Contribution of plasma homovanillic acid (HVA) to urine and cerebrospinal fluid HVA in the monkey and its pharmacokinetic disposition.

Homovanillic acid (HVA) labelled with two deuterium atoms (d₂-HVA) was used to determine the contribution of HVA in the blood to HVA in the urine and CSF of monkeys. During and after a six-hour intravenous infusion of d₂-HVA at a constant rate, the levels of both d₂-HVA and endogenous HVA (d₀-HVA) in plasma, urine, and CSF were determined by gas chromatography-mass spectrometry, and the relative enrichments of d₂-HVA in each of these fluids calculated. Results indicate that HVA in the urine is derived exclusively from the blood, with no contribution from renal metabolism of dopamine (DA). Furthermore, less than one percent of HVA in either lumbar or ventricular CSF is derived from circulating HVA. The plasma elimination curve of d₂-HVA was biexponential, with a terminal phase half-life ($\text{t}{}_{\mathrm{<mtext>1</mtext><mtext>2</mtext>}}$) of 44 minutes and an apparent volume of distribution of 0.8 liters/kg.     

Determination of homovanillic acid turnover in man

Homovanillic acid (HVA) labelled with five deuterium (d) atoms was used to determine the total body turnover of HVA, the size of the peripheral body pool of HVA and HVA elimination characteristics in five healthy men. After i.v. injection of 5.5 μmoles (1 mg) of HVA-d₅ the levels of HVA-d₅ and endogenous HVA (HVA-d_o) in plasma and urine were followed by mass fragmentography using HVA-d₂ as the carrier and internal standard. Following an initial distribution phase of 10–20 minutes the plasma elimination curve of HVA-d₅ was monoexponential with a mean $\text{T}{}_{\mathrm{<mtext>1</mtext><mtext>2</mtext>}}$ of 0.66 hrs. The apparent volume of distribution (V_D) approximated the volume of the body water. The content of HVA in the peripheral body pool calculated from the plasma levels of HVA-d_o and V_D was 3.4 moles. The urinary HVA excretion rate (mean 1.70 moles/hour) was 45% of the total body turnover, the recovery of urinary HVA-d₅ was 48% of the mean body clearance. Together the results indicate that about 50% of the HVA formed in the body is eliminated by mechanisms other than renal excretion.     

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.     

Effects of ethanol, fructose, and ethanol plus fructose infusions on plasma glucose concentration and glucose turnover in monkeys (Macaca fascicularis) as measured by [6-3H]glucose

Six adult, female, cynomolgus monkeys were fasted for 64 hr and then continuously infused with [6-3H]glucose to determine the rates of glucose turnover and clearance while they were also being infused with ethanol (110 mumol/min/kg), 1,3-butanediol (110 mumol/min/kg), fructose (30 mumol/min/kg) or ethanol plus fructose (110 and 30 mumol/min/kg) respectively. Both ethanol and 1,3-butanediol infusions decreased the glucose turnover rate (the steady-state input-output rate from the plasma glucose pool) and the plasma glucose concentration by halving the glucose production rate. In contrast, fructose infusions increased the glucose turnover rate and glucose concentration by increasing the glucose production rate by 20%. The plasma clearance rate of glucose was lowest when the animals were infused with ethanol plus fructose; this suggests that acetate from ethanol oxidation may have a glucose-sparing effect if normoglycemia is maintained.     

