An assessment of the contribution of clonidine metabolised from alinidine to the cardiovascular effects of alinidine.

Five healthy volunteers (mean age 20.6 years, mean weight 71 kg) received in random order on day 1 and day 8 a single dose of alinidine 40 mg, clonidine 0.1 mg or placebo and on days 2-7 alinidine 40 mg, clonidine 0.1 mg or placebo given three times a day with 1 week between treatment periods. Blood samples were taken for measurement of concentrations of alinidine and clonidine during alinidine administration and of clonidine during clonidine dosing. Heart rate and blood pressure were recorded in supine and standing positions and heart rate after 3 min exercise. Plasma concentrations of alinidine reached a maximum of 163.6 +/- 10.0 ng/ml 2 h after alinidine administration on day 1 and during chronic administration similar concentrations were achieved. Clonidine plasma concentrations reached 0.3 +/- 0.11 ng/ml 6 h after alinidine 40 mg on day 1, and during chronic administration of alinidine, increased to a steady state on day 5 with trough and 2 h values of 0.73 +/- 0.15 and 0.86 +/- 0.14 ng/ml respectively. After the first dose of clonidine on day 1, the maximum plasma concentration of clonidine was 0.32 +/- 0.1 ng/ml at 4 h, during chronic administration clonidine plasma concentration rose to 1.04 +/- 0.14 ng/ml 2 h after a dose on day 5. Alinidine produced a greater reduction in the exercise tachycardia than clonidine.(ABSTRACT TRUNCATED AT 250 WORDS)     

Activity of the triazole antifungal r126638 as assessed by corneofungimetry

BACKGROUND: R126638 is a novel triazole exhibiting potent in vitro and in vivo antifungal activity against fungal pathogens including dermatophytes and yeasts. OBJECTIVE: To determine the antifungal activity in time in the stratum corneum of healthy volunteers after oral intake of R126638 at a daily dose of 100 or 200 mg for 1 week. METHOD: Sixteen male volunteers were randomly allocated to oral treatment with either 100 or 200 mg of R126638 once daily for 1 week. Five cyanoacrylate skin surface strippings (CSSS) were obtained from the forearm of each subject before drug intake at day 1. CSSS were also collected during treatment at day 2 (24 h after the first drug intake, before the second drug intake), at day 4 (before the fourth drug intake) and at day 7 (10 h after the last drug intake). The post-treatment lingering effect was assessed at day 10 (3 days after treatment) and at day 14 (7 days after treatment). The corneofungimetry bioassay was performed on these CSSS to assess the antifungal profile of R126638. Cells of different fungal species (Trichophyton rubrum, Trichophyton mentagrophytes, Microsporum canis, Candida albicans and Malassezia globosa) were deposited and cultured for 10 days on CSSS in a sterile and controlled environment. The extent of fungal growth on the stratum corneum was determined using computerized image analysis. RESULTS: R126638 clearly reduced the growth of all tested fungal species. The onset of effects of R126638 was evidenced at day 4 when it reached statistical significance for 3 of 5 species. At day 7, significance was reached for 4 of 5 species. During the posttreatment period, R126638 remained effective for 4 of 5 species at day 10, and this activity persisted until day 14 for 2 of 5 species. CONCLUSION: A broad spectrum antifungal activity was rapidly expressed in the stratum corneum after oral intake of R126638. The drug likely reached the upper layers of the stratum corneum by diffusion and persisted in this location for at least 7 days after treatment.     

