Pseudomonas helleri sp. nov. and Pseudomonas weihenstephanensis sp. nov., isolated from raw cow's milk.

Analysis of the microbiota of raw cow's milk and semi-finished milk products yielded seven isolates assigned to the genus Pseudomonas that formed two individual groups in a phylogenetic analysis based on partial rpoD and 16S rRNA gene sequences. The two groups could be differentiated from each other and also from their closest relatives as well as from the type species Pseudomonas aeruginosa by phenotypic and chemotaxonomic characterization and average nucleotide identity (ANIb) values calculated from draft genome assemblies. ANIb values within the groups were higher than 97.3 %, whereas similarity values to the closest relatives were 85 % or less. The major cellular lipids of strains WS4917T and WS4993T were phosphatidylethanolamine, phosphatidylglycerol and diphosphatidylglycerol; the major quinone was Q-9 in both strains, with small amounts of Q-8 in strain WS4917T. The DNA G+C contents of strains WS4917T and WS4993T were 58.08 and 57.30 mol%, respectively. Based on these data, strains WS4917T, WS4995 ( = DSM 29141 = LMG 28434), WS4999, WS5001 and WS5002 should be considered as representatives of a novel species of the genus Pseudomonas, for which the name Pseudomonas helleri sp. nov. is proposed. The type strain of Pseudomonas helleri is strain WS4917T ( = DSM 29165T = LMG 28433T). Strains WS4993T and WS4994 ( = DSM 29140 = LMG 28438) should be recognized as representing a second novel species of the genus Pseudomonas, for which the name Pseudomonas weihenstephanensis sp. nov. is proposed. The type strain of Pseudomonas weihenstephanensis is strain WS4993T ( = DSM 29166T = LMG 28437T).

Brain Responses during the Anticipation of Dyspnea.

Dyspnea is common in many cardiorespiratory diseases. Already the anticipation of this aversive symptom elicits fear in many patients resulting in unfavorable health behaviors such as activity avoidance and sedentary lifestyle. This study investigated brain mechanisms underlying these anticipatory processes. We induced dyspnea using resistive-load breathing in healthy subjects during functional magnetic resonance imaging. Blocks of severe and mild dyspnea alternated, each preceded by anticipation periods. Severe dyspnea activated a network of sensorimotor, cerebellar, and limbic areas. The left insular, parietal opercular, and cerebellar cortices showed increased activation already during dyspnea anticipation. Left insular and parietal opercular cortex showed increased connectivity with right insular and anterior cingulate cortex when severe dyspnea was anticipated, while the cerebellum showed increased connectivity with the amygdala. Notably, insular activation during dyspnea perception was positively correlated with midbrain activation during anticipation. Moreover, anticipatory fear was positively correlated with anticipatory activation in right insular and anterior cingulate cortex. The results demonstrate that dyspnea anticipation activates brain areas involved in dyspnea perception. The involvement of emotion-related areas such as insula, anterior cingulate cortex, and amygdala during dyspnea anticipation most likely reflects anticipatory fear and might underlie the development of unfavorable health behaviors in patients suffering from dyspnea.

Congenitally altered motor experience alters somatotopic organization of human primary motor cortex.

Human motor development is thought to result from a complex interaction between genes and experience. The well-known somatotopic organization of the primate primary motor cortex (M1) emerges postnatally. Although adaptive changes in response to learning and use occur throughout life, somatotopy is maintained as reorganization is restricted to modifications within major body part representations. We report of a unique opportunity to evaluate the influence of experience on the genetically determined somatotopic organization of motor cortex in humans. We examined the motor "foot" representation in subjects with congenitally compromised hand function and compensatory skillful foot use. Functional magnetic resonance imaging (fMRI) and transcranial magnetic stimulation (TMS) of M1 revealed that the foot was represented in the classical medial foot area of M1 and was several centimetres away in nonadjacent cortex in the vicinity of the lateral "hand" area. Both areas had direct output to the spinal motor neurons innervating foot muscles and were behaviorally relevant because experimental disruption of either area by TMS altered reaction times. We demonstrate a unique, nonsomatotopically organized M1 in humans, which emerged as a function of grossly altered motor behavior from the earliest stages of development. Our results imply that during early motor development experience may play a more critical role in the shaping of genetically determined neural networks than previously assumed.

