Effects of ethanol on microsomal drug metabolism in aging female rats. II. Inhibition

The effect of aging on the inhibition by ethanol of drug metabolism activity was examined in liver microsomes of female Fischer 344 rats aged 4, 14 and 24 months. Inhibition of aniline hydroxylase activity in microsomes from 4-month-old females occurred at low concentrations of ethanol (0.1 mM) and was predominantly competitive. Aging was associated with a significant increase in apparent Km for aniline in the absence of ethanol (24 +/- 2, 20 +/- 2 and 32 +/- 1 microM in microsomes from 4-, 14- and 24-month-old rats, respectively) and a change from competitive to non-competitive inhibition by ethanol. Inhibition of benzphetamine N-demethylase activity occurred only at high concentrations of ethanol (100 mM) and was non-competitive in nature. There were no significant effects of aging on the kinetics of the reaction or the type of inhibition produced by ethanol. Microsomal ethanol oxidation rates were measured in liver microsomes of 4-, 15- and 25-month-old Fischer 344 rats of both sexes. Ethanol oxidation in males was greater than in females and was decreased significantly in old age. Ethanol oxidation in female rats was unaffected by aging. The results suggest that significant changes in drug/ethanol interactions can occur as a consequence of aging.     

Effects of ethanol on microsomal drug metabolism in aging female rats. III. In vivo

The effects of aging on ethanol inhibition of zoxazolamine metabolism in vitro and in vivo were studied in female Fischer 344 rats aged 4, 14 and 26 months. Zoxazolamine hydroxylase activity in freshly-isolated liver microsomes decreased significantly with age (1.88 +/- 0.32, 1.49 +/- 0.30 and 0.74 +/- 0.18 nmol/min per mg protein in young-adult, middle-aged and old rats, respectively). A substantial inhibition of zoxazolamine hydroxylation occurred in the presence of 40 mM ethanol. The extent of inhibition was the same in microsomes from all three age groups. The effect of aging on the duration of zoxazolamine paralysis in vivo reflected the effect of aging on zoxazolamine metabolism in vitro. Mean duration of paralysis following a standard 50 mg/kg dose of zoxazolamine increased significantly as a function of aging (0.5, 2.9 and 4.7 h in young-adult, middle-aged and old rats, respectively). Administration of ethanol (1.2 g/kg) 10 min before zoxazolamine treatment prolonged the duration of zoxazolamine paralysis in young-adult and middle-aged rats by about 2 to 2.5 h, but ethanol pretreatment did not affect paralysis time in old rats. Thus, the inhibitory effect of ethanol on zoxazolamine metabolism in vivo appeared to be attenuated in old age.     

Hepatic drug metabolism during development in food-restricted female Sprague-Dawley rats.

Hepatic drug metabolism was investigated in female Sprague-Dawley rats fed ad libitum (A) or a restricted diet (R) (implemented from age 1 month), at 1.5, 4.5 and 12 months to determine the short- and long-term effects of caloric restriction. Microsomal cytochrome P-450 content and NADPH cytochrome c reductase activity were not modified by age. While dietary restriction did not affect cytochrome P-450, it significantly increased NADPH cytochrome c reductase activity at all time periods when compared to corresponding A-fed groups. Aniline hydroxylase and aminopyrine N-demethylase activity tended to decrease with age in the A-fed groups but the differences did not prove to be statistically significant. A significant decrease of aminopyrine N-demethylase was observed with age in R rats. A significant reduction of aniline hydroxylase activity was noted in the R groups compared to age-matched A-fed controls. In contrast, aminopyrine N-demethylase activity increased significantly, but only at 1.5 months of age. Glutathione S-transferase activity was augmented between 1.5 and 4.5 months of age, and this was followed by a significant decrease at age 12 months in both A and R groups. Dietary restriction had no effect on this enzymatic activity. The microsomal cholesterol and phospholipid content as well as the cholesterol/phospholipid molar ratio changed significantly between 1.5 and 4.5 months of age but not between 4.5 and 12 months of age. These parameters were unaltered by dietary restriction. In conclusion, in the female Sprague-Dawley rat there are no statistically significant changes in hepatic microsomal components and drug metabolizing capacity between 1.5 and 12 months of age. Dietary restriction resulted in significant changes in enzymes related to drug metabolism which varied with the enzyme examined. In general, these changes were similar after short- or long-term dietary intervention.     

