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Biomedical subjects

J D Robinson

Publications and source records attributed to J D Robinson.

At least 145 records · Page 8Linked to original sources
PubMed1977

Amniotic fluid androgens and estrogens in midgestation.

Amniotic fluid androgen and estrogen levels associated with 48 male and 72 female fetuses between 14 and 20 weeks of gestation were measured. Amniotic fluid testosterone levels were significantly higher (P less than 0.001) in the male (224 +/- 11 pg/ml) than the female fetuses (39 +/- 2 pg/ml) with no overlap of values. Amniotic fluid androstenedione concentrations were also significantly higher (P less than 0.001) with male (1024 +/- 53 pg/ml) than female fetuses (668 +/- 39 pg/ml), but there was overlap. There was no difference between anmiotic fluid dehydroepiandrosterone levels for the two sexes. Estrone concentrations were slightly but not significantly higher with the male (353 +/- 33 pg/ml) than with female fetuses (331 +/- 28 pg/ml), while estradiol concentrations were significantly higher (P =0.002) with the female (96 +/- 8 pg/ml) than male (64 +/- 4 pg/ml) fetuses. It is interpreted that the higher amniotic fluid testosterone and androstenedione levels for the male fetuses reflect fetal testicular secretion. The significantly higher estradiol concentrations for the female fetuses may reflect early ovarian secretion.

Amniotic Fluid↗
PubMed1977

Radioimmunoassay for flupenthixol in plasma.

We describe a radioimmunoassay for the neuroleptic drug flupenthixol, suitable for routine monitoring of its concentration in blood. Antibodies for the assay were raised in a sheep against a 7-carboxyflupenthixol/ovalbumin conjugate. The resulting assay, with [3H] flupenthixol as the label, is capable of detecting 2.0 microgram of flupenthixol per liter, in a 100-microliter plasma sample. The antiserum shows no cross reactivity with tricyclic drugs and low interference from the major metabolites of flupenthixol. Concentrations in plasma after a single oral dose of flupenthixol have been followed in one volunteer. Peak values were reached after 3 h. Determinations of flupenthixol after fortnightly intramuscular depot injections of the sustained-release preparation, flupenthixol decanoate, showed the extent of fluctuations during this period.

Cross Reactions↗
PubMed1976

The (Na + K+)-dependent ATPase. Mode of inhibition of ADP/ATP exchange activity by MgC12.

Na+-dependent ADP/ATP exchange activity, of a (Na++K+)-dependent ATPase preparation from eel electric organ, was measured in terms of the incorporation of 14C into ATP during incubations with labeled ATP and [14C]ADP. Estimates of initial rates of exchange were possible by keeping changes in nucleotide concentrations, from both exchange and extraneous hydrolytic processes, to less than 10%. Under these conditions, increases in MgC12 concentration, from 0.2 to 3 mM, generally inhibited this exchange activity. The concentrations of free Mg2+, Mg-ATP, and Mg-adp present, with a range of MgC12, ATP, and ADP concentrations, were calculated from measured dissociation constants. Inhibition was associated with Mg-ATP as well as with Mg2+, at concentrations from 0.4 to 1 mM (Mg-ADP, in the same concentration range, probably inhibited also). The affinity of the enzyme for these inhibitors is in fair correspondence with demonstrated affinties for Mg2+, Mg-atp, and Mg-ADP at low affinity substrate sites, measured kinetically. These observations are considered in terms of a dimeric enzyme with high and low affinity substrates sites: ADP/ATP exchange being catalyzed at the high affinity sites, with inhibition occurring through occupancy by Mg2+, Mg-ATP, or Mg-ADP, of the low affinity sites, thereby pulling the reaction process away from those steps involved in exchange.

Adenosine Diphosphate↗
PubMed1976

Uptake and release of calcium by rat brain synaptosomes.