Detection of mutagenic activity of metronidazole and niridazole in body fluids of humans and mice.

After humans were treated at therapeutic doses with the trichomonacide metronidazole (Flagyl) and the antischistosomal agent niridazole mutagenic activity was demonstrable in their urines when tested with the histidine auxotroph of Salmonella typhimurium. Both compounds were active in the host-mediated assay in mice, and evidence of activity was found in the blood and urine of mice treated with niridazole but not with metronidazole.

Dyspnea catastrophizing and neural activations during the anticipation and perception of dyspnea.

Dyspnea is an aversive symptom in various diseases. High levels of negative affectivity are typically associated with increased dyspnea and changes in its neural processing. Recently, more dyspnea-specific forms of negative affectivity such as dyspnea catastrophizing were suggested to contribute to increased perception of dyspnea beyond effects of rather unspecific negative affectivity such as general anxiety levels. The involved neural mechanisms have not yet been explored. Therefore, the present retrospective analysis examined the associations of dyspnea catastrophizing with neural activations during the anticipation and perception of dyspnea. Sixty-six healthy volunteers underwent 20 blocks of inspiratory resistive load breathing with parallel acquisition of fMRI data. Loads inducing either severe or mild dyspnea (dyspnea conditions) were presented in alternating order, with each condition being visually cued (anticipation conditions). Dyspnea catastrophizing and general trait anxiety were measured with the Breathlessness Catastrophizing Scale (BCS) and the State-Trait Anxiety Inventory, respectively. Correlating the BCS scores with neural activations during the perception of dyspnea yielded no significant results. However, during the anticipation of dyspnea, BCS scores correlated positively with activations of the anterior cingulate cortex (ACC), even after controlling for general anxiety levels. These activations in the ACC were not related to concurrent respiratory parameters. Results suggest that dyspnea catastrophizing in healthy volunteers is associated with stronger ACC recruitment during dyspnea anticipation. Given the established role of the ACC in processing affective states, affect regulation, and antinociception, this might reflect increased affective and/or top-down modulatory processing in individuals with higher dyspnea catastrophizing when anticipating dyspnea.

Breathlessness amplifies amygdala responses during affective processing.

Breathlessness is an aversive symptom in many prevalent somatic and psychiatric diseases and is usually experienced as highly threatening. It is strongly associated with negative affect, but the underlying neural processes remain poorly understood. Therefore, using fMRI, the present study examined the effects of breathlessness on the neural processing of affective visual stimuli within candidate brain areas including the amygdala, insula, and anterior cingulate cortex (ACC). During scanning, 42 healthy volunteers, mean (SD) age: 29.0 (6.0) years, 14 female, were presented with affective picture series of negative, neutral, and positive valence while experiencing either no breathlessness (baseline conditions) or resistive-load induced breathlessness (breathlessness conditions). Respiratory measures and self-reports suggested successful induction of breathlessness and affective experiences. Self-reports of breathlessness intensity and unpleasantness were significantly higher during breathlessness conditions, mean (SD): 45.0 (16.6) and 32.3 (19.8), as compared to baseline conditions, mean (SD): 1.9 (3.0) and 2.9 (5.5). Compared to baseline conditions, stronger amygdala activations were observed during breathlessness conditions for both negative and positive affective picture series relative to neutral picture series, while no such effects were observed in insula and ACC. The present findings demonstrate that breathlessness amplifies amygdala responses during affective processing, suggesting an important role of the amygdala for mediating the interactions between breathlessness and affective states.

