Biomedical subjects

N Mons

Publications and source records attributed to N Mons.

43 records · Page 3Linked to original sources

PubMed1988

Visualization of L-dihydroxyphenylalanine in rat brain by using specific antibodies.

L-Dihydroxyphenylalanine (L-DOPA) was conjugated to different protein carriers with glutaraldehyde (G). During the synthesis of the catecholamine conjugates, precautions were taken in order to preserve the structure of L-DOPA. Reduced and non-reduced conjugates were injected to rabbits according to a specific immunization protocol. Anti-L-DOPA antibody affinity and specificity were evaluated by using ELISA tests. The most immunoreactive compounds were the non-reduced conjugate, L-DOPA = G = BSA and the reduced one, L-DOPA-G-BSA. The other conjugated catecholamines were poorly recognized or not at all. These antisera enabled us to specifically visualize the precursor of the catecholaminergic neurotransmitters which are: dopamine, noradrenaline and adrenaline in the G-fixed rat brains.

Animals↗

PubMed1987

Simultaneous detection of indoleamines and dopamine in rat dorsal raphe nuclei using specific antibodies.

Using a monoclonal antibody against dopamine and a rabbit antiserum against serotonin, 5-methoxytryptamine or tryptamine, we were able to achieve the simultaneous localization of two amines in glutaraldehyde-fixed sections of rat dorsal raphe nuclei. In this staining procedure, the first antigen was localized using 3,3'-diaminobenzidine (DAB), while the second antigen was stained using the 1-naphthol basic dye (2-NBD) method. The two antigens were localized in different cells or structures. No overlap of the staining was observed, thus indicating that dopamine is not localized with serotonin, 5-methoxytryptamine or tryptamine.

3,3'-Diaminobenzidine↗

PubMed1987

Monoclonal antibodies against glutaraldehyde-conjugated dopamine.

Four mice were immunized with dopamine (DA)-glutaraldehyde (G)--protein conjugates over a period of 8-10 weeks. Polyclonal antisera, obtained at various intervals, were tested using an enzyme-linked immunosorbent assay (ELISA). All had anti-conjugated DA antibodies. As soon as good antibody affinity was detected between 10(-10) and 10(-6) M, the mouse yielding the highest apparent affinity was killed, and the spleen was dissected out. Hybridomas were obtained from spleen cells fused with SP2/O/Ag myeloma cells. Supernatant culture media of hybridomas were tested for the presence of anti-conjugated DA antibodies with the ELISA method. Selected hybridomas giving good antibody affinity and specificity were then cloned by the limiting dilution technique. The resulting supernatant culture media were again tested by ELISA. Clones that gave a high antibody affinity (10(-10)-10(-8)M) for G-conjugated DA were used for histochemical localization of DA in rat brain. G-fixed rat brains were sectioned from the telencephalon to the mesencephalon, reduced with sodium borohydride, and prepared for peroxidase-antiperoxidase immunocytochemistry using supernatant (diluted 1:100) or ascites fluid (diluted 1:50,000). Dense networks of very fine fibers were observed in the striatum, septum, and cortex. Numerous immunoreactive cell bodies were found in the ventral tegmental area, the substantia nigra, the hypothalamus, and the dorsal raphe. The ELISA tests and adsorption controls suggested that the monoclonal antibody allowed highly specific detection of DA in tissues.

Aldehydes↗

PubMed1987

Specific antisera against the catecholamines: L-3,4-dihydroxyphenylalanine, dopamine, noradrenaline, and octopamine tested by an enzyme-linked immunosorbent assay.

Antisera were raised against L-3,4-dihydroxyphenylalanine (L-DOPA), dopamine (DA), noradrenaline (NA), and octopamine (OA). This was achieved by coupling each molecule to bovine serum albumin or human serum albumin using glutaraldehyde. The conjugated aromatic amines were kept in a reducing medium containing sodium metabisulfite. Antiserum specificity was tested using an enzyme-linked immunosorbent assay method for catecholamines. Competition experiments were done between the immunogen coated on the well plates and each catecholamine, either in the free state or in conjugated form, previously incubated with an antiserum. In each case, the nonconjugated compound was poorly recognized. The nonreduced conjugates of L-DOPA and DA were well recognized, whereas those of NA and OA were poorly immunoreactive. The cross-reactivity ratios established in the competition experiments allowed the specificity of the immune response to be defined. In each case, it was found to be high. The results suggest that the antibodies of L-DOPA and DA antisera recognize preferentially the catechol moiety, whereas for the anti-NA and anti-OA antibodies, the lateral chain is important.

