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Biomedical subjects

D G Morgan

Publications and source records attributed to D G Morgan.

At least 109 records · Page 6Linked to original sources
PubMed1986

Dietary pyridoxine and the susceptibility to limbic motor seizures in rats.

Dietary pyridoxine (PN) deficiency in adult female rats produced a 32% decrease in hippocampal gamma-aminobutyric acid content, measured by a radioreceptor assay. No spontaneous seizures were observed in pyridoxine-deficient animals, but the seizure latency after a systemic kainic acid challenge decreased by 35%. The results suggest that latency is a useful measure of limbic seizure susceptibility; this susceptibility can be manipulated by diet; and in adults pyridoxine-dependent mechanisms normally participate in preventing, rather than initiating, limbic seizures.

Animals↗
PubMed1986

[3H]Fluphenazine binding to brain membranes: simultaneous measurement of D-1 and D-2 receptor sites.

[3H]Fluphenazine was used to label both D-1 and D-2 dopamine receptors in mouse striatal membranes. The D-1 and D-2 specific binding of [3H]fluphenazine was discriminated by the dopamine antagonists SCH-23390 (D-1 selective) and spiperone (D-2 selective). Saturation analyses of these two sites yielded a D-1 receptor density in mouse striatum of 1,400 fmol/mg of protein and a D-2 receptor density of 700 fmol/mg of protein. The affinity of [3H]fluphenazine for the D-2 site was slightly greater than for the D-1 site; the equilibrium dissociation constant (KD) was 0.7 versus 3.2 nM, respectively. Assay conditions are described that reduce nonspecific binding of [3H]fluphenazine to acceptable levels (35% of total binding at 1 nM [3H]fluphenazine). By comparison of displacement curves from a series of dopaminergic and nondopaminergic ligands, the pharmacological specificity of [3H]fluphenazine binding in mouse striatum was demonstrated to be dopaminergic. Only small amounts of dopamine-specific (apomorphine-sensitive) [3H]fluphenazine binding were found in other brain regions. However, chlorpromazine displaced considerable [3H]fluphenazine from all brain regions, including cerebellum, suggesting the presence of a [3H]fluphenazine binding site with a phenothiazine specificity.

Animals↗
PubMed1986

Binding specificity of the periplasmic oligopeptide-binding protein from Escherichia coli.

The structural properties required for the binding of peptide substrates to the Escherichia coli periplasmic protein involved in oligopeptide transport were surveyed by measuring the ability of different peptides to compete for binding in an equilibrium dialysis assay with the tripeptide Ala-Phe-[3H]Gly. The protein specifically bound oligopeptides and failed to bind amino acids or dipeptides. Acetylation of the peptide amino terminus of (Ala)3 severely impaired binding, whereas esterification of the carboxyl terminus significantly reduced but did not completely eliminate binding. Peptides composed of L-amino acids competed more effectively than did peptides containing D-residues or glycine. Experiments with a series of alanyl peptide homologs demonstrated a decrease in competitive ability with increasing chain length beyond tripeptide. Competition studies with tripeptide homologs indicated that a wide variety of amino acyl side chains were tolerated by the periplasmic protein, but side-chain composition did affect binding. Fluorescence emission data suggested that this periplasmic protein possesses more than one substrate-binding site capable of distinguishing peptides on the basis of amino acyl side chains.

Acetylation↗
PubMed1985

Purification and characterization of a periplasmic oligopeptide binding protein from Escherichia coli.

We have purified and characterized an oligopeptide binding protein released from the periplasm of Escherichia coli W by mild osmotic shock. The purified protein was greater than 97% homogeneous as determined by either sodium dodecyl sulfate-polyacrylamide gel electrophoresis (Mr = 60,000) or isoelectric focusing (pI = 5.95). The binding protein has a Stokes radius of 30 A and a sedimentation coefficient (s(0)20,w) of 4.6 S. Based on these hydrodynamic studies, the native protein has a molecular weight of 56,000. The tripeptide, Ala-Phe-[3H]Gly, which is transported via the shock-sensitive sensitive oligopeptide permease, binds to the purified protein in dilute solution with a Kd of 0.1 microM and a stoichiometry of approximately 1 to 1. Results from this study support the hypothesis that this periplasmic oligopeptide binding protein functions in the initial recognition of peptide substrates for the oligopeptide permease system.