Kinetics of Homovanillie Acid and Determination of Its Production Rate in Humans

Abstract: Homovanillie acid (HVA) labeled with either two or five deuterium atoms (d₂-HVA or d₅-HVA) was used to label the peripheral body pool of endogenous HVA (d₀-HVA) in control humans and in neurological patients. Either d₂- or d₅-HVA was rapidly injected intravenously and concentrations of d₂- or d₅- and d₀-HVA in sequential samples of blood plasma were determined by gas chromatography-mass spectrometry (GC-MS). Parameters describing the distribution and elimination of HVA, as well as its pool size, turnover, and synthesis rate were then calculated. Results indicate that the decline of plasma d₂- or d₅-HVA with time was multiexponential in five out of six subjects. Basal levels of endogenous HVA averaged 14.4 ± 1.3 ng/ml in the control patients and 8.69 ± 2.4 ng/ml in the neurological patients. The biological half-life averaged 71.3 ± 8.0 min in the control subjects and 91.3 ± 15 min in the neurological patients. The apparent volume of distribution of HVA in the body was 31–100 liters. Plasma clearance was 243–679 ml/min. The size of the peripheral body pool, calculated from plasma kinetic parameters, was 392–673 μg. The HVA rate of production, calculated for the whole body, was 166–323 μg/h. This technique can be used to determine accurately the rate of HVA production by the whole body over short time intervals. Since sample size was very limited ( n = 3 per group) and effects of variables such as age and sex on these data have not been excluded, more thorough investigations are needed to document any differences between normal controls and neurological patients.     

Interrelations between sulfate-reducing and methane-producing bacteria in bottom deposits of a fresh-water lake. III. Experiments with 14C-labeled substrates

An ecological substrate relationship between sulfate-reducing and methane-producing bacteria in mud of Lake Vechten has been studied in experiments using ¹⁴C-labeled acetate and lactate as substrates. Fluoroacetate strongly inhibited the formation of ¹⁴CO₂ from [U-¹⁴C]-acetate and β-fluorolactate gave an inhibition of similar magnitude of the breakdown of [U-¹⁴C]-l-lactate to ¹⁴CO₂ thus confirming earlier results on the specific action of these inhibitors. The turnover-rate constant of l-lactate was 2.37 hr^-1 and the average l-lactate pool size was 12.2 μg per gram of wet mud, giving a turnover rate of 28.9 μg of lactate/gram of mud per hr. The turnover-rate constant of acetate was 0.35 hr^-1 and the average pool size was 5.7 μg per gram of wet mud, giving a rate of disappearance of 1.99 μg of acetate/gram of mud per hr. Estimations of the acetate turnover rate based upon the formation of ¹⁴CO₂ from [U-¹⁴C]-acetate or [1-¹⁴C]-acetate yielded figures of the same magnitude (range 0.45 to 1.74). These and other results suggest that only a portion of the lactate dissimilated is turned over through the acetate pool. The ratio of ¹⁴CO₂/¹⁴CH₄ produced from [U-¹⁴C]-acetate by mud was 1.32; indicating that 0.862 moles of CH₄ and 1.138 moles of CO₂ are formed per mole of acetate. From the rate of disappearance of acetate (0.027 μmoles/gram wet mud per hr) and the rate of methane production (0.034 μmoles/gram wet mud per hr), it may be concluded that acetate is an important precursor of methanogenesis in mud (approximately 70%). A substrate relationship between the two groups of bacteria is likely since ¹⁴CH₄ was formed from [U-¹⁴C]-l-lactate.     

Prostaglandins do not modulate the positive chronotropic and inotropic effects of sympathetic nerve stimulation and injected norepinephrine in the isolated blood perfused canine atrium

In isolated canine atrium, perfused with blood from a donor dog, the infusions of both prostaglandins (PG)I₂ and E₂ (0.1–1 μg/min) into the sinus node arterial cannula neither altered the sinus rate and developed tension nor the positive chronotropic and inotropic responses elicited by either electrical stimulation or by injected norepinephrine. Infusion of arachidonic acid (10–100 μg/min), a precursor of PGs, or indomethacin (15–20 μg/min), an inhibitor of PG synthesis, into the sinus node arterial cannula also failed to alter the increase in sinus rate or developed tension produced by either adrenergic stimulus in the isolated atria. When arachidonic acid, 100–300 μg/kg or PGI₂, 1 μg/kg, were injected into the jugular vein of the donor dog, they produced a fall in systemic blood pressure; this effect of arachidonic acid but not of PGI₂ was abolished by indomethacin, 1 mg/kg. During administration of either arachidonic acid or indomethacin to the donor dog, the positive chronotripic and inotropic responses to adrenergic stimuli in the isolated atria also remained unaltered. These data indicate that PGs do not modulate adrenergic transmission in the blood perfused canine atrium.     

