivum - CC corpus callosum - CD nucleus caudatus - CI capsula interna - CIS cortex insularis - CM centrum medianum thalami - COR corona radiata - CP commissura posterior - CSR colliculus superior - EM eminentia medialis - F fornix - GC substantia grisea centralis - GLM corpus geniculatum laterale, magnocellular part - GLP corpus geniculatum laterale, parvocellular part - GP globus pallidus - LD nucleus lateralis dorsalis thalami - LME lamina medullaris externa thalami - LMI lamina medullaris interna thalami - LP nucleus lateralis posterior thalami - MD nucleus medialis dorsalis thalami - ML nucleus lateralis corpus mammillaris - MM nucleus medialis corpus mammillaris - NC nucleus centralis thalami - NCI nucleus colliculi inferioris - NLL nucleus lemnisci lateralis - NR nucleus ruber - NSTH nucleus subthalamicus - N III nervus oculomotorius - PC nucleus paracentralis thalami - PCR pedunculus cerebri - PUT Putamen - PV nucleus paraventricularis hypothalami - R nucleus reticularis thalami - RU nucleus reuniens thalami - SM stria medullaris thalami - SMH nucleus supramammillaris hypothalami - SMT nucleus submedius thalami - SN substantia nigra - TO tractus opticus - VL nucleus ventralis lateralis thalami - VP nucleus ventralis posterior thalami - ZI zona incerta - II ventriculus lateralis - III ventriculus tertius     

Histochemical mapping of succinic dehydrogenase and cytochrome oxidase in the diencephalon and basal telencephalic centers of the brain of squirrel monkey (Saimiri sciureus)

Summary The distribution of succinic dehydrogenase (SDA) and cytochrome oxidase (Cy. O) was mapped in the various diencephalic nuclei and basal telencephalic centers of the squirrel monkey brain. Thirty thick formaldehyde fixed serial sections were also studied for the delineation of the various nuclei, but histochemical preparations proved equally useful for this purpose. Strong SDA and Cy. O activity were observed in the habenular, pulvinaris, anterior dorsalis and ventralis, lateralis dorsalis and posterior, and ventral posterior nuclei. The corpus geniculatum laterale and mediale also showed a strong reaction for these enzymes. The nucleus ventralis anterior, which occupies a very large area in the rostral part of the thalamus, showed moderately strong activity in the cellular patches and negligible activity in the thick fiber bundles passing through it. A comparatively weak reaction was observed in the midline thalamic nuclei. The nucleus caudatus and putamen, however, showed very strong SDA/Cy. O activity. The hypothalamic nuclei showed mild Cy. O and moderate SDA reactions, except for the nucleus ventromedialis hypothalami. The latter showed a little stronger enzyme reaction. The fibers of the internal capsule and the anterior commissure showed little SDA and mild Cy. O activity. The various nuclei of the amygdaloid complex showed similar histochemical reactions with moderate SDA and mild Cy. O activity.This work has been carried out with the aid of Grant No. 00165 from the Animal Resources Branch, National Institutes of Health, and a grant from the National Foundation for Neuro-Muscular Diseases and Nasa Grant NGR-11-001-016.     

Distribution of oxytocin and vasopressin neurons in the diencephalon of the Japanese horseshoe bat, Rhinolophus ferrumequinum. An immunohistochemical study.

The distribution of oxytocin (OXT) and vasopressin (VP) neurons in the diencephalon of the hibernating Japanese horseshoe bat, Rhinolophus ferrumequinum, was immunohistochemically investigated by the avidin-biotin complex method. Magnocellular OXT and VP neurons were localized mainly in the paraventricular nucleus and the supraoptic nucleus. In addition to these main nuclei, both kinds of magnocellular neurons were also found in the periventricular nucleus, perifornical area and lateral hypothalamic area. Extensively distributed parvocellular neurons containing only VP were observed in the rostral and middle portions of the suprachiasmatic nucleus. The size of OXT and VP magnocellular neurons was almost equal in the paraventricular and ventromedial supraoptic nuclei, whereas VP neurons were significantly larger than OXT neurons in the dorsolateral supraoptic nucleus. The OXT and VP cells in the ventral supraoptic nucleus showed a distinctive elliptical shape. Both OXT and VP fibers were distributed in the lateral habenular nucleus, stria medullaris thalami, lateral preoptic area, stria terminalis, and medial and supracapsular part of the bed nucleus of the stria terminalis. Moreover, OXT fibers were found in the substantia nigra, and VP fibers were noted in the nucleus reunions and the paraventricular nucleus of the thalamus.     