Biochemical and toxicological effects of combined exposure to 1,1,1-trichloroethane and trichloroethylene on rat liver and brain

1. Inhalation exposure of adult male rats to a mixture of 1,1,1-trichloroethane (500 p.p.m.) and trichloroethylene (200 p.p.m.) for four days 6 h daily resulted in an accumulation of 1,1,1-trichloroethane in perirenal fat. Further exposure on the fifth day caused a rapid increase in various organ contents of both solvents with secondary depression of brain RNA. 2. The four-day exposure doubled the RNA content of liver and caused a slight decline in the concentrations of glutathione in liver. 3. The amount of cytochrome P-450 in liver was increased, as well as the overall mono-oxygenase activity, measured with styrene as substrate. During continuing treatment on the fifth day, styrene mono-oxygenase activity decreased, the activity after 6 h being only about 50% of that at the beginning of the fifth day of exposure. 4. UDP-glucuronosyl transferase activity (measured in digitonin-activated microsomes) was doubled by the four-day combined exposure to 1,1,1-trichloroethane and trichloroethylene. 5. The changes during the fifth day of exposure, e.g. rapid increase in the concentrations of solvents in organs, the detection of trichloroethylene in tissues and depression of mono-oxygenase activity, obviously also occurred during the exposures on days 1 to 4 and reverted during each post-exposure period.     

Metabolism of sulfadiazine in neonatal and young pigs. Comparative in vivo and in vitro studies

Metabolism of sulfadiazine (SDZ) was studied in vivo and in vitro during postnatal development of piglets in order to examine whether in vitro metabolism approaches the in vivo situation. Experiments were performed in 1-day-, 8-day- and 60-day-old piglets. In vivo: 14C-SDZ was injected intravenously and urine and tissue samples collected after 3 hr. Urinary excretion data as well as data from liver and kidney tissue indicated a relatively high capacity for acetylation at birth, while the capacity for oxidation is low during the first week of life. At 60 days of age the acetylation and oxidation of SDZ is equal each accounting for about 20% of the amount excreted in urine. In vitro: Incubation of subcellular fractions of liver and kidney showed that acetylation of SDZ in liver reached maximum within 1 week. Oxidative activity was absent at 1 day, present at a low level at day 8, and at a high level at day 60. Neither acetylation nor oxidation of SDZ took place in kidney. The results show a close correlation between in vivo and in vitro results with respect to the developmental pattern seen in piglets during the postnatal period of life.     

Ontogenesis of choline acetyltransferase, tyrosine hydroxylase, monoamine oxidase and catechol-O-methyl transferase in the superior cervical ganglion of swine

Swine have ca 50 per cent of the adult choline acetyltransferase (CAT) activity in the superior cervical ganglia (SCG) at birth. This activity rapidly increases to adult levels by 7–14 days of age. Tyrosine hydroxylase (TH) activity is ca 10 per cent of adult levels at birth. This enzyme increases at the same rate as SCG growth from 0–14 days of age and then changes 4- to 5-fold from 14–39 days of age. Monoamine oxidase (MAO) and catechol-O-methyl transferase (COMT) activities increase at the same rate as ganglion growth from birth through 150 days of age. The TH and MAO, but not CAT or COMT, activities were reduced ca 30 per cent in neonatal piglets treated for 5 days with 6-hydroxydopamine (50 mg/kg day, s.c.). Catecholamine levels in heart and spleen were reduced > 90 per cent by 6-hydroxydopamine, but TH, MAO or COMT was not diminished. The physically mature neonatal swine may be less susceptible to chemical sympathetecomy by 6-hydroxydopamine than immature neonatal mice or rats.     

Lead toxicity in cyanobacterial porphyrin metabolism

The effect of Pb2+ on growth, tetrapyrrole photosynthetic pigment content, total free porphyrin, and 5-aminolevulinate dehydratase (ALA-D) activity of a cyanobacterium, Microchaete tenera, and its ability to sequester Pb2+ from the culture medium were studied. Pb2+ was assayed by graphite furnace atomic absorption spectrophotometry. M. tenera growth and chlorophyll a content were not affected by 0.5, 1.0, and 6.0 ppm of Pb2+. These treatments doubled the protein content and increased the phycobiliprotein content by four times after 7 days. The ALA-D activity decreased in all concentrations by 63% at day 7 and by 34% at day 14. As a consequence of ALA-D inhibition, total free porphyrin also decreased by 64% at day 7 and by 40% at day 14. The highest biomass lead uptake (7454 +/- 565 micrograms Pb2+/g dry weight) was observed at day 3 with 6.0 ppm of Pb2+ in the culture medium. Uptake coefficient was highest (3723 +/- 279 micrograms Pb2+ g-1 dry weight/ppm of applied Pb2+) with 1.0 ppm after 3 days. The increase in protein and antenna pigments on day 7 was probably a response to stress conditions and could explain why the toxic metal did not affect growth. ALA-D inhibition and high lead biomass content confirm the importance of this enzyme as a biological indicator for stress.     

