The relationship between 13, 14-dihydro-15-keto PGF2α and LH secretion in bulls

Half-life ($\text{t}\text{1}\text{2}$), volume of distribution (Vd)_and total body clearance (TBC) of 13, 14-dihydro-15-keto PGF_2α (PGFM) were measured in order to determine optimal sampling frequency for accurate measurement of PGFM. Three yearling Holstein bulls (349.2 ± 6.7 kg) and 3 yearling Holstein steers (346.7 ± 7.0 kg) were utilized in a 3 × 3 Latin square design. Animals were given 0, 25 or 50 μg PGF_2α I.V.; blood samples collected every 2 min and plasma PGFM determined. The $\text{t}\text{1}\text{2}$, Vd and TBC of PGFM were 2.3 ± .2 min, 43.3 ± 3.3 liters and 13.7 ± 1.9 liters/min, respectively and were similar for 25 and 50 μg doses. To determine the relationship between endogenous PGFM and LH secretion in bulls, blood samples were collected every 2 min for 12 h in 4 yearling Angus bulls (489.1 ± 11.6 kg). All animals elicited at least one LH surge and PGFM concentrations were measured in samples coincident with the LH surge. Mean plasma PGFM concentrations were greater prior to the LH surge than during the LH surge. In addition, mean plasma PGFM concentration and frequency of PGFM peaks appeared to increase prior to the LH surge suggesting an association between PGFM and pulsatile LH secretion in the bull.     

Elevation of plasma tyrosine after a single oral dose of L-tyrosine.

Plasma tyrosine concentrations in twelve normal, fasting human subjects were significantly elevated 2–8 hours after they ingested 100 mg/kg or 150 mg/kg tyrosine. Mean plasma tyrosine levels were maximal after 2 hours, rising from $\text{69 ± 3.9}\text{to}\text{154 ± 9.5}\text{nmols/ml}\text{(}\text{X}\text{±}\text{SEM}\text{)}$ after the 100 mg/kg dose and to 203 ± 31.5 nmols/ml after the 150 mg/kg dose (p ≤ 0.001 for both doses). The mean tyrosine ratio (defined as the ratio of plasma tyrosine concentration to the sum of the concentrations of six other neutral amino acids that compete for the same blood-brain barrier uptake system) increased from $\text{0.10 ± 0.02}\text{to}\text{0.28 ± 0.04 (}\text{X}\text{±}\text{SEM}\text{)}$ 2 hours after the 100 mg/kg dose (p ≤ 0.001) and to 0.35 ± 0.05 2 hours after the 150 mg/kg dose (p ≤ 0.005). No side effects of orally-administered L-tyrosine were noted.     

The effects of prostacyclin (PGI2) and 6-keto-PGF1α on bovine plasma progesterone and LH concentrations

Injections of 1 mg PGI₂ directly into the bovine corpus luteum significantly increased peripheral plasma progesterone concentrations within 5 min. Concentrations were higher in the PGI₂-treated heifers than in saline-injected controls between 5 and 150 min and at 3.5, 4, 5, and 7 h post-treatment. Levels tended to remain elevated through 14 h. Saline and 6-keto-PGF_1α were without effect on plasma progesterone levels. The luteotrophic effect of PGI₂ was not due to alterations in circulating LH concentrations. An $\text{in vitro}$ experiment assessed the effects of either PGI₂ alone or in combination with LH on progesterone production by dispersed luteal cells. Progesterone accumulation over 2 h for control, 5 ng LH, 1 μg PGI₂, 10 μg PGI₂, and 10 μg PGI₂ plus 5 ng LH averaged 99 ± 42, 353 ± 70, 152 ± 35, 252 ± 45, and 287 ± 66 ng/ml (n=4), respectively. Thus PGI₂ has luteotrophic effects on the bovine CL both $\text{in vivo}$ and $\text{in vitro}$.     