The contribution of metronidazole and two metabolites to the mutagenic activity detected in urine of treated humans and mice.

The urine of two patients receiving therapeutic doses of the trichomonacide, metronidazole, was analyzed for mutagenic activity using the histidine auxotroph TA1535 of Salmonella typhimurium. The activity detected in the urine was significantly higher than could be accounted for by the presence of the administered drug. Chromatographic analysis of the urine indicated the presence of the metabolite 1-(2-hydroxyethyl)-2-hydroxymethyl-5-nitroimidazole, which when tested in vitro with TA1535 was found to be ten times more active than metronidazole. An additional urinary metabolite, 1-acetic acid-2-methyl-5-nitroimadazole, was found to be inactive when similarly tested. The in vitro mutagenic activity of metronidazole and the two metabolites was unchanged by the addition of phenobarbital- or Aroclor-induced rat liver homogenate to the test system. In addition, metronidazole and the hydroxymethyl metabolite reverted S. typhimurium TA100 but not TA1537, TA1538, or TA98, and the acetic acid metabolite failed to revert any of the tester strains. In studies with mice, metronidazole was required in excess of the human dose in order for significant amounts of the hydroxymethyl metabolite to be detected in the urine. Urine from mice pretreated with the hepatotoxin, carbon tetrachloride, prior to the administration of metronidazole demonstrated approximately a 50% reduction in mutagenic activity, and the formation of the urinary metabolites was inhibited. These findings indicate the production of metabolites from the parent compound by the liver of the intact animal which could not be determined by use of the standard in vitro liver homogenate system.

Structural Brain Changes in Patients With COPD.

BACKGROUND: Patients with COPD suffer from chronic dyspnea, which is commonly perceived as highly aversive and threatening. Moreover, COPD is often accompanied by disease-specific fears and avoidance of physical activity. However, little is known about structural brain changes in patients with COPD and respective relations with disease duration and disease-specific fears. METHODS: This study investigated structural brain changes in patients with COPD and their relation with disease duration, fear of dyspnea, and fear of physical activity. We used voxel-based morphometric analysis of MRI images to measure differences in generalized cortical degeneration and regional gray matter between 30 patients with moderate to severe COPD and 30 matched healthy control subjects. Disease-specific fears were assessed by the COPD anxiety questionnaire. RESULTS: Patients with COPD showed no generalized cortical degeneration, but decreased gray matter in posterior cingulate cortex (whole-brain analysis) as well as in anterior and midcingulate cortex, hippocampus, and amygdala (regions-of-interest analyses). Patients' reductions in gray matter in anterior cingulate cortex were negatively correlated with disease duration, fear of dyspnea, and fear of physical activity. Mediation analysis revealed that the relation between disease duration and reduced gray matter of the anterior cingulate was mediated by fear of physical activity. CONCLUSIONS: Patients with COPD demonstrated gray matter decreases in brain areas relevant for the processing of dyspnea, fear, and antinociception. These structural brain changes were partly related to longer disease duration and greater disease-specific fears, which might contribute to a less favorable course of the disease.

Brain mechanisms of short-term habituation and sensitization toward dyspnea.

Dyspnea is a prevalent and threatening cardinal symptom in many diseases including asthma. Whether patients suffering from dyspnea show habituation or sensitization toward repeated experiences of dyspnea is relevant for both quality of life and treatment success. Understanding the mechanisms, including the underlying brain activation patterns, that determine the dynamics of dyspnea perception seems crucial for the improvement of treatment and rehabilitation. Toward this aim, we investigated the interplay between short-term changes of dyspnea perception and changes of related brain activation. Healthy individuals underwent repeated blocks of resistive load induced dyspnea with parallel acquisition of functional magnetic resonance imaging data. Late vs. early ratings on dyspnea intensity and unpleasantness were correlated with late vs. early brain activation for both, dyspnea anticipation and dyspnea perception. Individual trait and state anxiety were determined using questionnaire data. Our results indicate an involvement of the orbitofrontal cortex (OFC), midbrain/periaqueductal gray (PAG) and anterior insular cortex in habituation/sensitization toward dyspnea. Changes in the anterior insular cortex were particularly linked to changes in dyspnea unpleasantness. Changes of both dyspnea intensity and unpleasantness were positively correlated with state and trait anxiety. Our findings are in line with the suggested relationship between the anterior insular cortex and dyspnea unpleasantness. They further support the notion that habituation/sensitization toward dyspnea is influenced by anxiety. Our study extends the known role of the midbrain/PAG in anti-nociception to an additional involvement in habituation/sensitization toward dyspnea and suggests an interplay with the OFC.