Ontogenesis of enzyme systems deaminating different monoamines.

1 A detailed investigation into the postnatal development of the activity of the enzyme monoamine oxidase (MAO) in the rat and domestic pig was carried out. 2 MAO activity was measured in littermate male rats aged between 3 and 122 days belonging to six breeding colonies. The tissues studied were three brain regions in which monoamines may play a role in neuronal transmission (septum, hypothalamus, corpus striatum) and, for comparison, in the cerebellum. Liver, heart and adrenal glands were the peripheral organs studied. The following substrates were used to measure MAO activity in each tissue homogenate: kynuramine, tyramine, dopamine, tryptamine and 5-hydroxytryptamine (5-HT). 3 MAO activity towards kynuramine, tyramine and dopamine increased after birth in all brain regions and also in the liver, to reach maximal values between days 40 and 80. In the heart and the adrenal glands enzyme activity remained low up to 30-40 days and then increased steeply. This was the case in all litters examined. 4 All tissues deaminated more tyramine than dopamine. In the liver, the ratio of the quantities of tyramine deaminated/dopamine deaminated was approx. 2 at all ages. In the homogenates of whole brains (including or excluding the hypothalamus and striatum) this ratio was also 2 at all ages. In contrast in the isolated striatum and hypothalamus it was first much higher and reached a value of 2 only at an age of about 20 days. This may indicate an independent development of a dopamine and a tyramine deaminating enzyme system in discrete brain regions. It was suggested, that the low ability to deaminate dopamine in discrete brain regions may be due to the local presence of an enzyme inhibitor which becomes too diluted to be active in homogenates of whole brain. 5 Deamination of tryptamine in the striatum decreased between day 5 and 20 in 3 out of 4 colonies tested. There was a large fall in the deamination of 5-HT in all tissues of one group of rats, but in another 4 groups the tissues of the 5 day old rats deaminated smaller amounts of 5-HT than those of the older rats. 6 Purified hypothalamic mitochondria from 40 day old rats deaminated more tyramine and dopamine but not tryptamine per mg protein than those from 5 day old rats. 7 In the domestic pig there was a significant rise in the values in hippocampal MAO activity towards dopamine and tyramine from the foetus (55 day gestation) to the 1 week old piglet. A further steady rise up to week 6 was indicated, but this rise was not statistically significant. The difference between rat and pig probably reflected the much higher degree of maturity of the latter at birth. 8 In the hippocampus of the pig the ratio between the amount of tyramine deaminated/dopamine deaminated decreased from greater than 10 (foetus) to 4.8 in the 6 week old pig and 2 in the adult.     

Serum transforming growth factor-beta1 levels increase in response to successful anti-inflammatory therapy in ulcerative colitis

