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M L Tappaz

Publications and source records attributed to M L Tappaz.

At least 37 records · Page 2Linked to original sources

GABAergic neurons in dissociated cultures of rat hypothalamus, septum, and midbrain.

Cultures of dissociated tissue blocks from 18 day-old rat embryos containing either septum plus anterior hypothalamus, retrochiasmatic hypothalamus, or midbrain, were maintained for 8-29 days in vitro. GABAergic neurons were visualized by incubation with (3H) gamma-aminobutyric acid followed by autoradiography and by immunocytochemistry using an antiserum against glutamic acid decarboxylase. Positive neurons were of variable shapes and sizes. Labeled neurons in the three cultured regions represented 3.4, 1.1 and 1.7%, respectively, of all neurons. Apart from a few exceptions, autoradiographic labeling and immunocytochemical staining coincided. Glial cells did not stain and showed very low grain densities. Radioactivity could be released from preloaded cultures upon depolarization by 50 mM potassium and 100 microM Veratridine. The demonstration of GABAergic neurons in cultures that are known to contain a variety of peptidergic neurons opens up the possibility of studying the interaction between the two classes of neurons in vitro.

Animals↗

Anatomical relationships of dopaminergic and GABAergic systems with the GnRH-systems in the septo-hypothalamic area. Immunohistochemical studies.

Immunohistochemical double staining for gonadotropin releasing hormone (GnRH) and tyrosine hydroxylase (TH) or glutamic acid decarboxylase (GAD) reveals in the septo-preoptic-diagonal band complex of the rat brain close spatial associations between GnRH-immunoreactive perikarya and TH and GAD immunoreactive fibers. In the organum vasculosum laminae terminalis, no close spatial relationships could be observed between TH- or GAD-positive fibers and the GnRH-containing system. In contrast, in the median eminence substantial overlap exists in the distribution of GnRH with TH and GAD containing nerve fibers. This overlap is most intense for TH throughout the lateral palisade zone, while for GAD it is more restricted to the outermost portion of the external palisade zone. The results suggest that dopamine and GABA influence GnRH secretion via axosomatic contacts in the septo-preoptic-diagonal band complex, as well as via axo-axonic interactions in the median eminence, while no such interactions seem to exist in the organum vasculosum laminae terminalis. Since dopaminergic cell bodies in the ventral hypothalamus are closely apposed by GnRH and GAD containing fibers, the existence of feedback circuits among GnRH, dopamine and GABA systems is proposed.

Animals↗

Light- and electron-microscopic immunocytochemistry of glutamic acid decarboxylase (GAD) in the basal hypothalamus: morphological evidence for neuroendocrine gamma-aminobutyrate (GABA).

GABAergic cells and axon terminals were localized in the basal hypothalamus of differnet species (rat, mouse and cat), by means of an immunocytochemical approach using a specific and well-characterized antiserum to the GABA biosynthetic enzyme, glutamate decarboxylase. Light-microscopic visualization was performed with an indirect immunofluorescence method and electron-microscopic observations were made on material with pre-embedding staining and use of the peroxidase-antiperoxidase procedure. At the light-microscopic level, a dense immunofluorescent plexus was observed over both the medial and lateral parts of the external layer of the median eminence. The labelling extended from the rostal part of the median eminence up to the pituitary stalk. Over the subependymal and internal layers only a few immunoreactive dots were visible, except around the blood vessels where they appeared more concentrated. Immunoreactive varicosities could be found following the outlines of the capillary loops and lining tanycyte processes, especially in the median eminance midportion. At the electron-microscopic level, the immunolabelling was exclusively found over neuronal profiles in the median eminence. The latter represented a small fraction of the total number of varicosities visible on the same section. Labelled profiles typically contained numerous small clear synaptic vesicles and only a few or no dense-core vesicles. In the subependymal and internal layers, rare labelled endings were found close to ependymal cells or among transversally cut fibers, respectively. In the palisadic zone, elongated positive boutons were visible intermingled with bundles of unlabelled axons and glial or ependymal processes. In the neurohemal contact zone, immunoreactive endings were observed among unlabelled neurosecretory endings in close vicinity to fenestrated capillary perivascular space. Small moderately intense immunofluorescent varicosities were observed all over the hypothalamus. The density of the glutamate decarboxylase-positive network was higher than in most diencephalic regions. Intraventricular or topical injection of colchicine allowed the visualization of small lightly immunoreactive cells in the diffusion area of colchicine. In the arcuate nucleus labelled axonal endings containing small pleomorphic synaptic vesicles and sometimes a few dense-core vesicles were observed at the electron-microscopic level. Typical synaptic junctions were commonly found between positive endings and unlabelled perikarya, or more frequently, unlabelled dendrites. These findings show that glutamate decarboxylase-containing endings are localized ed in several strategic sites for potential GABAergic neuroendocrine regulations. The GABAergic endings found among neurosecretory endings in the neurohemal contact zone may provide the morphological support for the release of gamma-aminobutyrate into the portal blood flow as an hypothalamic hypophysiotropic hormone.

