Biomedical subjects
Publications and source records attributed to J Schmidt.
Parameters of dopamine turnover were measured after developing different forms of behaviour facilitation. The state of denervation supersensitivity is characterized biochemically by diminished dopamine turnover and enhanced receptor sensitivity whereas forms of innervation supersensitivity did not show such alterations.
Rats exposed to an intermittent long-term hypoxia in the early postnatal life show a marked hyperactivity in the open field test at day 15 and 38 of life. This effect disappears during maturity. The spontaneous nocturnal activity of adult animals oxygen deprived postnatally was found to be elevated. These results may be a possible consequence of the hypoxic injury during the growth spurt period of neuronal systems which play an important role in the control of locomotor behaviour. The method used and the results obtained seem to meet some requirements of an animal model for symptoms associated with the syndrome referred to as "Minimal Brain Dysfunction".
Exposure of rats to hypoxia results in a substantial decrease of dopamine release from striatum slices for several days. Nootropic drugs (piracetam, meclofenoxate hydrochloride, methylglucamine orotate, nicergoline) accelerate the restitution of posthypoxic release inhibition. In contrast, amphetamine is ineffective in this respect. The antihypoxic action of sedatives (diazepam, phenobarbital) prevents the decrease of dopamine release. Comparable results with free radical scavengers (cysteamine hydrochloride, sodium formiate, ouabain), ascorbic acid, natrii calcii edetas, selenium methionine and acetylsalicylic acid which protect dopamine release from hypoxically produced changes agree with and support the hypothesis of hypoxia induced free radical generation followed by phospholipid peroxidation altering particularly neuronal membrane function. On that account, dopamine release from rat striatum slices reflects not only the vulnerability of neuronal membrane function by hypoxia but also the preventive, protective and restitutive effects of antihypoxic drugs of different type and is able to contribute to discriminating drug investigation.
The relations between a perinatal exposure schedule and the outcome of dopamine (DA) release from striatum slices on the one hand and behaviour on the other, were studied in order to identify the vulnerable period. A mild chronic postnatal hypoxia (2nd-10th day of life) induces long-term effects on the DA release rate (FER) from striatum slices. A 10 h daily exposure to hypoxia for 5 days (2nd-6th day of life) induces the same long-term effects. Adult rats exposed to hypoxia (pO2 8.6 kPa) for 16 h respond with a drastic decrease of FER of DA. Exposure to early postnatal hypoxia prevented this drastic decrease of FER of DA when adult rats were again exposed to hypoxia. 3 ten h periods of hypoxia within the first 10 days of life were enough to the long lasting effects. Prenatal hypoxia as well as hypoxia during the adolescent period did not show any long-term effects on the dopaminergic system. Alterations in behaviour (decreased learning capacity as well as decreased reaction to stimuli after prolonged stress in adulthood) were found after 3-9 days of hypoxia between the 2nd and 10th day of life.
The effectiveness of drugs affecting aminergic triggered motor behaviour was investigated in adult rats oxygen deprived in the early postnatal life. Offsprings were exposed to hypobaric hypoxia (pO2 = 11.6 kPa) from the 2nd till the 10th postnatal day (10 h per day). Hypoxia exposed animals displayed a significant decrease of the locomotor apomorphine effect as well as of the amphetamine induced stereotyped behaviour. On the other hand, apomorphine stereotypies and the locomotor amphetamine effect proved to be unchanged after postnatal oxygen deprivation. The motility levels achieved after giving lysergic acid diethylamide 30 min before apomorphine administration were found to be considerably higher than after apomorphine alone and similar for both controls and hypoxia exposed animals. After unilateral dopamine injections into the nucleus accumbens no different rotational behaviour was detectable. The deviations in some of their responses to drugs affecting aminergically triggered behaviour in adult rats suggested a decreased behavioural effectiveness of dopamine agonists due to early postnatal hypoxia.
Piracetam (PIR) and dimethylaminoethanol (DMAE) were tested with respect to their ability to prevent changes of the locomotor behaviour and of the dopamine release from striatum slices after an intermittent postnatal hypoxia in the rat (2nd-10th day of postnatal life). The drugs given to pregnant rats (from 12th day of gestation) and continued after delivery to the newborns (till 10th day of postnatal life) displayed antihypoxic effects. Perinatal DMAE- but not PIR-treatment diminishes the consequences of postnatal oxygen deprivation regarding the motor activity as well as the dopamine release of adult animals. Hypoxia-caused open field hyperactivity of developing rats was found to be reduced to control values in PIR-treated rats. Only minor effects were seen after DMAE-treatment, but a marked depressant own effect on the explorative activity was detectable.