Rat brain synaptosomes, prepared by discontinuous Ficoll density gradient centrifugation, accumulated 45Ca during brief incubations in modified Krebs-Ringer media. Uptake of 45Ca was increased by 5 mM glutamate and 50 mM KCl, conditions that depolarize nerve cells; uptake of 22Na was also increased by these agents. With 0.2 mM diphenylhydantoin, the increased 45Ca uptake due to KCl was diminished, whereas that due to glutamate was less affected; conversely, with 0.15 muM tetrodotoxin the increased 45Ca uptake due to glutamate was diminished, whereas that due to KCl was less affected. Both diphenylhydantoin and tetrodotoxin diminished the augmented uptake of 22Na due to KCl and glutamate; thus the increased uptake of 45Ca under depolarizing conditions may be dissociated from the increased influx of sodium. Ruthenium red decreased the uptake of 45Ca under all conditions, as did procaine and the lanthanide Pr3+. Neither 5 mM glutamate nor 50 mM KCl increased 45Ca uptake by brain mitochondria under comparable experimental conditions, whereas ATP increased the uptake by mitochondria but not that by these synaptosomes. Altering the sodium gradient by equimolar substitution of lithium or choline for sodium in the medium increased 45Ca uptake, whereas 22Na uptake was decreased. Inhibiting the sodium pump by ouabain or strophanthidin also increased 45Ca uptake, and increased 22Na uptake as well. The increased uptake of 45Ca induced by ouabain was inhibited by diphenylhydantoin and tetrodotoxin. Measurements of the total calcium content showed that conditions producing an increased uptake of 45Ca also produced a net uptake of calcium, rather than merely accelerating a 45Ca-40Ca exchange. Experiments measuring the loss of previously accumulated 45Ca showed that directly decreasing the sodium gradient or inhibiting the sodium pump slowed the loss of 45Ca. These data are considered in terms of calcium influx through 'leak' pathways and gated channels (sensitive to membrane depolarization) and of net efflux dependent on a coupled sodium-calcium exchange mechanism.

Acetylcholinesterase↗
PubMed1976

Substrate sites for the (Na+ + K+)-dependent ATPase.

Kinetic studies on a rat brain (Na+ + K+)-dependent ATPase (EC 3.6.1.3) preparation demonstrated high-affinity sites for ATP, with a Km near 1 mum, and low affinity sites for ATP, with a Km near 0.5 mM. In addition, the dissociation constant for ATP at the low affinity sites was approached through the ability of ATP to modify the rate of photo-oxidation of the enzyme in the presence of methylene blue; a value of 0.4 mM was obtained. The temperature dependence of the Km values in these two concentration ranges also differed markedly, and the estimated entropy of binding was +27 cal/degree per mol at the high affinity sites, whereas it was -20 cal/degree per mol at the low affinity sites. Moreover, the relative affinities of various congeners of ATP as of the K+ -dependent phosphatase reaction of the enzyme indicated an interaction at the low-affinity sites for ATP: ATP, ADP, CTP, and the [beta-gamma] -imido analog of ATP all competed with Ki values near those for the ATPase reaction at the low affinity sites. Conversely, the Km for nitrophenyl phosphate as a substrate for the phosphatase reaction was near its Ki as a competitor at the low-affinity sites of the ATPase reaction. These observations are incorporated into a reaction scheme with two classes of substrate sites on a dimeric enzyme, manifesting idverse enzymatic and transport characteristics.

Adenine Nucleotides↗
PubMed1976

Amniotic fluid testosterone levels in midpregnancy.

Amniotic fluid testosterone levels were measured on specimens obtained between 12 and 25 weeks' gestation from 58 male-fetus and 77 female-fetus pregnancies. For the male fetuses the mean +/- SE amniotic fluid testosterone level of 223 +/- 10 pg/ml was significantly higher (P less than 0.001) than the concentration found for the female fetuses (40 +/- 2 pg/ml). The ranges were 104-424 and 18-82 pg/ml, respectively, for the same fetuses. In the male fetuses the highest mean level was found during the 17th gestational week, but the mean level observed during any 1 week was not significantly differenf from any other week for both sexes. These data are consistent with the concept that amniotic fluid testosterone levels may be a rapid and effective method for establishing fetal sex in utero during midgestation.