The architecture of the magnocellular hypothalamus.

The distribution pattern of the magnocellular elements has been described using enzyme histochemical and classical staining procedures. The main amount of magnocellular neurons is found within nuclei, which, in addition are interconnected by strips of large neurons. These strips are organized as oblique lamellae traversing the hypothalamic area. The upper most lamella forms the dorsal hypothalamic border.     

Histochemical distribution of acetylcholinesterase and simple esterases in the brain of squirrel monkey (Saimiri sciureus)

Summary The distribution of acetylcholinesterase (AChE) and simple esterases (SE) has been investigated in 15 thick fresh frozen sections of the squirrel monkey brain. Simple esterases are cellular enzymes, found in locations similar to those of acid phosphatase (AC), although not as abundantly. The neuropil in most of the nuclei of the brain shows stronger SE activity compared to the AChE reaction. The AChE activity is strong in nucleus caudatus and putamen, and appreciable quantities of this enzyme are found in nucleus fasciculus diagonalis band of Broca, habenular complex, nucleus interpeduncularis, cranial nerve nuclei, gray layers of colliculus superior, griseum pontis, nuclei olivaris inferior, cuneatus, gracilis, etc. The nucleus interpeduncularis has proved very interesting histochemically because it is rich in AChE and SE, as well as acid phosphatase, monoamine oxidase and lactic dehydrogenase. In the area postrema, the neurons give stronger SE activity than the parenchymal cells, while the AChE activity in both types of cells is similar. The molecular layer of cerebellum is stronger in AChE compared to SE, whereas the Purkinje cells and granule cells are strong in SE and show only negligible AChE activity. The lining cells of the choroid plexus, in addition to the ependymal cells, demonstrate a negligible to mild AChE reaction in comparison to moderately strong simple esterase activity. The significance of these observations has been discussed.This work has been carried out with the aid of Grant No. 00165 from the Animal Resources Branch, National Institute of Health and a grant (NGR-11-001-016) from The National Aeronautics and Space Administration. Thanks are due to Mrs. M. J. Nimnicht and Miss M. E. Rogero for their technical help.     

Enzyme histochemistry of the mesenteric and dorsal root ganglion cells of cat and squirrel monkey

Summary A detailed histochemical study has been made on the mesenteric ganglia of the cat, and dorsal root ganglia of the squirrel monkey by the use of appropriate histochemical techniques accompanied by appropriate controls for phosphatases, esterases, and oxidative enzymes. The different neurons of a particular ganglion show varied amounts of enzyme activity at a particular time depending upon the functional state of the neurons. SDH, CYO and LDH reaction is prominent in the cytoplasm of the neurons, gliocytes and satellite cells, whereas the MAO preparations generally show a weak reaction. The AK is prominent in the neuropil, cell membranes and peripheral part of cytoplasm, whereas ATPase activity has been observed in blood vessels as well. In AC preparations the area of lipofuscin concentration shows more intense reaction than the rest of the cytoplasm. The activity of AChE and BChE varies from mild, to moderate to strong. The TPPase preparations show morphologically different types and amounts of TPPase positive Golgi material even in the adjoining cells. The relationship between the TPPase Golgi material and various oxidative and dephosphorylating enzymes has been briefly discussed.Abbreviations used AC Acid phosphatase - AChE Acetyl-cholinesterase (specific) - AK Alkaline phosphatase - AMPase Adenosine monophosphatase (5-nucleotidase) - ATPase Adenosine triphosphatase - BChE Butyryl-cholinesterase (nonspecific) - CYO Cytochrome oxidase - DPN-D DPN-diaphorase - G6P Glucose-6-phosphatase - LDH Lactic dehydrogenase - MAO Monoamine oxidase - MDH Malic dehydrogenase - NAD-D NAD-diaphorase - SDH Succinic dehydrogenase - SE Simple esterase - TPPase thiamine pyrophosphatase T. R. Shanthaveerappa in previous publications.     