Amygdala response to anticipation of dyspnea is modulated by 5-HTTLPR genotype.

Dyspnea anticipation and perception varies largely between individuals. To investigate whether genetic factors related to negative affect such as the 5-HTTLPR polymorphism impact this variability, we investigated healthy, 5-HTTLPR stratified volunteers using resistive load induced dyspnea together with fMRI. Alternating blocks of severe and mild dyspnea ("perception") were differentially cued ("anticipation") and followed by intensity and unpleasantness ratings. In addition, volunteers indicated their anticipatory fear during the anticipation periods. There were no genotype-based group differences concerning dyspnea intensity and unpleasantness or brain activation during perception of severe vs. mild dyspnea. However, in risk allele carriers, higher anticipatory fear was paralleled by stronger amygdala activation during anticipation of severe vs. mild dyspnea. These results suggest a role of the 5-HTTLPR genotype in fearful dyspnea anticipation.

Catecholaminergic control of alpha-melanocyte-stimulating hormone (alpha MSH) release by frog neurointermediate lobe in vitro: evidence for direct stimulation of alpha MSH release by thyrotropin-releasing hormone.

The role of dopaminergic and adrenergic innervation of the intermediate lobe of amphibian pituitary in the release of alpha MSH has been studied in vitro. Neurointermediate lobes of frog (Rana ridibunda Pallas) have been perifused in amphibian culture medium (ACM) for 5-7 h. alpha MSH released in the effluent perifusate was measured by means of a sensitive and specific RIA. No significant morphological alteration of neurointermediate lobe cells was observed during the perifusion experiment, even at the electron microscopic level. The existence of dopaminergic receptors, responsible for an inhibition of frog melanotrophs, was shown using the dopaminergic agonists apomorphine (10(-6) M) and bromo-2-ergocryptine (10(-8) and 10(-7) M), which initiated a marked reduction of alpha MSH secretion. The effect of apomorphine was obliterated by the dopaminergic antagonist haloperidol. Haloperidol itself induced a dose-related stimulation, and the monoamine oxidase inhibitor nialamide (4 x 10(-3) M) inhibited alpha MSH secretion. In addition, haloperidol led to a complete reversal of the inhibitory effect of nialamide on alpha MSH secretion. These results demonstrate the existence, in the parenchyme of the intermediate lobe, of dopaminergic nerve fibers that are functionally active. The beta-adrenergic agonist isoproterenol was responsible for a dose-related stimulation of alpha MSH secretion; the stimulatory effect was reversed by the beta-adrenergic antagonist propranolol. TRH is a potent stimulator of alpha MSH secretion in amphibians. Since haloperidol and propranolol did not abolish the stimulation of alpha MSH release induced by TRH, it appeared that TRH action was not mediated via an inhibition of dopamine release or via a stimulation of adrenergic nerve fibers.

Histoautoradiographic detection of oxytocin- and vasopressin-binding sites in the telencephalon of the rat.