Bacterial Proteins↗
PubMed1985

Senescent change in tissue weight and immunoreactive beta-endorphin, enkephalin, and vasopressin in eight regions of C57BL/6J mouse brain and pituitary.

Tissue weights and immunoreactive (IR) content and concentration of beta-endorphin, enkephalin, and vasopressin were assayed for senescent change in anterior pituitary, neurointermediate pituitary, hypothalamus, hippocampus, striatum, dorsolateral cortex, and pons-medulla, as well as residual brain samples remaining after the other dissections. Groups of naive male C57BL/6J mice, 8-12 months old and 28-32 months old, served as subjects. Old mice exhibited significant decline in anterior pituitary and hippocampus weight. Significant increases with age were found in pons-medulla weight. IR beta-endorphin content decreased in hypothalamus and neurointermediate pituitary. IR enkephalin decreased in striatum and dorsolateral cortex. IR vasopressin content increased in hypothalamus and residual brain. Assays were replicated in later experiments using additional subjects, for total Ns of up to 54 mice. Although significant overall results were always consistent in direction from replication to replication, the magnitude of age change exhibited considerable variability. Such results suggest that single experiments on age changes in neuropeptides, particularly those giving negative results, should be carefully scrutinized and replicated before their acceptance as evidence for a transmitter's stability or instability throughout the lifespan.

Aging↗
PubMed1985

Arterial medial necrosis and hemorrhage induced in rats by intravenous infusion of fenoldopam mesylate, a dopaminergic vasodilator.

Fenoldopam mesylate, a selective, postsynaptic, dopaminergic vasodilator, was administered to rats for assessment of its clinical, toxicologic, and pathologic effects. Groups of 8 male and 8 female rats received 5, 25, 50, or 100 micrograms/kg/min by intravenous infusion for 24 hours. Groups of 12 male and 12 female rats received 2, 8, 16, or 20 mg/kg/day by intravenous injection once daily for 12 days. Tissues were examined by light microscopy. Rats infused for 24-hours with 5-100 micrograms/kg/min of fenoldopam had lesions of renal and splanchnic arteries characterized by medial necrosis and hemorrhage. None were seen in control rats or those administered the compound by intravenous injection. Arteries with four to five layers of medial smooth-muscle cells were most severely and frequently affected. Lesions were particularly severe in interlobular pancreatic arteries and subserosal gastric arteries. They occurred first at 4 hours, were present at low incidence at 8 hours, were induced in unrestrained rats, and were not caused by the experimental procedures employed. The nature and disposition of this novel arterial lesion in the rat suggests that its pathogenesis may be related to the pharmacologic activity of fenoldopam mesylate at the dopamine receptor.

Animals↗
PubMed1984

Hyperprolactinemia fails to increase striatal dopamine receptors in male and female C57BL/6J mice.

Mice were administered prolactin by either homotypic anterior pituitary grafts (4 weeks) or subcutaneous osmotic pumps containing ovine prolactin (1 week). The pituitary grafts produced considerably greater elevations of circulating prolactin than the osmotic pumps. In neither case was any change detected in striatal [3H]spiperone binding to dopamine receptors. These results are discussed in the context of contradictory evidence obtained with rats.

Animals↗
PubMed1984

Serotonin-2 binding sites in human frontal cortex and hippocampus. Selective loss of S-2A sites with age.

Serotonin (S-2) binding sites were characterized in human frontal cortex and hippocampus throughout the lifespan. We found that the S-2 binding sites (labeled by [3H]spiperone and displaced by ketanserin) consisted of two subtypes (S-2A and S-2B) which are discriminated by competition with methysergide. The total S-2 sites in frontal cortex had a Bmax of 360 fmol/mg protein, which is approximately twice that of the hippocampus (160 fmol/mg protein). The density of total S-2 sites decreased with age in the frontal cortex of normal adults (17-100 years, n = 24). The receptor loss was primarily in the S-2A subclass. This loss of S-2A sites occurred primarily after 60 years and decreased to 50% of young adult values by the 10th decade. In the hippocampus, S-2A binding sites decreased with age, but no effects on total S-2 or S-2B sites were detected. A study of infant brains suggested the S-2A subtype in frontal cortex increases postnatally. However, in the infant hippocampus the S-2 binding approximated adult levels.