Metabolism of labeled carnitine in the rat

In two series of rats, the concentration of carnitine in plasma was 39.9 and 37.8 μmol/ liter, in skeletal muscle tissue 2.97 and 3.26 μmol/g dry wt and the urinary excretion 3.2 and 2.4 μmol/24 h. The renal clearance of carnitine was calculated to 88 and 76 ml/24 h. L-[Me-¹⁴C]Carnitine and DL-[Me-¹⁴C]carnitine have been administered to rats. Only labeled l-carnitine has been found on chromatographic analysis of plasma, urine, and muscle tissue. The specific radioactivity of carnitine in plasma, urine, and muscle tissue has been followed for up to 16 days. A two-compartment metabolic model has been used to interpret the result of the experiment with labeled l-carnitine and the rate constants and compartment sizes have been calculated. The total body content of carnitine was 57 μmol (about 35 μmol/100 g body wt) and the daily turnover was about 7% of the body pool. The daily synthesis of carnitine in the rat is estimated to about 2 μmol/100 g body wt.     

Effects of 5-phenyloxazolidinedione on sodium-potassium-magnesium-activated adenosine triphosphatase activity in mouse brain

—10 μm 5-Phenyloxazolidinedione (PKO) stimulated Na–K–Mg-ATPase activity in mouse brain homogenates by 45%. This activation effect was also observed for trimethadione and dimethadione. Inhibition of Na–K–Mg-ATPase by 0.1 mm ouabain was reduced in the presence of PKO. When PKO (4 mg/kg) was administered to mice intravenously, maximum levels of 2 μg (10 nmol)/g were attained in the brain within 15 min. PKO had an ld ₅₀ of 872 mg/kg and an ed ₅₀ of 375 mg/kg for protection against metrazol-induced seizures in mice.     

Leishmania donovani: Oral efficacy and toxicity of formycin B in the infected hamster

Formycin B, a structural analog of inosine, was evaluated as an orally administrable antileishmanial agent. Against Leishmania donovani in hamsters, it achieved an 85–92% reduction in numbers of parasites in livers of infected animals after oral administration at 13 mg/kg/day for 4 days. Its efficacy by oral administration was approximately four to eight times that by intramuscular administration and four times that of the positive control drug Glucantime by intramuscular administration. The levels of formycin B in serum after the final oral administration of 26 mg/kg/day were 1.4 μg/ml at 1 hr and 0.3 μg/ml at 2 hr. The concentration in liver was greater (9.0 μg/ml at 1 hr) and declined more slowly. With this latter dosage or with 104 mg/kg/day there was no acute toxicity of formycin B to bone marrow or formed elements of the blood. The only statistically significant toxicity to the liver was a doubling of serum total bilirubin levels. Comparison of the in vivo efficacy of formycin B against L. donovani to the mild acute toxicity of the drug suggests that formycin B has potential as an oral agent against visceral leishmaniasis.     

Fluid volumes in rainbow trout, Salmo gairdneri: application of compartmental analysis

1. The measurement of fluid volumes by the indicator dilution technique and compartmental analysis was re-evaluated in free-swimming, undisturbed rainbow trout. 2. Plasma (33.5 ml/kg body wt) and blood (41.3 ml/kg body wt) volumes estimated by compartmental analysis from blood samples taken early (less than 5 min) after dye injection were 40% lower than volumes calculated by sampling late (greater than or equal to 80 min). 3. The rate of exchange of dye between plasma and interstitial fluid was high (48%/hr) compared to mammals (5%/hr) which supports the hypothesis that teleost capillaries have high protein permeability. 4. Total extracellular volume estimated using a single pool model (210.5 ml/kg body wt) of inulin kinetics was 20% higher than that calculated by a three pool model (172.8 ml/kg body wt).     