Animals↗

PubMed1987

First visualization of dopaminergic neurons with a monoclonal antibody to dopamine: a light and electron microscopic study.

A monoclonal antibody recently synthesized against dopamine (DA) was tested in rat and mouse brain sections after further treatment by PAP immunocytochemistry at the light and electron microscopic levels. Distribution of DA-immunoreactive cell bodies was examined in the substantia nigra (sn), the ventral tegmental area (vta), and the raphe nuclei. DA-immunoreactive fibers were investigated in two DA projection systems, the striatum and the septum. Many dopaminergic cell bodies were found in the sn and the vta. Some scattered DA neurons were encountered in the pars reticulata of the sn. The dorsal raphe and linearis raphe nuclei displayed sparse immunoreactive neurons and a dense plexus of DA fibers. Immunoreactive fibers were observed in the entire striatum, more dense in the ventral part. In the septum, immunonegative neurons were outlined by thin DA fibers in synaptic contact with their somata or dendrites. According to our observations, this DA monoclonal antibody seems to be a selective and sensitive tool for studying the dopaminergic neuronal circuitry at both histological and ultrastructural level.

Animals↗

PubMed1984

Isolation and characterization of yeast mutants blocked in mevalonic acid formation.

Yeast mutants defective in beta-hydroxy-beta-methylglutaryl-CoA synthase and acetoacetyl-CoA thiolase have been isolated. Mutants impaired in acetoacetyl-CoA thiolase range into two linked complementation units, erg 10 A and erg 10 B. Mutants deficient in beta-hydroxy-beta-methylglutaryl-CoA synthase belong to two unlinked complementation groups, erg 11 and erg 13. In strictly anaerobic growth conditions, mutants impaired in beta-hydroxy-beta-methylglutaryl-CoA synthase require mevalonic acid in addition to sterol and oleic acid, pointing out the role of mevalonic acid in other physiological function than ergosterol precursor. Growth of mutants impaired in acetoacetyl-CoA thiolase cannot be recovered by mevalonic acid supplementation, suggesting a role of acetoacetyl-CoA or thiolase not linked to sterol pathway.

Acetyl-CoA C-Acetyltransferase↗

PubMedDate not supplied

Evidence for the existence of L-dopa- and dopamine-immunoreactive nerve cell bodies in the caudal part of the dorsal motor nucleus of the vagus nerve.

The precise neurochemical nature of tyrosine hydroxylase-immunoreactive neurons lying in the caudal part of the dorsal motor nucleus of the vagus nerve of the rat has been identified by immunohistochemistry of the catecholamines themselves. This region corresponds precisely to the area where tyrosine hydroxylase has been previously shown to be colocalized with choline acetyltransferase. Adjacent serial cryostat sections from the medulla oblongata and from the cervical spinal cord were treated either for choline acetyltransferase immunohistochemistry, aromatic L-amino acid decarboxylase and tyrosine hydroxylase immunolabelling or for tyrosine hydroxylase, dopamine, noradrenaline and L-dihydroxyphenylalanine (DOPA) immunostaining. The procedure involved the peroxidase-antiperoxidase method and an intensified diaminobenzidine reaction with imidazole. While no noradrenaline-positive cells were detectable in the dorsal motor vagal nucleus, tyrosine hydroxylase-, dopamine- and DOPA-immunoreactive perikarya were seen in the medial half of this nucleus, caudally the obex level. These results led us to conclude that these tyrosine hydroxylase-positive cells were effectively of dopaminergic nature and therefore that dopamine is a neurotransmitter contained in some neurons of the dorsal motor vagal nucleus. In the light of previous data showing colocalization of tyrosine hydroxylase and choline acetyltransferase in neurons of this portion of the nucleus, colocalization of dopamine with acetylcholine appears most likely. This might shed some light on the physiological consequences of dopamine action at target parasympathetic organs, such as the gastrointestinal tract.

Animals↗