Amniotic Fluid↗
PubMed1975

Mechanisms by which Li+ stimulates the (Na+ and K+)-dependent ATPase.

The addition of LiCl stimulated the (Na+ + K+)-dependent ATPase activity of a rat brain enzyme preparation. Stimulation was greatest in high Na+/low K+ media and at low Mg-ATP concentrations. Apparent affinities for Li+ were estimated at the alpha-sites (moderate-affinity sites for K+ demonstrable in terms of activation of the associated K+-dependent phosphatase reaction), at the beta-sites (high-affinity sites for K+ demonstrable in terms of activation of the overall ATPase reaction), and at the Na+ sites for activation. The relative efficacy of Li+ was estimated in terms of the apparent maximal velocity of the phosphatase and ATPase reactions when Li+ was substituted for K+, and also in terms of the relative effect of Li+ on the apparent Km for Mg-ATP. With these data, and previously determined values for the apparent affinities of K+ and Na+ at these same sites, quantitative kinetic models for the stimulation were examined. A composite model is required in which Li+ stimulates by relieving inhibition due to K+ and Na+ (i) by competing with K+ for the alpha-sites on the enzyme through which K+ decreases the apparent affinity for Mg-ATP and (ii) by competing with Na+ at low-affinity inhibitory sites, which may represent the external sites at which Na+ is discharged by the membrane Na+/K pump that this enzyme represents. Both these sites of action for Li+ would thus lie, in vivo, on the cell exterior.

Adenosine Triphosphatases↗
PubMed1975

Interactions between K+ and ATP binding to the (Na+ + K+)-dependent ATPase.

K+ appears to decrease the affinity of the (Na+ + K+)-dependent ATPase (ATP phosphohydrolase, EC 3.6.1.3) for its substrate, Mg2+ - ATP, and Mg2+ - ATP, in turn, appears to decrease the affinity of the enzyme for K+. These antagonisms have been investigated in terms of a quantitative model defining the magnitude of the effects as well as identifying the class of K+ sites on the enzyme involved. K+ increased the apparent Km for Mg2+ - ATP, an effect that was antagonized competitively by Na+. The data can be fitted to a model in which Mg2+ - ATP binding is prevented by occupancy of alpha-sites on the enzyme by K+ (i.e. sites of moderate affinity for K+ accessible on the "free" non-phosphorylated enzyme, in situ on the external membrane surface). By contrast, occupancy of these alpha-sites by Na+ has no effect on Mg2+ - ATP binding to the enzyme. On the other hand, Mg2+ - ATP decreased the apparent affinity of the enzyme for K+ at the alpha-sites, in terms of (i) the KD for K+ measured by K+-accelerated inactivation of the enzyme by F-, and (ii) the concentration of K+ for half-maximal activation of the K+-dependent phosphatase reaction (which reflects the terminal hydrolytic steps of the overall ATPase reaction). These data fit the same quantitative model. Although this formulation does not support schemes in which ATP binding effects the release of transported K+ from discharge sites, it is consistent with observations that K+ can inhibit the enzyme at low substrate concentrations, and that Li+, which has poor efficacy when occupying these alpha-sites, can stimulate enzymatic activity at high K+ concentrations by displacing the inhibitory K+.

Adenosine Triphosphatases↗
PubMed1975

Functionally distinct classes of K+ sites on the (Na+ + K+)-dependent ATPase.