THE EFFECT OF HYDROCORTISONE ON HeLa CELL GROWTH

HeLa Chessen cells have a doubling time of 18 hr when grown in MEM containing 10% calf serum and antibiotics. When hydrocortisone (1.7 μg/ml) is added to exponentially distributed cells in log growth in this medium, a new pattern of growth begins to emerge after 10–12 hr. This pattern is characterized by a transitional state lasting for about 6 hr, and then a new doubling time of about 35 hr is maintained thereafter. Hydrocortisone removes about 5% of the cells from the proliferative pool and extends the generation time of proliferating cells to about 30 hr. The extension of the generation cycle appears to occur almost entirely in late G₁. Cells grown as clones (average 6 cells/clone) prior to the addition of hydrocortisone, undergo these changes with doses as low as 0.00017 μg/ml of medium. When the average clone size is 1.5 cells per clone, the drug concentration must be 0.017 μg/ml or higher to initiate this response. The HeLa S₃ strain continues to grow with an 18-hr doubling time in the presence of hydrocortisone after a temporary delay in growth occurring between the 12th and 16th hour.     

Isolation of extensin precursors by direct elution of intact tomato cell suspension cultures

Dilute salt solutions eluted peroxidase and hydroxyproline-rich glycoproteins (HRGP's) very rapidly (60 % within 10s) from the surface of intact tomato cells grown in suspension culture. Further purification of the HRGPs based on (a) their solubility in 10% trichloroacetic acid and (b) chromatography on carboxymethyl cellulose, gave two components (P1 and P2) rich in serine, tyrosine, lysine and arabinosylated hydroxyproline. The sum of the hydroxyproline arabinoside profiles of P1 and P2 approximated that of the wall. P1, unlike P2, was histidine-rich and also contained proline. Significantly, isodityrosine (IDT) was absent from P1 and P2 but present in cell wall hydrolysates where, the Hyp:IDT molar ratio was ca 15: 1. In cells 4 days after subculture, ³H-proline pulse-chase data indicated turnover of P1 and P2 presumably resulting from covalent attachment to the wall as neither P1 nor P2 appeared in the growth medium. At day four the cell mean generation time (MGT) was 4.6 days, the cell hydr oxyproline content was 0.7 % (w/w), the half lives of P1 and P2 were both ca 12 hr, and the combined CaCl₂ elutable P1 and P2 precursor pools contained ca 400 μg Hyp/g cells (dry weight). Calculated from the MGT and Hyp content, the cell demand was 44.μg Hyp/g cells (dry weight)/hr. The precursor pool size was therefore sufficient for 9 hours growth. However the pool turnover calculated from half life and pool size was 5.6 %/hr or 22.4μg Hyp/g cells (dry weight)/hr. Thus the supply of P1 and P2 precursors met > 50 % of the cell wall demand. Corroborative experiments showed that after depletion of the P1 and P2 pools by salt elution, washed cells resuspended in growth medium repleted the precursor pools at a rate corresponding to a synthesis of 43μg Hyp/g cells (dry weight)/hr, or 98 % of the demand. These data allow us to make the following suggestions: P1 and P2 represent monomeric extensin precursor subunits. Salt elution of P1 and P2 indicates their ionic binding by pectic carboxyl groups. The rapidity of elution indicates a high diffusivity of these extended rodlike macromolecules through the cell wall. This may imply a preferred orientation for P1 and P2 perpendicular rather than parallel to the plane of the wall. The lack of IDT in P1 and P2 implies that IDT forms in muro, possibly via peroxidase. We speculate that some of these IDT residues may crosslink an extensin precursor ‘tweft’ around a cellulose microfibrillar ‘twarp’. Such formation of heteromultimeric extensin interpenetrated by microfibrils would create a mechanically coupled extensin-cellulose network.     

The contribution of serum triacylglycerol to hepatic triacylglycerol turnover in the starved rat.