Adolescent↗
PubMed1984

Contamination of serotonin-2 binding sites by an alpha-1 adrenergic component in assays with (3H)spiperone.

(3H)Spiperone binds to two sites in mouse cortical membranes. These binding sites are discriminated by methysergide and prazosin, but not by butaclamol, lysergic acid diethylamide, or ketanserin. One of these sites is serotonergic in nature and is the authentic S-2 binding site. The other component is adrenergic and corresponds to the alpha-1 adrenoreceptor. This alpha-1 component may be present in other S-2 binding assays using (3H)spiperone, or (3H)ketanserin. No (3H)spiperone binding to dopaminergic D-2 sites was found in mouse cortex. Methods of avoiding alpha-1 contamination of S-2 binding assays are suggested.

Animals↗
PubMed1984

Chronic domperidone fails to increase striatal spiperone binding sites despite hyperprolactinemia: comparison with chronic haloperidol.

Mice were administered the dopaminergic antagonists haloperidol or domperidone in their drinking water for 2 or 21 days. Serum prolactin levels and striatal 3H-spiperone-binding sites (D-2 receptors) were compared to vehicle-treated controls. While only domperidone elevated serum prolactin levels, only haloperidol increased the density of striatal 3H-spiperone-binding site. The failure of domperidone, a potent D-2 receptor antagonist, to increase striatal receptor number is attributed to its poor penetration of the blood-brain barrier. These results indicate that peripheral effects of neuroleptic drugs, including prolactin elevation, are not sufficient for the development of dopaminergic supersensitivity in the central nervous system.

Animals↗
PubMed1981

Brain pyruvate dehydrogenase: phosphorylation and enzyme activity altered by a training experience.

The active portion of the alpha subunit of pyruvate dehydrogenase in rat frontal cortex was elevated after a training experience. No change in total pyruvate dehydrogenase activity was observed. The phosphorylation in vitro of pyruvate dehydrogenase (band F-2) was also elevated after training. Since activation of pyruvate dehydrogenase requires its dephosphorylation, the following sequence is proposed. Training alters frontal cortex and reduces the phosphate content of pyruvate dehydrogenase in vivo; this leads to enzyme activation; and an increase in back-titration of sites available for phosphorylation in vitro.

Animals↗
PubMed1981

Human brain protein phosphorylation in vitro: cyclic AMP stimulation of electrophoretically-separated substrates.

In vitro phosphorylation of electrophoretically-separated brain proteins was studied in human frontal cortex obtained 3-16 h post-mortem from 13 patients ages 3 days-82 years with extensive, mild or no neuropathological involvement. In 12 of the 13 cases, cyclic AMP increased incorporation of phosphate into acid-precipitable protein. Analysis of the autoradiographic profiles of separate proteins indicated that phosphorylation of the doublet of molecular weight 86-80,000 was stimulated by cyclic AMP in certain samples. This doublet corresponded to the cyclic AMP stimulated doublet from rat frontal cortex we have termed band D-1,2 (proteins Ia and Ib of Ueda and Greengard). Of special interest was the fact that, while co-migration was observed in the other phosphoprotein bands studied, band D-1,2 of humans consistently migrated slightly less than rat protein band D-1,2. This difference was not a function of post-mortem time, subcellular fraction or buffer used in the reaction phosphorylation assay. The use of post-mortem tissue was not a contributing factor as the retardation in band D-1,2 migration was still observed when post-mortem rat brain was used for comparison. In two human post-mortem samples, there was no measureable band D-1,2 phosphorylation even in the presence of cyclic AMP. This was the case in both homogenate and crude synaptosome/mitochondrial preparations. Band F-1 (mol. wt. = 47,000) was not observed in any of the human samples studied. This is consistent with prior studies in rat which show that band F-1 phosphorylation is not detected in post-mortem brain, Band F-2 (mol. wt. 41,000) recently identified as pyruvate dehydrogenase, was lightly phosphorylated under the reaction conditions used in this study.