OBJECTIVE: To investigate serum levels of transforming growth factor-beta1 and interferon-gamma in active ulcerative colitis and to assess changes during treatment. METHODS: We prospectively evaluated serum from 25 patients with untreated active ulcerative colitis and 19 healthy controls. Disease activity score (DAI), serum transforming growth factor-beta1 and interferon-gamma levels were measured at baseline and after 7 days of conventional treatment. Disease activity score and transforming growth factor-beta1 were also assessed at 42 days. RESULTS: Baseline transforming growth factor-beta1 levels were significantly higher in patients than in controls (P < 0.02). On the 7th day, transforming growth factor-beta1 levels increased only in patients who responded (P < 0. 01); variations in transforming growth factor-beta1 levels and disease activity score were inversely correlated (r=- 0.72, P < 0. 001). At day 42, serum transforming growth factor-beta1 decreased significantly compared with the 7th day (P < 0.05). While in controls, interferon-gamma was undetectable; untreated patients had higher, widely variable, levels. At day 7, responders had higher interferon-gamma values than unresponsive cases. Variations in interferon-gamma correlated moderately with changes in transforming growth factor-beta1 (r=0.53, P < 0.05). Cytokine response did not depend upon the type of treatment. CONCLUSIONS: Both transforming growth factor-beta1 and interferon-gamma may play a role in the injury-repair process in active ulcerative colitis. Variations in circulating transforming growth factor-beta1 levels in the first week of treatment seem to be related to the therapeutic response.     

Development of the hepatic mixed function oxidase system in a marsupial, the quokka (Setonix brachyurus).

The development of the mixed function oxidase system has been studied in pouched and young-at-heel joey quokkas. Cytochrome P-450 was first detectable at 35 days, rose sharply to one-third of the adult level by 53 days, and reached levels in the adult range between 155 and 255 days. NADPH-Cytochrome c reductase was approximately one-fifth the adult level at 45 days and , by 80–105 days of life, had reached values within the range for adult animals. Benzpyrene hydroxylase activity was barely detectable at 35 days; its development lagged behind that of the components of the cytochrome P-450 system and adult levels were not seen until 200 days. Electron microscopy showed that hepatocytes in 20- and 35-day joeys contained some short chains of rough endoplasmic reticulum. Areas of smooth endoplasmic reticulum were first evident at 45 days and at subsequent stages (80, 100, and 150 days and adult) increasing concentrations of both smooth and rough endoplasmic reticulum were present. Development of the mixed function oxidase system in the quokka liver is slow and resembles more closely the pattern of development in human fetal liver than that in many common laboratory animals.     

The inducibility and ontogeny of rat liver UDP- glucuronyltransferase toward furosemide

Furosemide (F) conjugation with glucuronic acid is the main pathway of F metabolism in humans and experimental animals. In order to study rat liver microsomal UDP-glucuronyltransferase (UDP-GT) activity towards F we developed an in vitro assay in which the conjugation product, furosemide 1-0-acyl glucuronide (FG) was separated and quantitatively determined by reverse phase high pressure liquid chromatography. The optimal conditions of the reaction were established and the apparent K_m for F and UDP-glucuronic acid (UDPGA) were 0.22 and 1.76 mM, respectively. Substrate inhibition of UDP-GT toward F occured at F concentrations higher than 1.5 mM. Developmental changes in F glucuronidation were compared to the ontogeny of UDP GT activity toward two other acceptors, 1-napthol and esterone that are known to have different patterns of maturation. F glucuronidation was 26% of adult activity at 18 days of gestation, reached 48% at birth and gradually increased to 250% of adult activity at 22 days of age. Glucuronidation of 1-napthol and estrone attained 87% and 44% of adult activity at 22 days of gestation, 37% and 66% in six-day-old rats and 100% and 427% of adult activity in 22-day-old rats, respectively. The effect of 3-methylcholantrene (3-MC), phenobarbital (PB) and pregnenolone-16α-carbonitrile (PCN) on F UDP-GT was studied and compared to their effect on 1-napthol and estrone glucuronidation. PB, 3-MC and PCN increased F-UDP-GT activity to 208%, 282% and 342% of vehicle-treated animals, respectively, while F pretreatment did not affect the conjugation of F. In comparison, 1-napthol glucuronidation was preferentially induced by 3-MC (4.4-fold of control) while estrone glucuronidation was induced by PB and PCN (4.9- and 2.5- fold of control, respectively). These studies suggest that several forms of UDP-GT activities, which differ in their ontogeny and inducibility patterns, are involved in the glucuronidation of F in vitro.     