Localization of oxytocin- and vasopressin-binding sites has so far been studied in the rat brain by means of film autoradiographs. The disposal of iodinated ligands with high specificity has allowed us to develop histoautoradiography on emulsion-coated sections and to reinvestigate on a microscopic scale the distribution of these sites in the telencephalon (septum, striatopallidal system, amygdala and hippocampus). This technique showed that oxytocin and vasopressin labelling presented distinct distributions and coincided with delimited zones, corresponding to anatomical subdivisions defined on cytoarchitectural and immunocytochemical bases. Vasopressin sites were seen in the dorsal and intermediate parts of the lateral septum and the juxtacapsular nucleus of the bed nucleus of the stria terminalis. Oxytocin sites were located in the ventral and intermediate parts of the lateral septum, the oval and the principal nuclei of the bed nucleus of the stria terminalis and the septofimbrial nucleus. In the striatopallidal system, vasopressin sites were found in the accumbens nucleus and the fundus striati, whereas oxytocin sites were in the accumbens nucleus, the head, and the posterolateral parts of the caudate-putamen, the striatal cell bridges, and the olfactory tubercle. In the amygdala, vasopressin sites were not found, but oxytocin sites were located in the central, medial, and basomedial nuclei. In the hippocampus, vasopressin sites were located in the dentate gyrus (polymorph and molecular layers), and oxytocin sites, in the subiculum (molecular and pyramidal layers) and in the field CA1 of Ammon's horn (lacunosum moleculare and pyramidal layers). The localization of the binding sites at the microscopic level permitted us to reinvestigate whether or not correlation existed in a same area between innervation, electrophysiological effects, and presence of binding sites.

Distribution and morphometric characteristics of oxytocin- and vasopressin-immunoreactive neurons in the rabbit hypothalamus.

The distribution, morphological features, and morphometric characteristics of cell bodies producing oxytocin (OT) and vasopressin (AVP) were studied in the rabbit hypothalamus by means of a conventional immunoperoxidase method. The aim of the present study was to determine the existence or not of a species-specific OT-cell group that might be involved in the dense OT innervation of the intermediate lobe in the leporidae. No OT-cell group clearly distinct from the hypothalamic paraventricular (PVN) and supraoptic (SON) nuclei was found, even in colchicine-treated animals. Most immunoreactive perikarya were found within these nuclei. In addition, small AVP neurons occurred in the suprachiasmatic nucleus. In the SON, the predominant, tightly packed AVP cells occupied the ventral part of the nucleus, whereas OT neurons were dorsolaterally located. The PVN presented a loose organization without any obvious subdivision. OT cells, which predominated, occupied the medial part of the nucleus. The PVN had a prominent rostral anterobasal extension composed mainly of OT cells. Laterally to the nucleus, numerous large AVP neurons, with few and smaller OT cells, dispersed along the neurosecretory tract without forming definite cell clusters. AVP cell bodies had a rough granular aspect contrasting with the smooth and fine one of OT cells. Spinelike processes were rarely observed on the perikarya, except on large scattered AVP neurons, but frequently covered the proximal dendrites of both types of neurons. Throughout the hypothalamus, OT neurons had definitely smaller mean somal areas and were more homogeneous in size than AVP cells.

Co-localization of tyrosine hydroxylase, GABA and neuropeptide Y within axon terminals innervating the intermediate lobe of the frog Rana ridibunda.

Possible co-existence of gamma-aminobutyric acid (GABA), catecholamines, and neuropeptide Y (NPY) in the same nerve terminals of the frog intermediate lobe was investigated by immunocytochemistry at the electron microscopic level. Co-localization of GABA and tyrosine hydroxylase (TH) was studied by using a double immunogold labeling procedure. Co-localization of glutamate decarboxylase (GAD) and NPY was studied by combining, respectively, the peroxidase-antiperoxidase method and a radioimmunocytochemical labeling procedure. Catecholamines and GABA were systematically co-localized in nerve endings of the pars intermedia. Most of the NPY-immunoreactive fibers also contained GAD-like immunoreactivity. However, a few NPY-positive nerve terminals were not immunoreactive for GAD. These data provide evidence for co-existence of a regulatory peptide (NPY) and several neurotransmitters (i.e., GABA and catecholamines) within the same axon terminals in the intermediate lobe. Since GABA, dopamine, and NPY have all been shown to inhibit the activity of frog melanotrope cells, the present findings suggest that these neuroendocrine factors may interact either at the pre-synaptic or post-synaptic level.