Aged↗
PubMed1981

A membrane-impermeant, cleavable cross-linker. Dimers of human erythrocyte band 3 subunits cross-linked at the extracytoplasmic membrane face.

We have synthesized diisethionyl-3,3'-dithiobispropionimidate (DIDIT), a new membrane-impermeant, cleavable protein cross-linking reagent designed for probing protein organization at one face of a membrane. Rabbit muscle aldolase were reacted in solution with DIDIT and the products were electrophoresed in sodium dodecyl sulfate-polyacrylamide gels. When electrophoresed under nonreducing conditions, the gels contain bands corresponding to oligomers of aldolase, while pretreatment with dithiothreitol to cleave the cross-link prior to electrophoresis results in gels containing primarily the band corresponding to aldolase monomer. These experiments demonstrate that DIDIT is a cleavable protein cross-linker. Reaction of isolated human erythrocyte membranes with DIDIT leads to extensive cross-linking of spectrin, band 3, and band 6, and residual hemoglobin, consistent with results previously obtained with permeant cross-linkers. In contrast, when intact human erythrocytes are cross-linked with DIDIT, hemoglobin and the cytoplasmic face membrane proteins are not cross-linked, but band 3, which is accessible at the extracytoplasmic face of the membrane, is cross-linked to dimers.

Animals↗
PubMed1981

In vivo phosphorylation following [32P]orthophosphate injection into neostriatum or hippocampus: selective and rapid labeling of electrophoretically separated brain proteins.

Intracranial injections of [32P]orthophosphate readily label a number of brain phosphoproteins as resolved by polyacrylamide gel electrophoresis. The majority of these in vivo labeled phosphoproteins co-migrate with phosphoproteins that are labeled in vitro by incubation of brain membranes with [32P]ATP. Two of the major in vitro labeled phosphoproteins with apparent molecular weights of 47,000 (band F1) and 41,000 (band F2) are rapidly labeled in vivo. Since they are rapidly dephosphorylated in vitro, this suggests a high rate of phosphate turnover. The electrophoretic pattern of in vivo labeled phosphoproteins did not appear to be altered by the method of sacrifice (focused microwave irradiation, decapitation or liquid nitrogen immersion) or by the state of the animal at the time of labeling (awake or lightly anesthetized with pentobarbital). The reduction of phosphatase activity during tissue processing at 0 degree C may account for the similarities observed with different sacrifice methods. Removal of phospholipids or polynucleotides had little effect on the in vivo labeled 32P-containing bands. However, alkaline hydrolysis or protease treatment uniformly reduced the radioactivity in the labeled bands. These findings suggest that the 32P-containing bands consist of phosphoester linkages to serine or threonine residues. The present evidence emphasizes that previously characterized in vitro labeled brain phosphoproteins are, in fact, labeled in the awake, freely-moving animal.

Adenosine Triphosphate↗
PubMed1979

Incorporation of intrastriatally injected[3H]fucose into electrophoretically separated synaptosomal glycoproteins. I. Turnover and molecular weight estimations.

The radioactivity profiles of electrophoresed neostriatal P2 fraction glycoproteins were examined at a series of times (2.5, 3 and 4 h; 1, 5 and 10 days) following intracranial injections of [3H]fucose into the neostriatum. Ten major fucosylglycoprotein peaks were discerned in these profiles and certain aspects of their metabolism were characterized. The half-life of fucosylglycoproteins in the P2 fraction was estimated to be 9.7 days. The half-lives of the individual glycoprotein peaks ranged from 4.9 to 17.9 days. The apparent molecular weights of the glycoprotein peaks obtained by our procedures ranged from 32,000 to 180,000 daltons. One peak (peak VIII) incorporated radioactivity primarily at short intervals following the injection. The time course of [3H] fucose incorporation into this peak suggests involvement in the transport, activation and/or incorporation of fucose in brain. Since intracranial injections of [3H]fucose are incorporated into proteins in the cell body, synaptosomal fractions from caudate neurons alone are labeled by this technique. This may be useful in separating pre- and postsynaptic glycoprotein biochemistry. Finally, we tentatively propose that the glycoprotein peaks observed in neostriatum may be identical to previously isolated glycoproteins of known function or subcellular location.

Animals↗