Selective specificity of calcium-binding proteins calbindin and calretinin expression in the magnocellular neurosecretory hypothalamic nuclei of tortoises and turtles

We have studied the distribution of calcium-binding proteins in the magnocellular neurosecretory nuclei of nonapeptidergic neurosecretory nuclei of the preoptic–hypothalamic complex in a tortoise (Testudo horsfieldi) and a pond turtle (Emys orbicularis) using immunohistochemistry. We have found that different types of cells in the paraventricular and supraoptic nuclei predominantly express calbindin and, to a lesser extent, calretinin, but not parvalbumin. The selective calbindin/calretinin control of the neurohormone secretion in these hypothalamic nuclei is an evolutionary conservative feature typical of reptiles and mammals.     

The function of the hypophysis without preoptic neurosecretory control

Summary In a large number of adult female and male specimens of Rana temporaria, a total extirpation of the magnocellular neurosecretory preoptic nuclei was performed. Several months after operation, no regeneration of the neurosecretory nuclei had occurred. The results showed 1. that all A.F.-positive neurosecretory nerve fibres of the hypothalamo-hypophysial region originate from nerve cell bodies localized in the magnocellular preoptic nuclei; 2. that the magnocellular preoptic nuclei do not play an important part in the inhibitory control of the activity of the pars intermedia of Rana temporaria; 3. that the possible role of the magnocellular preoptic nuclei in the seasonal development of the gonads and of the secondary sexual characteristics of Rana, could be only of secondary importance; 4. that, in accordance with previous experiments, in Rana, ovulation is dependent on preoptic hypothalamic structures. Some results of the present and of other experiments can be interpreted as supporting the hypothesis of a possible dual higher control over the gonadotropic centre (Dierickx, 1965b, 1966, 1967) of the pars ventralis of the tuber cinereum.A part of these studies has been reported previously in abstract form (Dierickx, 1963a, 1963b, 1965a).     

Corticotropin-releasing factor mRNA in the hypothalamus is affected differently by drinking saline and by dehydration

Corticotropin-releasing factor (CRF) stimulates the synthesis and release of adrenocorticotropin in the anterior pituitary and may help maintain fluid and electrolyte balance. 'Salt-loaded' rats had an increase in CRF mRNA in hypothalamic magnocellular neurons of the paraventricular and supraoptic nuclei and a decrease in message in the parvocellular paraventricular neurons. After salt-loaded rats were adrenalectomized, CRF mRNA increased in the parvocellular cells. In contrast to salt loading, water deprivation lead to a decrease in CRF mRNA in magnocellular and parvocellular neurons. These results show that CRF synthesis within separate populations of hypothalamic neurons is regulated differently under various conditions.     

Immunocytochemical localization of somatostatin-containing neurons in the rat hypothalamus

Summary The rat hypothalamus was studied at the light microscopic level with the use of single and double immunocytochemical staining methods. It was shown that the rat supraoptic and paraventricular hypothalamic nuclei, and their accessory neurosecretory nuclei, do not contain magnocellular somatostatin neurons. The distribution of the hypothalamic parvocellular somatostatin cells is described. The parvocellular component of the rat hypothalamic paraventricular nucleus is, at least partly, composed of somatostatin cells: they form a fairly well circumscribed periventricular cell mass. The rat suprachiasmatic nuclei contain separate somatostatin neurons and vasopressin neurons. Scattered somatostatin cells are present in the entire arcuate nucleus. In addition to the periventricular somatostatin cells located in the preopticanterior hypothalamic area and in the arcuate nucleus, the rat hypothalamus also contains numerous scattered somatostatin cells located distant from the third ventricle.This investigation was supported by a grant from the Belgian Nationaal Fonds voor Geneeskundig Wetenschappelijk Onderzoek     

Histological and histochemical observations on the neurosecretory cells in the diencephalon of Chthonerpeton indistinctum and Ichthyophis paucisulcus (Gymnophiona,Amphibia)