Comparison of two weeks versus one week of prenatal ethanol exposure in the rat on gonadal organ weights, sperm count, and onset of puberty.

Sprague-Dawley dams from Harlan Ind. (Indianapolis, IN) were administered a fortified ethanol liquid diet containing 35% ethanol derived calories for two weeks (E-2) beginning on day 7 or one week (E-1) beginning on day 13 of gestation and continuing through parturition. Control dams were pair-fed an isocaloric liquid diet containing no ethanol during these periods or remained on lab chow and water. E-2 dams consumed an average of 13.52 g ethanol/kg bwt during the first week of exposure (days 8-14) and 12.50 g ethanol/kg bwt the second week (days 14-20). E-1 dams consumed significantly less than E-2 dams during the second week (9.75 g/kg; p < 0.0001). Although the lower consumption in E-1 dams led to a significant decrease in maternal weight gained during the few days of pregnancy compared to E-2 dams, birthweights of E-1 offspring were significantly heavier than those of E-2 offspring (p < 0.05). No effect of ethanol was detected on anogenital distance at birth in either sex. Puberty was delayed in female offspring of both E-1 and E-2 dams (p < 0.01) as measured by age of vaginal opening. These data suggest that the primary teratogenic actions of ethanol in the rat on fetal growth, as well as delayed puberty in females, occur in the last week of gestation. In adult E-2 males, testis weight was significantly heavier than all other groups when indexed to body weight. No effect of prenatal ethanol exposure was observed on the indexed weights of prostate, epididymis, or seminal vesicles.(ABSTRACT TRUNCATED AT 250 WORDS)     

Development of phase II xenobiotic metabolizing enzymes in differentiating murine clara cells

Glutathione S-transferases (GSTs) and epoxide hydrolases (EHs) protect cells from exogenous insult by detoxifying electrophilic compounds. Little is known about these enzyme systems during postnatal lung development. This study was designed to help establish whether the heightened neonatal susceptibility of the lung to bioactivated cytotoxicants is the result of inadequate ability to detoxify reactive intermediates. We compared the distribution of immunoreactive protein and enzymatic activity of GSTs and EHs in isolated distal airways during pre- and postnatal development in lungs of mice from 16 days gestation to 9 weeks postnatal age (adult). GST alpha, mu, and pi class protein expression in fetal and postnatal lung varied by isozyme and age. Isozymes alpha and mu are expressed at low levels before birth, high levels on postnatal day 7, low levels between postnatal days 14 and 21, high levels at postnatal day 28, and slightly lower levels in adults. Immunoreactive protein of isozyme pi has a peak expression on gestational day 18 and again on postnatal day 4, is undetectable at postnatal day 21, and is at peak levels in the adult mouse lung. GST activity in distal airways increased with age. Microsomal EH protein expression increased in intensity with age, while activity was similar in airways from all ages. We conclude that in the mouse lung (1) cellular expression of glutathione S-transferase varies by age and isozyme and does not increase with increasing age, (2) airway glutathione S-transferase activity increases with increasing age and does not correlate with immunoreactive protein expression, and (3) airway microsomal epoxide hydrolase activity does not increase, even though immunoreactive protein expression does increase with age.     

Neonatal exposure to toluene: effects on the development of liver microsomal cytochrome P-450 and serum hormone levels in the rat