Elimination kinetics and protein binding of theophylline in the rabbit

The detailed elimination kinetics of theophylline were studied in 27 rabbits. Each received a 10 mg/kg intravenous bolus of aminophylline. The theophylline half-life ($\text{T}\text{1}\text{2}$) was 3.8 ± 0.63 hr. The apparent volume of distribution (V_D) and total body clearance (TBC) for theophylline were 439 ± 60 ml/kg and 81.0 ± 17.3 ml/kg·hr respectively. Theophylline protein binding was determined in 10 animals. The mean bound fraction was 74.3 ± 3.9% (range, 68.3–78.0%); the fraction bound was concentration indifferent over a serum concentration range of 5–20 μgm/ml.     

Potentiation of bradykinin-induced vascular permeability increase by prostaglandin E2 and arachidonic acid in rabbit skin

The activity of prostaglandins (PG) in producing vascular permeability was quantitated by dye extraction method in skin of anaesthetized rabbits. PGE₁ and PGE₂ (0.01–10 μg) produced increase in vascular permeability. Activity was approximately equal to that of histamine (Hist) and $\text{1}\text{20}$ of that of bradykinin (BK) on a weight basis. The activity of PGF_1α and PGF_2α was only $\text{1}\text{20}$ of that of PGE₁ or PGE₂.In spite of the relatively low potency of PGE₁ and PGE₂ in the rabbit, near threshold doses (0.1 or 1 μg) of PGE₂ could potentiate permeability responses to bradykinin (0.1 μg) by 10 or 100-fold, respectively. Equivalent doses (0.1 or 1 μg) of histamine could not potentiate the bradykinin responses. Arachidonic acid (AA) at 1 μg, produced a 10-fold potentiation in the permeability response to bradykinin (0.1 μg). Pretreatment of the rabbits with indomethacin (20 mg/kg, i.p.) reduced the responses of BK (0.1 μg) + AA (1 μg) down to a similar magnitude of those seen with bradykinin alone. However, indomethacin did not block responses to either, BK alone, BK + PGE₂, or BK + Hist. Various doses (1, 10, 100 and 300 μg) of arachidonic acid alone also produced increase in cutaneous vascular permeability, although its potency was only $\text{1}\text{3}$–$\text{1}\text{8}$ of that of PGE₂. This activity of arachidonic acid was attributed in part to its bioconversion to PGE₂, since its activity was significantly reduced by the prostaglandin antagonist, diphloretin phosphate (DPP) (60 mg/kg, i.v.) and by indomethacin (20 mg/kg, i.p.), which blocks conversion of arachidonic acid to prostaglandins. Arachidonic acid may owe some of its permeability increaseing effects to histamine release, since its effects were also reduced by the antihistamine, pyrilamine (2.5 mg/kg, i.v.).     

Refinement of the coomassie blue method of protein quantitation. A simple and linear spectrophotometric assay for less than or equal to 0.5 to 50 microgram of protein

The Coomassie brilliant blue G assay for proteins described by Bradford (1976) (Anal. Biochem.72, 248) was reexamined. It was found that the extinction coefficient of the dye-protein complex solution remained constant over the protein concentration range of 0.8 to 10 μg/ml of solution. This unchanging extinction coefficient, $\text{A}{}_{595}\text{=}\text{0.60 ± 0.01}\text{10}\text{μ}\text{g}$ of protein/ml of solution, enhances both the sensitivity and versatility of the assay. Selection of a volume of dye-reagent (0.5 to 5.0 ml) which dilutes the protein sample to a final concentration of 0.8 to 10 μg/ml permits the application of Beer's Law for accurate determinations of ≤0.5 to 50 μg of protein. A combination of Bradford's study and the present one indicates that most common laboratory reagents and chemicals exert little or no influence on the A₅₉₅ of the dye-reagent.     

Tryptophan uptake by Mycobacterium tuberculosis H37Rv--inhibition by isoniazid.

The effect of various sub-inhibitory concentrations of isoniazid on tryptophan uptake by $\text{Mycobacterium tuberculosis}$ H₃₇Rv grown $\text{in vitro}$ and $\text{in vivo}$ was studied. Uptake, measured after 3 minutes of drug exposure was inhibited mildly by 0.1 μg/ml and 0.2 μg/ml concentration and completely by 0.3 μg/ml. However, with the minimal inhibitory concentration (MIC)⁷ of 0.5 μg/ml, not only inhibition but also a strong efflux of the preformed tryptophan pool were observed. The results are discussed in the light of the theory that isoniazid interferes with the cell wall mycolate synthesis.     