Induction of an auto-anti-IgE response in rats I. Effects on serum IgE concentrations

With a view to specifically suppressing the IgE isotype, rats of high (BN) and low (PVG.RT1u) IgE-responding phenotypes were immunized with a highly purified rat IgE myeloma (IR2) in an attempt to induce an anto-anti-IgE response. Rat IgE antibodies against epsilon determinants were detected in the serum of IR2-immunized animals using a solid-phase (plate) radioimmunoassay. The auto-anti-IgE antibodies detected were found to bind to IR2, to a second rat IgE myeloma (IR162) and to mouse monoclonal IgE but not rat IgG. The specificity of the anti-epsilon binding was shown by inhibition studies. The raising of an auto-anti-IgE response in PVG.RT1u rats severely depleted the serum level of circulating IgE for at least 8 weeks. In BN rats, immunization with IR2 caused marked fluctuations in serum IgE levels. The rats in both strains remained healthy throughout the experiment. The rate and route of IgE break down was not altered in anti-IgE-producing rats. The relevance of the present model in understanding and possibly controlling allergic disorders is considered.     

Effects of ovariectomy and steroid replacement on hypothalamic LHRH content in aging female rats

To examine the role of age on the hypothalamic LHRH response to ovariectomy (ovx) and steroid replacement, young cycling (3-4 months) and old constant estrous (18-20 months) rats were ovariectomized. Two weeks later, rats were treated for 5 days with estradiol benzoate (E, 5 micrograms/kg), progesterone (P, 10 mg/kg), E + P (5 micrograms E + 10 mg/kg) or corn oil, after which time they were killed for determination of hypothalamic LHRH content. The young and old ovx rats had similar levels of LHRH in the medial basal (MBH) and anterior (AH) hypothalamus, but E treatment was only effective in increasing MBH-LHRH content in the young animals. There was no significant effect of P alone or in combination with E. In the second experiment, similar results were seen using a single dose of E (1 microgram/rat) in young and old ovx rats. In addition, the radioimmunoassay of LHRH using two different antibodies binding to different portions of LHRH gave similar results in young and old rats, suggesting that the LHRH peptide was being processed similarly in the two age groups. In conclusion, it appears that hypothalamic LHRH content of young and old ovx rats does not differ with age despite a marked attenuation of serum LH levels with age. The hypothalamus from old rats, however, is less responsive to steroid stimulation of LHRH content.     

Effects of estradiol and estradiol-progesterone treatment on gonadotropin secretion in castrate aging female rats

Estradiol-17β and estradiol-17β-progesterone were administered to aging female rats immediately after castration. Estradiol (1.6 μg) alone suppressed the castration hypersecretion of FSH in mature animals to 70% of the castration controls, but in 360-day-old and older animals, the inhibition was less than 10%. There was a decrease in the castration hypersecretion of LH by exogenous estradiol-17β with increasing age, but the pattern of suppression was similar in all animals, regardless of age. When a constant dose of estradiol-17β of 0.4 μg/kg body wt. was administered to castrate rats with varying doses of progesterone, it was shown that after 270 days of age the negative feedback effect of these steroids on FSH secretion was not found. Whereas. when progesterone of varying doses was administered with a constant dose of 0.8 μg/kg BW of estradiol-17β, it was shown that after 180 days of age the positive feedback effect of these steroids on FSH was absent. After 270 days of age the augmentation of LH secretion by estradiol-17β and a middle dose of progesterone was not apparent even though the negative feedback on LH was effective. A defect in positive and negative feedback effects of gonadal steroids on gonadotropin secretion may be causative to age-related changes in the estrous cycle of rats.     

Effects of maleate on carbohydrate metabolism in rats.

Intraperitoneal administration of maleate produced an increase in blood alpha-ketoacid, acetoacetate, and free fatty acids. The effect of this treatment on blood glucose levels depended on whether the rats were fed or fasted. In fed rats it was accompanied by slight, transient hyperglycemia connected with depletion of liver glycogen stores. In fasted animals moderate hypoglycemia was observed. The in vivo conversion of various precursors into blood glucose was not inhibited, suggesting that maleate does not affect hepatic gluconeogenesis. Neither was a direct effect on liver glycogenolysis observed. On the other hand, maleate inhibited renal gluconeogenesis from various substrates and stimulated anerobic glycolysis in kidney cortical alices. The data are interpreted in terms of increased utilization and decreased production of glucose by the kidney followed by secondary changes in liver carbohydrate metabolism.     

In vivo and in vitro inhibition of hepatic microsomal drug metabolism by ketoconazole.