Summary In the diencephalon of two species of Gymnophiona (Amphibia) two neurosecretory nuclei were examined with histological (Alcian Blue, Aldehyde Fuchsin, Brookes Trichrome stain) and enzyme histochemical techniques (acid phosphatase, -naphthyl acetate esterase, acetylcholinesterase (AChE)). In the preoptic nucleus two categories of secretory neurons were distinguished: large and medium sized neurons. The perikarya of both cell types contain very little neurosecretory material. The Alcian Blue method stained the medium sized neurons faintly but selectively. The tractus praeopticohypophyseus is marked by the presence of Herring bodies, which, however, are relatively scarce. The neurohypophysis, in contrast, contains large amounts of neurosecretory material. Both cell types of the preoptic nucleus are characterized by their very strong AChE and -naphthylacetate esterase activity. The AChE also marks the tractus praeoptico-hypophyseus. In the large neurons acid phosphatase is present around the nucleus; in the medium sized neurons this enzyme is concentrated close to the origin of the axon. In the dorso-caudal hypothalamus a small group of neurons is stained with Alcian-Blue. These neurons, which also contain AChE, are located immediately under the ependyma which seems to be specialized in this region.The financial support of the Deutsche Forschungsgemeinschaft is gratefully acknowledged (We 380/5)     

Colocalization of neuropeptide Y (NPY)-like and FMRFamide-like immunoreactivities in the brain of the Atlantic salmon (Salmo salar)

Summary The colocalization of the peptides neuropeptide Y (NPY) and Phe-Met-Arg-Phe-NH₂ (FMRFamide) in the brain of the Atlantic salmon was investigated with double immunofluorescence labeling and peroxidase-antiperoxidase immunocytochemical techniques. Colocalization of NPY-like and FMRE amide-like immunoreactivities was observed in neuronal cell bodies and fibers in four brain regions: in the lateral and commissural nuclei of the area ventralis telencephali, in the nucleus ventromedialis thalami, in the laminar nucleus of the mesencephalic tegmentum, and in a group of small neurons situated among the large catecholaminergic neurons in the isthmal region of the brainstem. All cell bodies in these nuclei were immunoreactive to both NPY and FMRF. We consistently observed larger numbers of FMRF-immunoreactive than NPY-immunoreactive fibers. In the nucleus ventromedialis thalami NPY- and FMRFamide-like immunoreactivities were colocalized in cerebrospinal fluid (CSF)-contacting neurons. NPY-immunoreactive, but not FMRF-immunoreactive, neurons were found in the stratum periventriculare of the optic tectum, and at the ventral border of the nucleus habenularis (adjacent to the nucleus dorsolateralis thalami). Neurons belonging to the nucleus of the nervus terminalis were FMRF-immunoreactive but not NPY-immunoreactive. The differential labeling indicates, as do our cross-absorption experiments, that the NPY and FMRFamide antisera recognize different epitopes. Thus, it is probable that NPY-like and FMRFamide-like substances occur in the same neurons in some brain regions.     

Distribution of osmoregulatory peptides and neuronal-glial configuration in the hypothalamic magnocellular nuclei of desert rodents

The desert rodents Psammomys obesus and Gerbillus tarabuli live under extreme conditions and overcome food and water shortage by modes of food and fluid intake specific to each species. Using immunohistochemistry and electron microscopy, we found that the hypothalamic magnocellular nuclei, and in particular, their vasopressinergic component, is highly and similarly developed in Psammomys and Gerbillus. In comparison to other rodents, the hypothalamus in both species contains more magnocellular VP neurons that, together with oxytocin neurons, accumulate in distinct and extensive nuclei. As in dehydrated rodents, many magnocellular neurons contained both neuropeptides. A striking feature of the hypothalamic magnocellular system of Psammomys and Gerbillus was its display of ultrastructural properties related to heightened neurosecretion, namely, a significant reduction in glial coverage of neuronal somata and dendrites in the hypothalamic nuclei. There were many neuronal elements whose surfaces were directly juxtaposed and shared the same synapses. Their magnocellular nuclei also showed a high level of sialylated isoform of the Neural Cell Adhesion Molecule (PSA-NCAM) that underlies their capacity for neuronal and glial plasticity. These species thus offer striking models of structural neuronal and glial plasticity linked to natural conditions of heightened neurosecretion.     