Lactating Sprague-Dawley rats and their pups were exposed on postnatal days 1-7, 6 h/day, to 80, 500 and 1000 ppm toluene, respectively, by inhalation. Exposure to 80 ppm toluene decreased the liver microsomal AHH activity and the rate of 7 alpha- and 6 beta-hydroxylation of androstenedione in 8-day-old-pups. On the other hand, neonatal exposure to 500 or 1000 ppm toluene resulted in a significant increase in AHH and 7-ethoxyresorufin O-deethylase activities and in the formation of 16-oxygenated metabolites of androstenedione in 8-day-old animals. Exposure to toluene increased the cytochrome P-450 content at all 3 dose levels in male but not in female pups. Twenty-one days after neonatal exposure no such effects were seen in young animals of either sex. In 56-day-old male rats, however, neonatal exposure to 80 ppm toluene resulted in a decreased rate of 6 beta-hydroxylation of androstenedione and a reduced AHH activity. No such effects were seen in female rats of the same age. Neonatal exposure to toluene affected the body and liver weights in 8-day-old pups of both sexes but had no effect on these parameters in 21-day-old animals of either sex. Exposure to 80 ppm toluene during the neonatal period gave a significantly increased body weight of 56-day-old male but not of female rats of the same age although this treatment increased liver weight in both sexes at this age. Serum testosterone levels were decreased in 21-day-old male rats following neonatal exposure to 80 or 500 ppm toluene and in 56-day-old male rats exposed neonatally to 1000 ppm toluene. In conclusion, exposure to toluene during the first week of life caused significant changes in various liver microsomal cytochrome P-450 dependent enzyme activities in 8-day-old pups, whereas the long-term effects on liver metabolism of the adult animal were small.     

Behavioral changes induced in developing rats by an early postnatal exposure to lindane

The purpose of this study was to determine whether the behavioral developmental pattern was altered by an early postnatal exposure to lindane. Male and female offspring of Wistar rats were daily orally administered with a nonconvulsant dose of lindane (10 mg/kg) during 7 days either the 1st or the 2nd postnatal week days. Effects on pups were evaluated with a reduced developmental neurotoxicological test battery. Body weight evolution, neuromotor reflexes (surface righting, cliff avoidance and tail hang reflex) and spontaneous motor activity were analyzed from day 1 after birth up to day 28. The body weight pattern was unaffected by treatment with lindane and no signs of overt toxicity were observed. Lindane-treated pups showed an increased positive response of the neuromotor reflexes. Furthermore, lindane produced hyperactivity, especially manifested between days 12 and 16. A peak of activity was reached at day 16 in lindane-treated groups, while control animals had a maximum between days 20 and 24. These results suggest that low nonconvulsant doses of lindane may induce behavioral changes in developing rats.     

Aversive properties of theκ opioid agonist U50,488 in the week-old rat pup

Opioids have been shown to produce analgesia and to be reinforcing during the first week of life in the rat. Opioids also have analgesic actions in both the infant and adult, but can be aversive in the mature animal. We examined the aversive effects of the opioid agonist U50,488 during the first postnatal week in the rat pup in three ways. In the first experiment, U50,488, injected peripherally (1.0–30.0 mg/kg), was paired with an odor and pups were tested 8 h later for positional preference for avoidance of that odor. This task is similar to conditioned preference/aversion tests used with adult animals. Both 3- and 7-day-old pups learned to avoid the odor adulterated side at the two higher doses. When exposed to odors previously associated with U50,488, pups at both ages decreased locomotor activity. In a second experiment, acute treatment with U50,488 increased ultrasonic distress vocalizations (USV) equally at 3 and 7 days of age, increased locomotor activity, and decreased rectal temperature. Neither of the latter two effects was correlated with the increase in USV production. The third experiment showed that conditioned odor cues increased USV 8 h later in 3- and 7-day-old pups at 1.0–10.0 mg/kg without changes in activity or rectal temperature. The results from these studies suggest that U50,488 can produce aversions in the neonatal rat pup as it does in the adult.Supported by U.S.P.H.S. grant DA-06600     