Spatial requirements for insulin-sensitive sugar transport in rat adipocytes

(1) Alkyl sugar inhibition of d-allose uptake into adipocytes has been used to explore the spatial requirements of the external sugar transport site in insulin-treated cells. α-methyl and β-methyl glucosides show low affinity indicating very little space around C-1. The high affinity of d-glucosamine (K_i = 9.05 ± 0.66 mM) is lost by N-acetylation. $\text{N-}\text{Acetyl-}\text{d}\text{-glucosamine}$ shows no detectable affinity, indicating that a bulky group at C-2 is not accepted. Similarly $\text{2,3-}\text{di}\text{-O-}\text{methyl-}\text{d}\text{-glucose}$ (K_i = 42.1 ± 7.5 mM) has lower affinity than $\text{3-O-}\text{methyl-}\text{d}\text{-glucose}$ (K_i = 5.14 ± 0.32 mM) indicating very little space around C-2 but much more around C-3. A reduction in affinity does occur if a propyl group is introduced into the C-3 position. The K_i for $\text{3-O-}\text{propyl-}\text{d}\text{-glucose}$ is 11.26 ± 2.12 mM. $\text{6-O-}\text{Methyl-}\text{d}\text{-galactose}$ (K_i = 87.2 ± 17.9 mM) and $\text{6-O-}\text{propyl-}\text{d}\text{-glucose}$ (K_i = 78.07 ± 12.6 mM) show low affinity compared with d-galactose and d-glucose, indicating steric constraints around C-6. High affinity is restored in $\text{6-O-}\text{pentyl-}\text{d}\text{-galactose}$ (K_i = 4.66 ± 0.23 mM) possibly indicating a hydrophobic binding site around C-6). (2) In insulin treated cells $\text{4,6-O-}\text{ethylidene-}\text{d}\text{-glucose}$ (K_i = 6.11 ± 0.5 mM) and maltose (K_i = 23.5 ± 2.1 mM) are well accommodated by the site but trehalose shows no detectable inhibition. These results indicate that the site requires a specific orientation of the sugar as it approaches the transporter from the external solution. C-1 faces the inside while C-4 faces the external solution. (3) To determine the spatial and hydrogen bonding requirements for basal cells 40 μM $\text{3-O-}\text{methyl-}\text{d}\text{-glucose}$ was used as the substrate. Poor hydrogen bonding analogues and analogues with sterically hindering alkyl groups showed similar K_i values to those determined for insulin-treated cells. These results indicate that insulin does not change the specificity of the adipocyte transport system.     

Photodynamic alteration of cornea endothelium: Relation to bicarbonate fluxes and oxygen concentration

Corneas were mounted in flux chambers and endothelial bicarbonate fluxes were determined following sensitization of endothelial cells with 5 · 10^?6 M rose bengal and exposure to light. Corneas exposed to light demonstrated an increased passive bicarbonate flux compared to corneas not photosensitized. Active bicarbonate flux was reduced after 5 min of light exposure, but not after 1 min of light exposure. The increase in passive bicarbonate flux was prevented by the addition of 200 μg/ml catalase to the bathing solution; however, catalase had no effect on the photodynamic alteration of active flux. Neither 10 mM ascorbic acid nor 1.012 g/l glutathione prevented the photodynamically induced increase in passive flux. Perfusion of corneas with 5 · 10^?6 M rose bengal dissolved in a sucrose-substituted Krebs-Ringer bicarbonate solution with a $\text{p}{}_{\mathrm{<mtext>o</mtext>}}{}_2$ of $\text{124 ± 4.0}\text{mmHg}$ and exposed to light swelled at rates more rapid than corneas treated in a similar fashion but perfused with a solution with a $\text{P}{}_{\mathrm{<mtext>o</mtext>}}{}_2$ of $\text{20 ± 4.6}\text{mmHg}$. This study demonstrates that photodynamically induced corneal endothelial cell alteration results in increased passive bicarbonate flux, a time-dependent decrease in active bicarbonate flux, is oxygen dependent, and is at least in part secondary to H₂O₂ produced by the dismutation reaction of the superoxide free radical.