Quantitative autoradiographic mapping of neurohypophysial hormone binding sites in the rat forebrain and pituitary gland--I. Characterization of different types of binding sites and their distribution in the Long-Evans strain.

Oxytocin and vasopressin binding sites were localized and characterized by quantitative autoradiography on consecutive sections of Long-Evans rat forebrains and pituitary glands, incubated in the presence of 5 nM [3H]oxytocin or 5 nM [3H]vasopressin. In the forebrain, two types of neurohypophysial hormone binding sites were thus defined. (1) Oxytocin/vasopressin sites with similar nanomolar-range affinities for [3H]oxytocin and [3H]vasopressin; both tritiated peptides were displaced from these sites in the presence of 10 microM of either oxytocin or vasopressin. The main areas bearing such sites were the ventral subiculum, several nuclei of the amygdala, the ventromedial hypothalamic nucleus, the bed nucleus of the stria terminalis and the olfactory tubercle. (2) Selective vasopressin sites, binding [3H]vasopressin with nanomolar-range affinity and [3H]oxytocin with a much lower affinity; these sites were not labelled in the presence of 5 nM [3H]oxytocin, and 10 microM oxytocin displaced [3H]vasopressin binding by 80%. Such sites occurred in several thalamic nuclei, in the dopaminergic A13 cell group of the zona incerta, the suprachiasmatic nucleus, the fundus striati and the lateral septal nucleus. No selective oxytocin sites were detected. Different oxytocin and vasopressin binding characteristics were found in the hypothalamo-neurohypophysial system. In the paraventricular and supraoptic nuclei and in the pituitary neural lobe the [3H]vasopressin binding density was twice that of [3H]oxytocin; vasopressin was always more potent than oxytocin in displacing both [3H]vasopressin and [3H]oxytocin binding from those sites. Interaction of the tritiated peptides with neurophysins cannot be completely ruled out in these locations. The present data are discussed in correlation with the functional roles of the neurohypophysial peptides in the brain and the pharmacological characteristics of their receptors.

Quantitative autoradiographic mapping of neurohypophysial hormone binding sites in the rat forebrain and pituitary gland--II. Comparative study on the Long-Evans and Brattleboro strains.

The anatomical distribution and pharmacological characteristics of the different types of neurohypophysial hormone binding sites were compared in the forebrains and pituitary glands of Long-Evans rats and its mutant Brattleboro strain, genetically deficient in vasopressin. Quantitative autoradiography on sections incubated in the presence of 5 nM of either [3H]oxytocin or [3H]vasopressin revealed the presence of the same types of sites in the brains of both strains but noticeable variations in their densities were found in several areas. In the forebrain, oxytocin/vasopressin sites, which bind both peptides with similar high nanomolar affinities, had the same locations and densities in the ventral subiculum, in several nuclei of the amygdala, the bed nucleus of the stria terminalis and the olfactory tubercle. The density of such sites was, in contrast, lower in the ventromedial hypothalamic nucleus of the Brattleboro rat. Selective vasopressin sites which bind [3H]vasopressin with a nanomolar-range affinity and [3H]oxytocin with a much lower affinity showed more variations. They were not found in the Brattleboro rat thalamus but were highly concentrated in several thalamic nuclei in the Long-Evans rat. Conversely, their densities were higher in the dopaminergic A13 cell group of the zona incerta and the suprachiasmatic nucleus of the Brattleboro rat. Their densities were similar in the lateral septal nucleus and in the fundus striati of both strains. In the hypothalamo-neurohypophysial system, [3H]oxytocin and [3H]vasopressin binding occurred in the Long-Evans rat with characteristics different from those found in other brain areas. In the Brattleboro rat, no [3H]vasopressin binding and only low [3H]oxytocin binding, restricted to the magnocellular nuclei, were found.(ABSTRACT TRUNCATED AT 250 WORDS)

Three distinct thyrotropin-releasing hormone-immunoreactive axonal systems project in the median eminence-pituitary complex of the frog Rana ridibunda. Immunocytochemical evidence for co-localization of thyrotropin-releasing hormone and mesotocin in fibers innervating pars intermedia cells.