Activities of drug metabolizing enzymes in bovine colon epithelial cell cultures

The metabolic competence of cultured bovine colon epithelial cells was evaluated by determining activities of phase I and II enzymes in colonocytes cultured for different intervals (maximum of 10 days) compared with activities measured in freshly isolated cells. Cytochrome P450 1A1-associated 7-ethoxyresorufin O-deethylase (EROD) activity was detectable in freshly isolated colonocytes and in colon cells maintained in culture for up to 5 days. In contrast to liver samples, cytochrome P450 3A4-associated 7-benzyloxyresorufin O-debenzylase (BROD) activity was not detectable in bovine colon cells. Prostaglandin H synthase-mediated production of prostaglandin E₂ was found in freshly isolated and also in cultured colonocytes. Both isoenzymes (COX 1 and COX 2) were detected in cultured cells. To examine phase II metabolic potency, activities of N-acetyltransferases 1 and 2, of phenol and amino sulfotransferases, of glutathione S-transferases alpha, mu, pi and theta and of UDP-glucuronyltransferase were measured. N-Acetyltransferase (NAT) activity (substrate p-aminobenzoic acid, PABA, a diagnostic substrate for the human NAT-1 enzyme) was stable under culture conditions and during the observed culture period comparable to that of freshly isolated cells. In contrast, sulfamethazine, a specific substrate for NAT-2, was not acetylated, neither in bovine colon cells nor in bovine liver samples. Whereas activity of amino sulfotransferase (substrate 2-naphthylamine) decreased continuously during the entire culture period, the activity of phenol sulfotransferase (substrate 1-naphthol) decreased only slowly. Activity of total glutathione S-transferases (alpha, mu, and pi) (substrate 1-chloro-2,4-dinitrobenzene) decreased after 2 days in culture, but was stable during the following culture period. Activity of glutathione S-transferase theta (substrate epoxy-3-nitrophenoxypropane) changed during the culture period. At the beginning and the end (after 10 days) of the culture period maximum activity was measured. Activity of UDP-glucuronyltransferase increased during the culture period reaching a maximum after 7 days. The results show that cultured bovine epithelial colon cells express several enzyme activities required for the biotransformation of xenobiotics.     

Duration of pleconaril effect on cytochrome P450 3A activity in healthy adults using the oral biomarker midazolam.

The objective of this study was to evaluate the duration of oral pleconaril (a picornavirus inhibitor) effect on intestinal and hepatic cytochrome P450 (P450) 3A activity as assessed by oral midazolam. Healthy adults received oral midazolam (0.075 mg/kg) on days 1 (baseline), 7, 9, 13, 20, 27, and 34. Oral pleconaril (400 mg) three times daily for 15 doses was administered on days 2 through 7. Blood samples were collected during each day of midazolam dosing to determine plasma midazolam concentrations. On days 5, 6, and 7, blood samples were collected to determine plasma pleconaril concentrations. Midazolam pharmacokinetics were determined by noncompartmental analyses, with bioequivalence assessed by least-squares geometric mean ratios (LS-GMR) and 90% confidence intervals (90% CI). Eighteen subjects completed the study. Midazolam C(max) (LS-GMR; 90% CI) decreased 24% on day 7 (0.76; 0.66-0.87). Midazolam oral clearance increased 53% on day 7 (1.53; 1.38-1.69). Midazolam oral clearance remained different on days 9 (1.38; 1.25-1.52) and 13 (1.19; 1.07-1.31) versus day 1. Midazolam volume of distribution (1.82; 1.57-2.11) and elimination half-life (1.19; 1.03-1.38) were also different on day 7 in comparison with day 1. Oral pleconaril increased intestinal and hepatic CYP3A activity. The duration of increased CYP3A activity by pleconaril was at least 6 days (but no longer than 13 days) after pleconaril discontinuation.     