The present study was undertaken to evaluate quantitatively the turnover of serum triacylglycerol (triglyceride) in the starved rat and to determine whether serum triacylglycerol recycled to liver contributes a significant fraction of the total hepatic triacylglycerol turnover. Serum was labelled in vitro with [3H]trioleoylglycerol (glycerol [3H]trioleate) to provide uniform labelling of all lipoprotein species. By using the curves describing disappearance of isotope from serum and its appearance in liver, rate constants for movement of triacylglycerol out of serum (0.29 min-1) and the uptake of serum triacylglycerol by liver (0.22 min-1) were calculated. The total rate of movement (flux) of triacylglycerol in these processes, the product of rate constant and serum pool size, was calculated to be 0.39 and 0.29 mg/min per 100 g body wt. respectively. A model is postulated for whole-body triacylglycerol metabolism consistent with the present data as well as most observations in the literature. From the model it can be predicted that: (1) the entire turnover of liver triacylglycerol in the starved rat can be accounted for on the basis of contributions from serum non-esterified fatty acid and serum triacylglycerol; (2) the entire turnover of the serum triacylglycerol pool can be accounted for quantitatively on the basis of contributions from intestine and liver; (3) the release rate for triacylglycerol from liver should be 0.34 to 0.35 mg/min per 100 g body wt.; (4) triacylglycerol synthesized by liver from non-esterified fatty acid of serum and by intestine can account quantitatively for the irreversible disposal rate of triacylglycerol from serum.     

Separate mechanisms of induction for ornithine decarboxylase by triiodothyronine and aminophylline

A single dose of aminophylline (200 μmol/kg, i.p.) or triiodothyronine (T₃, 300 μg/kg, i.p.) resulted in the induction of ornithine decarboxylase (ODC) in rat liver with maximal activity 10-fold and 6-fold above controls, respectively, 4 hr after the administration of the drug or hormone. After either agent, the induction of ODC was blocked by either cycloheximide or actinomycin D. The same concentrations of aminophylline and T₃ administered simultaneously produced an additive 16-fold increase in ODC activity. After T₃ administration, the cyclic AMP-dependent protein kinase activity ratio was unaltered at all times measured. After aminophylline, the protein kinase activity ratio was elevated by 15 min and remained elevated for 2 hr. Somatostatin administration (50 μg/100 g), which lowers plasma growth hormone to 30% of control, had no effect on the ability of T₃ to induce ODC. These data suggest separate routes of induction of ODC in response to aminophylline and T₃. Aminophylline induction occurs via cycyclic AMP-mediated event whereas T₃ does not involve ccyclic AMP but results from a direct nuclear interaction.     

Size and specific activity of the UTP pool and overall rates of RNA synthesis in early mouse embryos

Mouse embryos from the one-cell to the blastocyst stage were cultured for 2 hr in the presence of 5 μM [³H]uridine or 10 μM [³H]adenosine, and the size and specific activity of the UTP and ATP pools were determined by an Escherichia coli RNA polymerase assay using synthetic poly(dA-dT) as template. The total UTP pool increased in size and specific activity with development from 0.05 pmole (0.06% labeled) in the one-cell stage to 0.54 pmole (27% labeled) in the blastocyst stage. The total ATP pool remained relatively constant in size at about 1 pmole/embryo, but increased in specific activity from 2.6 to 52% from one-cell to blastocyst. The turnover of the [³H]UTP pool was also examined under pulse-chase conditions in eight-cell and morula-stage embryos. The UTP pool decayed with approximately first-order kinetics up to 20 hr of chase, but the rate of decay was slower in eight-cell embryos (t_0.5 = 5.5 hr) than in morulae (t_0.5 = 2.8 hr). The observed specific activities of the UTP pools were used to calculate the overall rates of uridine incorporation into acid-precipitable material during early development. The rate of uridine incorporation per embryo increased from 3.6 × 10^?3 pmole/2 hr in the two-cell embryo to 1.8 × 10^?1 pmole/2 hr in the blastocyst. The rate of RNA synthesis per cell over a 2-hr period was estimated at 2.5 pg in the two- to four-cell embryo, 5 pg in the eight-cell, and 10 pg in the morula-early blastocyst.     