Animals↗

Central GABAergic innervation of neurointermediate pituitary lobe: biochemical and immunocytochemical study in the rat.

Activity of glutamic acid decarboxylase GluDCase, the biosynthetic enzyme of gamma-aminobutyric acid (GABA) was measured in low-speed homogenate supernatant of the neural and intermediate (neurointermediate) lobe (28--30 pmol of CO2 per microgram of protein per hr) and of the anterior lobe (2--4 pmol of CO2 per microgram of protein per hr). In the neurointermediate lobe, stalk transection reduced the GluDCase activity by more than 95%. By using an antiserum to rat brain GluDCase and the unlabeled antibody--peroxidase method of Sternberger, GluDCase immunoreactivity was localized in many terminals within the neurointermediate lobe of the hypophysis. In pars intermedia, immunoreactive terminals occurred in apposition to secretory cells and to glial cells and were near nonimmunoreactive axonal profiles; in pars neuralis they were apposed to pituicytes and to unlabeled axons including the neurosecretory terminals and were along fenestrated portal capillaries. GluDCase immunoreactive axons terminals exhibited diverse morphological features and would not have been identified as a distinct population without the GluDCase antiserum. No GluDCase-immunoreactivity was found in the anterior pituitary lobe. Stalk transection abolished GluDCase immunoreactivity in the neurointermediate lobe. These data provide biochemical and morphological evidence for a central GABAergic innervation of neural and intermediate lobes of the hypophysis.

Animals↗

Decrease of glutamate decarboxylase (GAD)-immunoreactive nerve terminals in the substantia nigra after kainic acid lesion of the striatum.

Antiserum sheep 3 against rat brain glutamate decarboxylase (GAD) was employed for the immunohistochemical localization of GAD-immunoreactive nerve terminals in the substantia nigra (SN). To test whether the antiserum specifically localized GAD-containing axon terminals, the effect of kainic acid-induced striatal lesions on the reactive nerve endings in the SN was investigated. Seven days after the injection of 1 microgram kainic acid into the striatum, a 65% decrease in GAD-enzyme activity occurred in the ipsilateral SN. On immunohistochemical examination there was correspondingly a marked reduction of GAD-positive terminals. The parallel decrease in biochemical and immunohistochemical GAD activity indicates that antiserum sheep 3 can be used as a specific immunohistochemical probed for GAD-containing elements.

Animals↗

On the origin of substance P and glutamic acid decarboxylase (GAD) in the substantia nigra.

Knife cuts in the frontal plane separating the anterior part of the caudate-putamen from the globus pallidus resulted in marked decreases in substances P levels in the reticular part of the substantia nigra. More caudal knife cuts were required in order to effect maximal decreases in nigral glutamic acid decarboxylase levels. Thus, there is a clear anatomical dissociation between the striatal neurons which project to the reticular part of the substantia nigra and which contain SP, and the more caudally located GAD-containing striatal and pallidal neurons, all of which travel through the globus pallidus on their way to the substantia nigra.