Assessment of some clinical and laboratory variables for early diagnosis of cumulative copper poisoning in sheep

Sixteen male Suffolk lambs fed a 8 ppm Cu basal diet were randomly assigned to 2 groups: 12 copper-loaded (CL) and 4 controls (C). The CL sheep were drenched initially with 3 mg Cu/kg bw daily for a week. Every week an additional dose of 3 mg Cu/kg bw was included in the drench until signs of copper poisoning appeared; the control sheep were drenched with saline solution. The onset of copper poisoning occurred between 42 and 55 d. Food intake and body weight were recorded daily. Blood samples were collected weekly to measure the activity of the liver enzymes gamma-glutamyltransferase (gammaGT), aspartate aminotransferase (AST), sorbitoll dehydrogenase (SDH) and acid phosphatase (AF). The following changes were significantly recorded in the CL sheep in the weeks or days previous to the hemolytic crisis: higher levels of gammaGT were found on the -28th d increasing slowly but continuously until the hemolytic crisis; SDH fluctuated during the period presenting higher levels on the -28th, -14th and -7th d; AST and AF activities increased from the -14th and -7th d respectively; sharp decreases in the activities of SDH and AF at the hemolytic crisis; lower feed intake and body weight gain from the -7th d; and sheep ceased eating concentrates from the -9th d and became anoretic the day before the hemolytic crisis. Plasma copper concentration increased only the day before the hemolytic crisis. There were no changes in respiratory and heart rates, rectal temperature or rumen movements throughout the pre-hemolytic phase. The higher the amount of cumulative copper drenched, the higher was the gammaGT and AST activities. It was concluded that gammaGT followed by AST are the best enzymes to assess copper-load in sheep during the pre-hemolytic phase. Sheep fed copper-rich diets with high plasma activity of these enzymes, decreased feed consumption and subtle loss of body weight are most likely to present with a hemolytic crisis in a few days.     

A pharmacological study of chlorphenesin carbamate. Tolerance to chlorphenesin carbamate (author's transl)]

Tolerance to chlorphenesin carbamate (CPC) was investigated from the viewpoints of action of CPC, serum free CPC concentration, the activity of UDP-glucuronyltransferase and the content of cytochrome P-450. CPC was administered once daily for 7 or 14 days. In mice, the hypnotic action of hexobarbital injected 24 hours after the last administration of CPC and the motor incoordinating action of CPC significantly decreased on the 7th day, but slightly recovered on the 14th day. Serum free CPC concentration also decreased on the 7th day, but recovered on the 14th day. A significant relationship between the motor incoordinating action of CPC and serum free CPC concentration was observed. Therefore, the recovery of CPC effect on the 14th day was considered to be due to the recovery from the induction of drug-metabolizing enzymes. On the other hand, in rats, the weekly alteration of the motor incoordinating action of CPC was similar to that observed in mice. Serum free CPC concentration on the 7th and 14th days was lower than that on the 1st day, and enzyme induction was observed during CPC administration. Notwithstanding the low level of serum free CPC concentration, the recovery of CPC effect was observed on the 14th day and such was considered to be due to habituation to the rotarod. In mice and rats, it was demonstrated that the intensity of CPC effect was dependent on serum free CPC concentration to the extent that enzyme induction played an important role in the development of tolerance. From these results, the tolerance to CPC is attributed to induction of drug-metabolizing enzymes in liver microsomes.     

Effect of rifampicin on haem and bilirubin metabolism in man.

Haem and bilirubin metabolism was studied in seven healthy volunteers during 4 weeks treatment with rifampicin 600 mg at night. The serum unconjugated bilirubin concentration increased 2-3 fold in the first 24 h of treatment (P less than 0.01) and subsequently fell to below pretreatment values (P less than 0.05) during the third and fourth weeks of rifampicin therapy. In each subject, the activity in leucocytes of delta-aminolaevulinic acid (ALA) synthase increased and that of protoporphyrinogen (proto) oxidase decreased during the first week of therapy. The mean ALA synthase activity was most markedly increased on day 4 being seven-fold its pretreatment value (P less than 0.01), and the mean proto oxidase activity most depressed on day 2 at 50% its pretreatment value (P less than 0.02). There was increased urinary excretion of porphobilinogen (PBG) during the first week of therapy and increased excretion of porphyrins and PBG during the third week of treatment. The increase in ALA synthase activity and haem precursor excretion can be explained by the combination of increased haem demand for haemoprotein induction and partial block in haem synthesis due to reduced proto oxidase activity.