Autoradiographic studies with 3H dihydrotestosterone in the brain of sex reversed mice, heterozygous for androgen insensitive testicular feminization (Tfm). A comparison with normal female mice and sex reversed male mice

Specific binding sites for 3H dihydrotestosterone are demonstrated by autoradiography in brain nuclei of sex reversed mice heterozygous for testicular feminization (Tfm) which are phenotypically intersexes with testes and accessory sex glands that consist of a mosaic of androgen insensitive Tfm cells which lack specific dihydrotestosterone binding and androgen sensitive normal cells. The nuclear group evaluated include: nucleus (n.) septi lateralis, n. interstitialis striae terminalis, n. medialis amygdalae, the hypothalamic n. arcuatus, n. ventromedialis lateralis, n. pre-mammillaris ventralis, n. preopticus medialis, and nuclei of the cranial nerves VII, X, and XII. In the sex reversed males and the female, used as controls, the frequency of neurons with specific DHT binding show a distinct male-female difference in the caudal part of the arcuate nucleus. In the sex reversed Tfm heterozygotes, in all brain nuclei studied, the frequency of labeled neurons is reduced. The extent of reduction of androgen binding in the different brain nuclei varies among as well as within individual sex reversed Tfm heterozygotes, suggesting variations of the ratio of normal to Tfm neurons in sex reversed Tfm heterozygotes. The differentially reduced androgen binding of different brain systems corresponds to a differentially reduced androgen dependent behaviour reported in the literature.     

Distribution of NT-IR perikarya in the brain of the guinea pig with special reference to cardiovascular centers in the medulla oblongata

Summary The occurrence and distribution of neurotensin-immunoreactive (NT-IR) perikarya was studied in the central nervous system of the guinea pig using a newly raised antibody (KN 1). Numerous NT-IR perikarya were found in the nuclei amygdaloidei, nuclei septi interventriculare, hypothalamus, nucleus parafascicularis thalami, substantia grisea centralis mesencephali, ventral medulla oblongata, nucleus solitarius and spinal cord. The distribution of NT-IR perikarya was similar to that previously described in the rat and monkey. In the gyrus cinguli, hippocampus and nucleus olfactorius, though, no NT-IR neurons were detected in this investigation. Additional immunoreactive perikarya, however, were observed in areas of the ventral medulla oblongata, namely in the nucleus paragigantocellularis, nucleus retrofacialis and nucleus raphe obscurus.The relevance of the NT-IR perikarya within the ventral medulla oblongata is discussed with respect to other neuropeptides, which are found in this area, and to cardiovascular regulation.Abbreviations abl nucleus amygdaloideus basalis lateralis - abm nucleus amygdaloideus basalis medialis - acc nucleus amygdaloideus centralis - aco nucleus amygdaloideus corticalis - ahp area posterior hypothalami - ala nucleus amygdaloideus lateralis anterior - alp nucleus amygdaloideus lateralis posterior - ame nucleus amygdaloideus medialis - atv area tegmentalis ventralis - bst nucleus proprius striae terminalis - CA commissura anterior - CC corpus callosum - cgld corpus geniculatum laterale dorsale - cglv corpus geniculatum laterale ventrale - cgm corpus geniculatum mediale - CHO chiasma opticum - CI capsula interna - co nucleus commissuralis - cod nucleus cochlearis dorsalis - cp nucleus caudatus/Putamen - cs colliculus superior - cu nucleus cuneatus - dmh nucleus dorsomedialis hypothalami - DP decussatio pyramidum - em eminentia mediana - ent cortex entorhinalis - epi epiphysis - FLM fasciculus longitudinalis medialis - fm nucleus paraventricularis hypothalami pars filiformis - FX fornix - gd gyrus dentatus - gp globus pallidus - gr nucleus gracilis - hl nucleus habenulae lateralis - hm nucleus habenulae medialis - hpe hippocampus - ift nucleus infratrigeminalis - io oliva inferior - ip nucleus interpeduncularis - LM lemniscus medialis - MT tractus mamillo-thalamicus - na nucleus arcuatus - nls nucleus lateralis septi - nms nucleus medialis septi - npca nucleus proprius commissurae anterioris - ns nucleus solitarius - n III nucleus nervi oculomotorii - nt V nucleus tractus spinalis nervi trigemini - ntm nucleus mesencephalicus nervi trigemini - osc organum subcommissurale - P tractus cortico-spinalis - PC pedunculus cerebri - PCI pedunculus cerebellaris inferior - pir cortex piriformis - pol area praeoptica lateralis - pom area praeoptica medialis - prt area praetectalis - pt nucleus parataenialis - pvh nucleus paraventricularis hypothalami - pvt nucleus paraventricularis thalami - r nucleus ruber - re nucleus reuniens - rgi nucleus reticularis gigantocellularis - rl nucleus reticularis lateralis - rm nucleus raphe magnus - ro nucleus raphe obscurus - rp nucleus raphe pallidus - rpc nucleus reticularis parvocellularis - rpgc nucleus reticularis paragigantocellularis - sch nucleus suprachiasmaticus - SM stria medullaris thalami - snc substantia nigra compacta - snl substantia nigra lateralis - snr substantia nigra reticularis - ST stria terminalis - tad nucleus anterior dorsalis thalami - tam nucleus anterior medialis thalami - tav nucleus anterior ventralis thalami - tbl nucleus tuberolateralis - tc nucleus centralis thalami - tl nucleus lateralis thalami - tmd nucleus medialis dorsalis thalami - TO tractus opticus - TOL tractus olfactorium lateralis - tpo nucleus posterior thalami - tr nucleus reticularis thalami - trs nucleus triangularis septi - TS tractus solitarius - TS V tractus spinalis nervi trigemini - tvl nucleus ventrolateralis thalami - vmh nucleus ventromedialis hypothalami - vh ventral horn, Columna anterior - zi zona incerta Supported by the Deutsche Forschungsgesellschaft (DFG) SFB 90, Carvas     