Animals↗

Origin of glutamate-decarboxylase (GAD)-containing cells in discrete hypothalamic nuclei.

Glutamate decarboxylase activity (GAD) was assayed in discrete hypothalamic nuclei in the rat following lesions of the major afferent pathways from hippocampus, amygdala, midbrain, septum, thalamus and globus pallidus. None of these lesions led to a marked decrease in GAD activity in selected nuclei. After total deafferentation of the medial-basal hypothalamus GAD remained unchanged in the median eminence but fell markedly in the ventromedial and arcuate nuclei. In these two nuclei a decrease of GAD still occurred following partial deafferentation from lateral and posterior hypothalamus, but not from anterior and preoptic areas. These results indicated that most GAD-containing cells have their origin inside of the hypothalamus. In this region GABAergic neurons are likely to be short interneurons providing intrahypothalamic connections. Such connections are suggested from lateral and posterior hypothalamus onto the medial basal nuclei.

Afferent Pathways↗

Glutamate decarboxylase (GAD) and gamma-aminobutyric acid (GABA) in discrete nuclei of hypothalamus and substantia nigra.

The distribution of L-glutamate decarboxylase activity (GAD) and gamma-aminobutyric acid (GABA) was investigated in the hypothalamic nuclei and in parts of the substantia nigra in the rat. GAD varied markedly among these areas. The reticular part of the nigra showed the highest activity two-fold higher than any other nucleus. Among the hypothalamic nuclei, a 5-fold difference was found between the poorest and richest nuclei. High GAD was measured in the preoptic, anterior and dorsomedial nuclei. Low activity was found in arcuate and supraoptic nuclei. The lowest GAD activity was measured in the median eminence with only half the activity of the whole brain homogenate. This suggests that GABAergic neurones might not be involved in neuroendocrine regulation at the median eminence level. GABA was determined using the sensitive cycling microassay. The rats were killed by microwaves, procedure which was found to inactivate enzymatic processes within two sec without affecting the level or the distribution of GABA. Postmortem increments during the first 3 min following decapitation ranged up to 5 times the endogenous levels, and were proportional to the GAD activity of the corresponding nucleus. This confirms that GAD is the limiting factor in GABA synthesis and suggests that GABA turnover might be rapid. Endogenous GABA showed a uniform distribution within hypothalamic nuclei and nigra. No relationship appeared between endogenous GABA levels and GAD activities in the various nuclei. These results suggest that while GABA synthesis is likely to occur in non-evenly distributed nerve cells, most GABA may be stored in surrounding cells, presumably glia cells.

Aminobutyrates↗

Effect of surgical isolation of the hypothalamus on its neurotransmitter content.

The concentrations of norepinephrine, dopamine-beta-hydroxylase, dopamine, tyrosine hydroxylase, phenylethanolamine-N-methyltransferase, serotonin, tryptophan hydroxylase, histamine, glutamic acid decarboxylase, and choline acetyltransferase were determined in selected hypothalamic nuclei and in the median eminence after deafferentation of the medial basal hypothalamus. Norepinephrine and dopamine-beta-hydroxylase fell markedly while dopamine and tyrosine hydroxylase did not. Serotonin also decreased in all regions studied; histamine decreased in none. Choline acetyltransferase, phenylethanolamine-N-methyltransferase, and glutamic acid decarboxylase declined in some areas, but not in others.

Animals↗

Distribution of glutamate decarboxylase in discrete brain nuclei.

The L-glutamate decarboxylase (GAD) levels of 70 rat brain nuclei were determined. The distribution of GAD is uneven. High AAD activity was found in the substantia nigra, the colliculi, some hypothalamic nuclei (e.g. dorsomedial) some nuclei of the limbic system (e.g. tractus diagonalis and accumbens) and the medial forebrain bundle. The reticular part of the substantia nigra had the highest activity, two-fold higher than any other nucleus. Low GAD activity was found in the nuclei of the pons, the hypothalamic median eminence and the reticular formation. Moderate activity was measured in most areas studied. The biochemical mapping of GAD is discussed in relation to possible GABA pathways in the brain.

Animals↗