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We have studied putative nicotinic acetylcholine receptors in the optic lobe of the newborn chick, using 125I-labeled alpha-bungarotoxin, a specific blocker of acetylcholine receptors in the neuromuscular junction, and [3H]acetylcholine, a ligand which in the presence of atropine selectively labels binding sites of nicotinic character in rat brain cortex (Schwartz et al., 1982). [3H]Acetylcholine binds reversibly to a single class of high affinity binding sites (KD = 2.2 X 10(-8) M) which occur at a tissue concentration of 5.7 pmol/g. A large fraction (approximately 60%) of these binding sites is solubilized by Triton X-100, sodium cholate, or the zwitterionic detergent 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonate. Solubilization increases the affinity for acetylcholine and several nicotinic drugs from 1.5- to 7-fold. The acetylcholine-binding macromolecule resembles the receptor for alpha-bungarotoxin present in the same tissue with respect to subcellular distribution, hydrodynamic properties, lectin binding, and agonist affinity rank order. It differs from the toxin receptor in affinity for nicotinic antagonists, sensitivity to thermal inactivation, and regional distribution. The solubilized [3H]acetylcholine binding activity is separated from the toxin receptor by incubation with agarose-linked acetylcholine, by affinity chromatography on immobilized Naja naja siamensis alpha-toxin, and by precipitation with a monoclonal antibody to chick optic lobe toxin receptor.
Afterload reduction is an accepted therapeutic principle in the management of acute aortic (Ai) and mitral insufficiency (Mi). The question whether acute and chronic converting-enzyme inhibition by captopril has a beneficial hemodynamic effect in chronic Ai and Mi has been investigated in 17 patients with Ai and 10 with Mi. Ejection and regurgitation fraction (RF) were measured by radionuclide ventriculography (RNV) before, after 25 mg captopril and after 3-5 months of long-term treatment. The humoral response of the renin-angiotensin system (RAS) was quantified by analysis of angiotensin I and II. Captopril lowered under acute and chronic treatment RF in Ai and Mi by 32%. Angiotensin II levels decreased by the same order of magnitude. Acute and chronic vasodilation was followed by a distinct but well tolerated fall in blood pressure, especially in patients with Mi. These favourable hemodynamic effects of captopril make this therapy an adjunct but not an alternative to valve replacement.
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The antitumor agent, cis-dichlorodiammine Pt(II), is cytotoxic to Ehrlich cells in culture. These cells contain a substantial amount of metallothionein in the absence of inducers of the protein. At concentrations of drug which cause 60% inhibition of cell proliferation, most of the platinum is found in the cytosol. Of this about 30% is bound in the metallothionein fraction. Isolated rat liver metallothionein reacts slowly with hydrolyzed cis-dichlorodiammine Pt(II). Thus, metallothionein is a major cellular site of binding of the platinum complex at concentrations which inhibit tumor growth.
The entire M RNA segment of the phlebovirus Rift Valley fever virus (RVFV) has been molecularly cloned and the complete nucleotide sequence determined. The RNA is 3884 nucleotides in length, corresponding to a molecular weight of 1.38 X 10(6), having a base composition of 27.3% A, 25.4% G, 27.2% U, and 20.1% C. Sequences present at the 3' and 5' termini of the molecule are largely complementary for some 51 residues and can form a stable duplex structure when the potential secondary structure of the entire molecule is considered. A single major open reading frame, capable of encoding 1206 amino acids (131,845 Da), was found in the viral-complementary sequence ("positive" polarity). Amino-terminal amino acid sequencing of the purified viral glycoproteins G1 and G2 allowed for the positioning of the coding sequences for these polypeptides within this major open reading frame in the following orientation with respect to the genomic M RNA: 3'-G2-G1-5'. From the predicted amino acid composition of the two mature viral glycoproteins, both were found to have a high cysteine content (G2, 6%; G1, 5%). Sequences within the open reading frame capable of encoding up to 23,000 Da of polypeptide were found in addition to those required for the viral glycoproteins. The potential contribution of these sequences to the coding capacity of the M RNA, viral protein processing, and intracellular protein distribution is discussed.