Central connection of the optic, oculomotor, trochlear and abducens nerves in medaka, Oryzias latipes

Medaka (Oryzias latipes) is one of the few vertebrate experimental animals in which inbred lines have been established. It is also a species that has advanced in genetic studies in a manner comparable to zebrafish. This fish is therefore a good model for studying functional organization of the nervous system, but anatomical analysis of its nervous system has been limited to embryonic stages. In the present study, we investigated anatomy of cranial nerves in adult fish focusing on the visual function, using an inbred strain of medaka. Cranial nerves of medaka were labeled using biocytin, revealing a central distribution of retinofugal terminals, retinopetal neurons, and oculomotor, trochlear and abducens motor neurons. The optic nerve of the adult medaka was of a complete decussation type. Retinofugal terminals were located in 8 brain nuclei, the suprachiasmatic nucleus, nucleus pretectalis superficialis, nucleus dorsolateralis thalami, area pretectalis pars dorsalis (APd), area pretectalis pars ventralis (APv), nucleus of the posterior commissure (NPC), accessory optic nucleus, and the tectum opticum. Retinopetal neurons were identified in 6 brain nuclei, the ganglion of the terminal nerve, preoptic retinopetal nucleus, nucleus dorsolateralis thalami, APd, APv, and NPC. The oculomotor neurons were mostly labeled ipsilaterally and were located dorsomedially, abutting the fasciculus longitudinalis medialis in the mesencephalon. The trochlear nucleus was located contralaterally and dorsolaterally adjacent to the fasciculus longitudinalis medialis in the mesencephalon. The abducens nucleus was located ipsilaterally in a ventrolateral part of the rhombencephalic reticular formation. These results, generally similar to those in other teleosts, provide the basis for future behavioral and genetic studies in medaka.     

Interaction of neuronal NOS and catecholamines in regulation of expression of proteins of apoptosis by vasopressinergic hypothalamic neurons

The work studied vasopressinergic neurons of hypothalamic supraoptic and paravenricular nuclei of the wild type mice and the neuronal nitric oxide synthase (nNOS) gene knockouted mice at a decrease of the brain catecholamine (CA) level caused by administration of the blocker of activity of tyrosine hydroxylase alpha-methyl-paratyrosine (alpha-MPT) and at the CA level decrease on the background of functional activity of the vasopressinergic neurons caused by dehydration of animals. There were analyzed changes in the number of neurons in both magnocellular hypothalamic nuclei expressing proapoptotic proteins caspase-8 and caspase-9, p53, and antiapoptotic protein Bcl-2. The disturbance of the CA-ergic innervation was shown to be a strong damaging factor leading to apoptosis of neurons regardless of the presence of nNOS in the cells. However, at disturbance of the CA-ergic innervation due to the 5-day mouse dehydration, no death of neurons by apoptosis was revealed. Thus, it is possible that functional activation prevents the hypothalamic vasopressinergic neurons from death at a decrease of the CA level in brain. The main difference of the nNOS gene knockouts is the absence of activation of the Bcl-2 expression under all used actions. This confirms our suggestion about interaction of CA and NO in triggering of expression of the antiapoptotic protein Bcl-2.     