Fetotoxicity of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) for rhesus macaques (Macaca mulatta)

The fetotoxic and teratogenic potential of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) for rhesus macaques (Macaca mulatta) was tested through oral administration to monkeys early in pregnancy. A single or divided dose, 1 μg of TCDD/kg of body weight, was followed by abortion in 13 of 16 pregnant monkeys treated between days 20 and 40 of gestation. One of four aborted at 0.2 μg/kg, and two of two at 5 μg/kg. None of the mothers given 0.2 μg/kg showed signs of toxicity. Eight of the monkeys aborting at 1 μg/kg showed clinical toxicity 44 to 111 days after aborting, and three died. Both given 5 μg/kg became toxic soon after abortion and died. No malformations except for two minor palatal abnormalities of questionable significance were found in the six fetuses that were not aborted at doses of 0.2 and 1.0 μg/kg. These results indicate (1) that TCDD is fetotoxic at doses that frequently have delayed toxicity to the mother, but that conclusions about teratogenicity cannot be drawn, and (2) that pregnant rhesus females are more sensitive to the toxic effects of TCDD than any species tested but the guinea pig.     

Action of glucagon and atropine on mucosal blood flow of resitng fundic pouches of dogs.

Clearance of plasma aminopyrine (an index of mucosal blood flow, MBF) into acid (0.1 N HCl) instilled into Heidenhain pouches was about 200 ml per 30 min under basal conditions. Glucagon 50 μg/kg s.c. significantly decreased the MBF from 200 to 132 ml per 30 min at one hr; the decrease lasted about 60 min. Infusion of glucagon 50 μg/kg i.v. for 1 hr produced a delayed reduction of MBF lasting for more than 90 min. Under the same experimental conditions, atropine 50 μg/kg reduced MBF from 200 to 166 ml per 30 min upon subcutaneous administration and showed no significant effect by i.v. infusion. The increase in residual volume of the pouch caused by glucagon could not account for all the decrease in clearance of aminopyrine. We conclude that glucagon reduces gastric mucosal blood flow under basal conditions.     

Prostacyclin-induced hypothermia: Involvement of central histamine H2-receptors

Some of the biological activities of prostacyclin (PGI₂) are known to be mediated through cyclic AMP (cAMP). The purpose of this study was to assess the involvement of histamine and serotonin receptors as well as cAMP in the PGI₂-induced hypothermia in conscious guinea pig. Intracerebroventricular administration of 50–500 μg/kg PGI₂ produced a dose-related hypothermia, whereas its stable metabolite 6-keto prostaglandin F₁α had an insignificant effect. Low central doses (10–50 μg/kg) of dibutyryl cAMP (DBC) were hyperthermic, but high doses (100–500 μg/kg) caused hypothermia. Theophylline and low doses of DBC given centrally attenuated the PGI₂-induced hypothermia. Mepyramine and methysergide did not antagonize the effects of PGI₂ or DBC. However, central administration of metiamide (10–100 μg/kg) attenuated the hypothermic responses to both PGI₂ and DBC. These results suggest that histamine H₂-receptors are involved in the hypothermia induced by PGI₂.     

Sodium 5-(3′-pyridinylmethyl)benzofuran-2-carboxylate (U-63557A), a new,selective thromboxane synthase inhibitor: Intravenous and oral pharmacokintics in dogs and correlations with exsitu thromboxane B2 production

The pharmacokinetics of a new, selective thromboxane synthase inhibitor, sodium 5-(3′-pyridinylmethyl)benzofuran-2-carboxylate were determined for single dose, bolus intravenous injections (1, 3, and 10 mg/kg); for continuous 24 hr infusions (10 and 30 μg/kg/min); and for oral doses of gelatin encapsulated powdered drug (3, 10, and 30 mg/kg). Drug disappeared biexponentially after intravenous administration, and plasma concentrations were proportional to the dose. Absorption of drug occurred rapidly after its oral administration; peak plasma levels occurred 1–2 hours after ingestion, and circulating drug was detectable within 30 minutes. For all experiments, inhibition of cellular thromboxane B₂ production, $\text{ex situ}$, corresponded with plasma drug levels and its reactivation corresponded with disappearance of the drug indicating that it was not accumulated by platelets.