The localization of thyrotropin-releasing hormone-immunoreactive structures was investigated in the hypothalamo-hypophyseal complex of the frog, Rana ridibunda, by light and electron microscopy using the conventional indirect immunoperoxidase technique and the immuno-gold technique, respectively. The localization of mesotocin-, vasotocin- and neurophysin-immunoreactive elements was compared to that of thyrotropin-releasing hormone either by comparing homologous fields on serial sections or by staining the same section with two different antibodies. Thyrotropin-releasing hormone-immunoreactive perikarya occurred mainly in the anterobasal periventricular area and dorsal extension of the preoptic nucleus, and in the lateral zone of the infundibular nucleus. In the anterobasal preoptic nucleus, the distribution of thyrotropin-releasing hormone-immunoreactive perikarya partially overlapped that of vasotocin- and mesotocin-containing neurons; however, co-localization of thyrotropin-releasing hormone with either nonapeptide could not be detected there. In contrast, in the caudal extension of the preoptic nucleus, thyrotropin-releasing hormone- and mesotocin-like immunoreactivities were frequently co-localized in the same neurons. In the external zone of the median eminence, abundant networks of thyrotropin-releasing hormone- and vasotocin-immunoreactive nerve fibers were found in the vicinity of portal capillaries, while mesotocin-immunoreactive axons were only found in the internal zone. Using the immuno-gold technique at the electron microscopic level, three distinct thyrotropin-releasing hormone-immunoreactive systems were identified in the median eminence-neurointermediate lobe complex. (1) In the external zone of the median eminence, a conspicuous population of pericapillary endings contained 100-nm dense core vesicles immunoreactive solely for thyrotropin-releasing hormone. (2) In the neural lobe of the pituitary, thyrotropin-releasing hormone immunoreactivity occurred on secretory vesicles in a subpopulation of the mesotocinergic axons containing 160-nm secretory granules; co-localization with vasotocin was never seen. (3) In the intermediate lobe, thyrotropin-releasing hormone- and mesotocin (or neurophysin I)-immunoreactivities were systematically found in the same 120-nm dense core vesicles; these thyrotropin-releasing hormone-/mesotocin-immunoreactive axon terminals frequently made synaptic contacts with melanotropic cells. The possible modulatory effect of mesotocin on thyrotropin-releasing hormone-induced alpha-melanocyte-stimulating hormone secretion was investigated using perifused frog neurointermediate lobes. Administration of graded doses of mesotocin (from 10(-10) to 10(-5) M) did not affect the spontaneous release of alpha-melanocyte-stimulating hormone. In addition, mesotocin (10(-7) and 10(-6) M) did not modify thyrotropin-releasing hormone-evoked alpha-melanocyte-stimulating hormone release.(ABSTRACT TRUNCATED AT 400 WORDS)

GABAergic innervation of somatostatin-containing neurosecretory cells of the anterior periventricular hypothalamic area: a light and electron microscopy double immunolabelling study.