Intranuclear coupling of hypothalamic magnocellular nuclei by glutamate synaptic circuits

Magnocellular neurons of the supraoptic nucleus (SON) and paraventricular nucleus (PVN) display bursting activity that is synchronized under certain conditions. They receive excitatory synaptic inputs from intrahypothalamic glutamate circuits, some of which are activated by norepinephrine. Ascending noradrenergic afferents and intrahypothalamic glutamate circuits may be responsible for the generation of synchronous bursting among oxytocin neurons and/or asynchronous bursting among vasopressin neurons located in the bilateral supraoptic and paraventricular nuclei. Here, we tested whether magnocellular neurons of the PVN receive excitatory synaptic input from the contralateral PVN and the region of the retrochiasmatic SON (SONrx) via norepinephrine-sensitive internuclear glutamate circuits. Whole cell patch-clamp recordings were performed in PVN magnocellular neurons in coronal hypothalamic slices from male rats, and the ipsilateral SONrx region and contralateral PVN were stimulated using electrical and chemical stimulation. Electrical and glutamate microdrop stimulation of the ipsilateral SONrx region or contralateral PVN elicited excitatory postsynaptic potentials/currents (EPSP/Cs) in PVN magnocellular neurons mediated by glutamate release, revealing internuclear glutamatergic circuits. Microdrop application of norepinephrine also elicited EPSP/Cs, suggesting that these circuits could be activated by activation of noradrenergic receptors. Repetitive electrical stimulation and drop application of norepinephrine, in some cases, elicited bursts of action potentials. Our data reveal glutamatergic synaptic circuits that interconnect the magnocellular nuclei and that can be activated by norepinephrine. These internuclear glutamatergic circuits may provide the functional architecture to support burst generation and/or burst synchronization in hypothalamic magnocellular neurons under conditions of activation.     

Interaction of neuronal NOS and catecholamines in regulation of expression of proteins of apoptosis by vasopressinergic hypothalamic neurons

The work deals with studies on vasopressinergic neurons of hypothalamic supraoptic and paravenricular nuclei in the wild type mice and the neuronal nitric oxide synthase (nNOS) in the gene knockouted mice at a decrease of the brain catecholamine (CA) level caused by administration of the blocker of activity of tyrosine hydroxylase α-methyl-paratyrosine (α-MPT) and at the CA level decrease on the background of functional activity of the vasopressinergic neurons caused by dehydration of animals. There were analyzed changes in the number of neurons in the magnocellular hypothalamic nuclei expressing proapoptotic proteins caspase-8 and caspase-9, p53, and antiapoptotic protein Bcl-2. Disturbance of the CAergic innervation was shown to be a strong damaging factor leading to apoptosis of neurons regardless of the presence of nNOS in the cells. However, at disturbance of the CAergic innervation due to the 5-day mouse dehydration, no death of neurons by apoptosis was revealed. Thus, it is possible that functional activation prevents the hypothalamic vasopressinergic neurons from death at a decrease of the CA level in brain. The main difference of the nNOS gene knockouts is the absence of activation of the Bcl-2 expression under all used actions. This confirms our suggestion about interaction of CA and NO in triggering of expression of the antiapoptotic protein Bcl-2.     

Comparison of cholinergic systems in Ep neocortical regions of cats with high and low cognitive ability

Cats were behaviorally tested for the ability to solve the abstraction and generalization tasks. Fractions of light (C) and heavy (D) synaptosomes of the associative temporal (Ep) areas were prepared, and subfractions of synaptic membranes and synaptoplasm were isolated. Choline acetyltransferase (ChAT) and acetylcholinesterase (AChE) activity and the content of protein and protein sulphydric (SH-) groups were measured in synaptic subfractions. All the studied characteristics were lower in subfractions C of cats with higher cognitive abilities. In subfractions D, the ChAT activity was correlated neither with ChAT activity in the respective C fraction, nor with cognitive abilities of cats. It is suggested that cholinergic terminals originating from neurons of the basal magnocellular nuclei are concentrated in the C fractions, and those from the cortical cholinergic neurons are concentrated in the D fractions. Physiological significance of the "deficiency" of cholinergic inputs of the Ep areas from the basal magnocellular nuclei in animals with higher cognitive abilities is discussed.