K+ interactions with a rat brain (Na+ + K+)-dependent ATPase and the associated K+-dependent nitrophenyl phosphatase activity were examined. Classes of sites for K+ were distinguished, initially, on the basis of affinity estimated by kinetic analysis in terms of KO.5 (the concentration for half-maximal activation), and by K+-accelerated enzyme inactivation by F-minus, which permits evaluation of a dissociation constant for K+, KD. Moderate-affinity sites ("alpha sites"), with a KD near 1 mM, were demonstrable for the phosphatase activity and for the "free" enzyme. High-affinity sites ("beta sites"), with a KD near 0.1 mM, were seen for the overall ATPase activity and under conditions in which enzyme phosphorylation by substrate also occurs. Further differentiation between alpha and beta sites was made in terms of (i) the characteristic changes in affinity with pH, and (ii) the efficacy of Li+ relative to K+, Rb+, Cs+, and Tl+ at these two classes of sites. Low-affinity sites ("gamma sites") through which K+ inhibits enzymatic activity were also detectable, with a KD around 140 mM. These data are incorporated into a model for the reaction sequence to accommodate both transport processes and certain K+/ATP antagonisms.

Adenosine Triphosphatases↗
PubMed1975

Specific modifications of the Na+,K+-dependent adenosine triphosphatase by dimethyl sulfoxide.

DMSO inhibits the Na+, K+-ATPase, but stimulates the associated K+-phosphatase activity. For the ATPase, DMSO acts as an uncompetitive inhibitor toward both ATP and Na+, whereas it increases the K0.5 for K+. From measurements of the dissociation constant (Km) of these ions in the ligand states that correspond to the ATPase reaction, it can be shown that DMSO has little effect on the affinity for Na+, but decreases the affinity for K+ of the enzyme-phosphate intermediate (the form that has the highest affinity for K+). By contrast, DMSO decreases the Km for the phosphatase substrate (nitrophenyl phosphate) without affecting the Vmax. Moreover, DMSO decreases the K0.5 for K+ and also the Kd for K+ in the ligand states that correspond to the phosphatase reaction (which have only a moderate affinity for K+, since the acyl phosphate intermediate is absent in this pathway). These data may be incorporated into a reaction mechanism for the Na+, K+-ATPase. Initially the enzyme is phosphorylated to form an acyl phosphate intermediate, in steps that require Na+ and Mg-2+. At this stage the affinity of K+ is markedly increased (from the moderate affinity seen in the "free" enzyme and the phosphatase reaction). When K+ is bound, the phosphate group is transferred to the hydrolytic site where P-i is ultimately released. DMSO acts at the point at which the acyl phosphate group or the phosphatase substrate enters the hydrolytic site, inhibiting one and facilitating the other. At this stage the affinity for K+ is also changing, and DMSO apparently selects an enyme conformation of intermediate affinity. Ion transport may occur by a gate mechanism in an overall system that operates on a half-of-the-sites active enzyme pattern in which ATP hydrolysis may alternate between the dimeric subunits of the enzyme.

Adenosine Triphosphatases↗
PubMed1975

Pharmacokinetics of a single dose of phenytoin in man measured by radioimmunoassay.

1. Serum concentrations of phenytoin were studied by radioimmunoassay in five normal volunteers following a single oral dose of phenytoin sodium (100 mg). 2. Two distinct peaks were found at 2.5-3.5 and 10-12 h after ingestion. Maximum serum concentrations ranged from 1.56 mug/ml. The mean plasma half-life of the drug under these conditions was 9.81 +/- 0.66 (s.e. mean) hours.

Administration, Oral↗
PubMed1975

Development of a radioimmunoassay for etorphine (preliminary communication).

A radioimmunoassay for etorphine has been developed using antibodies raised in a rabbit against an etorphine-3-hemisuccinate-bovine serum albumin conjugate. Using tritiated etorphine as the label, the antiserum could be used at a final dilution of 1:4 and had an avidity of 1.8 times 10-9 L.M.-1. The sensitivity of the assay was 0.88 plus or minus 0.04 ng etorphine per ml of serum and there was no cross-reactivity with any of the six opiate alkaloids examined over the range 0-100 ng/ml.

Animals↗