Double immunolabelling on semithin sections revealed glutamate decarboxylase immunopositive dots surrounding somatostatin-containing cell sections in the rat periventricular hypothalamic area. Up to 12 appositions were observed per cell section with an average number of 2-3 and a unimodal distribution. At the electron microscopical level pre-embedding staining of glutamate decarboxylase showed that most immunoreactive elements consisted of immunolabelled axonal endings. Most of these glutamate decarboxylase immunopositive boutons were found within the neuropil where they frequently made synapses on unidentified dendrites. Some of them were apposed to somatostatin-containing cell bodies that were identified according to the presence of immunolabelled granules using combined immunogold post-embedding staining. In many instances glutamate decarboxylase immunoreactive endings were also found to be involved in synaptic contact with somatostatin-labelled perikarya, or neuronal processes. These contacts provide the morphological basis for a direct GABAergic control of the somatostatin-containing cells regulating the secretion of growth hormone.

Pharmacological characteristics and anatomical distribution of [3H]oxytocin-binding sites in the Wistar rat brain studied by autoradiography.

Oxytocin-binding sites were detected by autoradiography on rat brain sections incubated in the presence of the [3H]oxytocin. These sites were characterized pharmacologically using quantitative autoradiography. High pressure liquid chromatography controls of the incubation media indicated that labelling was due to the intact [3H]oxytocin molecule. Pharmacological analysis of different locations (central amygdaloid nucleus, ventral subiculum and ventromedial hypothalamic nucleus) showed that the sites detected had a high affinity for oxytocin and also for arginine-vasopressin. In contrast, some areas known to bind vasopressin intensely, such as suprachiasmatic and lateral septum nuclei, had little or no affinity for oxytocin. Autoradiographs revealed [3H]oxytocin-binding sites in already known brain areas (olfactory centres, ventral subiculum, central amygdaloid nucleus, bed nucleus of the stria terminalis) albeit with more extensive labelling of some of these formations, in particular, the amygdaloid complex. In addition, specific [3H]oxytocin-binding sites were found in areas not yet reported to bind oxytocin, such as the paraventricular thalamic and caudate nuclei. In the hypothalamus, specific binding sites were not detected in the supraoptic and paraventricular nuclei: the only structure labelled was the ventrolateral part of the ventromedial nucleus. Discrepancies between the concentrations of [3H]oxytocin-binding sites, the known distribution of oxytocin-containing endings and electrophysiological data indicate that autoradiography, under our conditions, apparently only reveals some of the oxytocin receptors in the brain. Thus, in the hypothalamus, no relationship can be established between the known effect of oxytocin on oxytocinergic magnocellular neurons and detection of specific [3H]oxytocin-binding sites. Autoradiography may reveal mainly oxytocin-binding sites in areas receiving diverse "parasynaptic" information, where oxytocin might play a modulatory role rather than exerting rapid, short-term effects of the neurotransmitter type.

Oxytocinergic innervation of autonomic nuclei controlling penile erection in the rat.

In the rat, spinal autonomic neurons controlling penile erection receive descending pathways that modulate their activity. The paraventricular nucleus of the hypothalamus contributes oxytocinergic fibers to the dorsal horn and preganglionic sympathetic and parasympathetic cell columns. We used retrograde tracing techniques with pseudorabies virus combined with immunohistochemistry against oxytocin and radioligand binding detection of oxytocinergic receptors to evidence the oxytocinergic innervation of thoracolumbar and lumbosacral spinal neurons controlling penile erection. Spinal neurons labelled with pseudo-rabies virus transsynaptically transported from the corpus cavernosum were present in the intermediolateral cell column and the dorsal gray commissure of the thoracolumbar and lumbosacral spinal cord. Confocal laser scanning microscopic observation of the same preparations revealed close appositions between oxytocinergic varicosities and pseudorabies virus-infected neurons, suggesting strongly the presence of synaptic contacts. Electron microscopy confirmed this hypothesis. Oxytocin binding sites were present in the superficial layers of the dorsal horn, the dorsal gray commissure and the intermediolateral cell column in both the thoracolumbar and lumbosacral segments. In rats, stimulation of the paraventricular nucleus induces penile erection, but the link between the nucleus and penile innervation remains unknown. Our findings support the hypothesis that oxytocin, released by descending paraventriculo-spinal pathways, activates proerectile spinal neurons.