We have employed a new pseudosubstrate, beta-(2-furyl)propionyl coenzyme A (FPCoA), to study the functional properties of two enzymes, fatty acyl-CoA dehydrogenase from porcine liver and fatty acyl-CoA oxidase from Candida tropicalis, involved in the oxidation of fatty acids. Previous studies from our laboratory have shown that the dehydrogenase exhibits oxidase activity at the rate of dissociation of the product charge-transfer complex. This raises the question of the difference in functionality between these two flavoproteins. To investigate these differences, we have compared the pH dependence of product formation, the isotope effects using tetradeuterio-FPCoA, and the spectral properties and chemical reactivity of the product charge-transfer complexes formed with the two enzymes. The pH dependencies of the reaction of FPCoA with electron-transfer flavoprotein (ETF) for the dehydrogenase and of the reaction of FPCoA with O2 for the oxidase are quite similar. Both reactions proceed more rapidly at basic pH values while substrate binds more tightly at acidic pH values. These data for both enzymes are consistent with a mechanism in which enzyme is involved in protonation of the carbonyl group of substrate followed by base-catalyzed removal of the C-2 proton from substrate. The C-2 anion of substrate may then serve as the active species in reduction of enzyme-bound flavin. The deuterium isotope effects for both enzyme systems are primary across the entire pH range, assuring that the chemically important step of substrate oxidation is rate limiting in these steady-state kinetic experiments. The two enzymes differ in the chemical reactivity of their product charge-transfer complexes.(ABSTRACT TRUNCATED AT 250 WORDS)
A simple procedure is described to separate reticulocytes of different maturity in high yield. It is shown that exhaustion of supply of mitochondria susceptible to degradation by the lipoxygenase-ATP-dependent proteolysis system limits the extent of breakdown of mitochondria during in vitro maturation. The susceptibility of mitochondria depends on the maturity of the reticulocytes. Incubation in the presence of calcium ions and calcium ionophore leads to full susceptibility of mitochondria in immature reticulocytes but has no effect on those in mature reticulocytes which are already fully susceptible to degradation. Conditions which lead to rapid degradation of mitochondria do not affect the behaviour of the reticulocyte count. There appears to be no obligatory connection between the breakdown of mitochondria and of ribosomes.
The nod C gene of Rhizobium meliloti encodes a protein of mol. wt. 44 000 which is highly conserved in at least three Rhizobium species. In order to overproduce this protein, a gene fusion of lambda cI repressor sequences to a large fragment of nod C was constructed. The fusion was placed under control of the tac promoter on plasmid pEA305 to yield pJS1035. IPTG-induced Escherichia coli cells harbouring pJS1035 accumulated the cI-nod C hybrid protein up to 19% of total cellular protein. The synthesis of the hybrid protein drastically inhibits the growth rate of the bacterium. The fusion protein was purified by gel and hydroxyapatite chromatography in the presence of SDS. Antibodies raised against the purified fusion protein precipitated the mol. wt. 44 000 nod C proteins of R. meliloti and of the broad-host range Rhizobium strain NGR234, which were both expressed in E. coli mini-cells. The hybrid protein is associated with the outer membrane of E. coli cells, and the cI-nod C fusion protein appears to be an integral membrane protein. Nodulation of alfalfa by R. meliloti and of clover by R. trifolii was markedly inhibited (approximately 50%) by the addition of antibodies against the hybrid protein to plant growth medium and inoculum.
A conventional in vitro test assay was used to determine maximal bactericidal capabilities of human granulocytes. By means of a mathematical model the maximal phagocytosis and killing activity could be calculated for S. aureus and P. aeruginosa serving as test organisms. The evaluation allowed moreover the determination of the optimal bacterial load and also of critical bacterial concentrations leading to a complete depression of observable granulocyte killing functions. In contrast to other studies frozen suspensions of bacteria were used allowing the employment of identical microorganisms within a complete series of experiments. On average one granulocyte was found to ingest a maximum of 17 CFU of S. aureus with 9 CFU killed under optimal ratios of bacteria per granulocyte. For P. aeruginosa the granulocyte function reached peak values of 96 CFU ingested and 62 CFU killed per one granulocyte. The new assay might provide a highly reproducible method for clinical assessment of granulocyte dysfunctions in various diseases.
The expression of RNA tumor virus was studied in BDF 1 mice after leukemogenic treatment with a single dose of methylnitrosourea (MNU) and in leukemic thymuses by a cell ELISA using antibodies against the viral glycoprotein gp 70 and by co-culture for the detection of eco- and xenotropic virus. The majority of the thymomas were positive for gp 70; ecotropic, but not xenotropic infectious virus could be detected in some of them. Early after MNU application the thymus and the bone marrow were positive for gp 70 in some animals. Later, after a phase with positive results with spleen cells, the bone marrow and the spleen were negative again. Only the thymus of some mice were positive during the last weeks before the first leukemias appeared.
Sensitive cocultivation techniques were applied to study the radiation-induced activation of endogenous retroviral genomes in different mouse strains by the alpha-emitting radionuclide 224Radium. Activated infectious C-type retroviruses were detected in spleen, bone marrow and bone tissues of C57BL/6-, BALB/c- and NMRI mice. The titres of high-dose-irradiated animals were higher than those found in low-dose-irradiated animals. Infectious retrovirus could be detected with a dose of 13.2 rad (maximum dose rate 0.9 rad/day) in the skeleton, and a dose of 4.2 rad (maximum dose rate 0.3 rad/day) in the spleen. The virus activation pattern was different in the three mouse strains. These data indicate that activation of endogenous retroviral genomes by alpha-irradiation shows a dose-effect relationship and a dependence on the genetic background of the mouse.