Ketoconazole (KC), a broad spectrum antifungal drug, has been recognized recently as a cause of hepatic injury. The mechanism of the adverse reaction remains unclear: a metabolic idiosincrasy has been suggested. However as a substituted imidazole, KC might be expected to interfere with the hepatic microsomal mixed function oxidases. Ethylmorphine N-demethylase (E-DM) and aniline hydroxylase (A-OH) activities were determined in rat liver microsomes in the presence of increasing amounts of KC. Both were inhibited in an exponential fashion. The E-DM inhibition was almost complete at concentrations greater than 250 microM and was of the mixed type. A much weaker effect was observed for A-OH. A significant inhibition of E-DM was also observed when KC was administered in vivo to rats either orally for 7 days at the dose of 100 mg/kg/day (P less than 0.02) or intraperitoneally for 4 days at the dose of 50 or 100 mg/kg day (P less than 0.01 or P less than 0.001 respectively). A-OH activity was significantly reduced (P less than 0.01) only after ip administration of 100 mg/kg/day of the drug for 4 days. Neither the amount of cytochrome P-450 nor NADPH cytochrome c reductase activity were affected at the doses considered. These data show that KC interferes with hepatic oxidative drug metabolism and suggest that this mechanism might be involved in the unwanted side effects of therapy with KC.     

The effect of aging on the hepatic metabolism of sulfobromophthalein in BN/Bi female and WAG/Rij male and female rats

The effect of aging on sulfobromophthalein (BSP) metabolism was studied in three groups of rats—BN/Bi female and WAG/Rij male and female rats—of different ages ranging from 3 to 30 months. Under Nembutal anesthesia, BSP biliary transport maximum (T_m) and relative storage capacity (S) were determined by a single infusion rate method by directly determining T_m from bile samples collected through a common bile duct cannula. T_m values expressed as μg of BSP per min per g of liver were highest in the youngest rats (3-month-old) as compared with the older rats (12-, 24-, 30-month-old) for all three rat groups. T_m gradually decreased as age increased and at the age of 24 or 30 months reached a value of 66–70% of the highest values for 3-month-old rats. The percentage of conjugated BSP in the bile measured during the T_m period remained essentially unchanged with age in all three rat groups. S values, expressed as mg of BSP stored per mg of BSP per ml of plasma per g of liver, remained unchanged (BN/Bi female) or even increased (WAG/Rij male and female) with age. As a consequence, S values expressed per rat were higher in older age groups than in the youngest one for all three rat groups. In contrast with previous reports by other authors on man and rats, the BSP T_m appears to decraese with age regardless of rat strain and sex, while S does not show such a decrease.     

Passive air-pouch anaphylaxis in rats. I. Induction of anaphylaxis

Induction of an experimental passive anaphylaxis of the air-pouch type, passive air-pouch anaphylaxis, was carried out in an attempt to induce a reproducible anaphylaxis model suitable for quantitative studies. Rats were injected subcutaneously with 10 ml of air into the dorsal skin to make an air-pouch and with 2 ml of antiserum at an appropriate dilution for passive sensitization, and then 5 ml of air was removed. The challenge with 5 ml of antigen solution into the air-pouch 48 h later provoked mast cell degranulation and increased vascular permeability induced by released histamine. Treatment with monovalent hapten prior to the antigen challenge almost completely inhibited histamine release and plasma exudation to levels similar to those in the nonsensitized group. In this model, mast cell-dependent late-phase allergic reaction, such as leukocyte migration or the increase of plasma exudation following mast cell degeneration, was not observed.     

Effects of aging and phenobarbital on the rat liver microsomal drug-metabolizing system

Significant declines in the non-induced activities of liver microsomal drug-metabolizing enzymes and in the amount of cytochrome P-450 occur between maturity (16 months) and senescence (27 months) in male Fischer 344 rats, whereas there are essentially no differences between very young (1 month) and mature animals. Several hepatic responses to chronic phenobarbital administration also demonstrate marked age-dependent changes. The livers of young and mature animals exhibit: (1) greater hepatomegaly; (2) faster rates of induction and post-induction recovery of microsomal mixed function oxidase enzyme activities and hemoprotein concentration; and (3) higher maximally induced levels of these components in comparison to senescent rats. When considered with information from previous studies, the present data suggest that the age-related decline in liver drug metabolism may be due to qualitative and/or quantitative changes in the structural and/or functional components of the hepatic microsomal mixed function oxidase system.     

Effects of ethanol consumption on maternal behavior in the female rat

Female rats consuming 30% ethanol (v/v) for 30 days prior to and during gestation, and during the post-partum period were tested for maternal behavior with either their own pups or pups provided by normal foster mothers. When tested with their own pups, ethanol consuming females displayed significant deficits in maternal behavior. However, when a second group of ethanol consuming females were tested with normal pups, the females displayed maternal behavior comparable to that of control females. These data suggest that gestational ethanol consumption per se does not produce a deficit in the ability to display maternal behavior. Possibly, due to ethanol related changes in the pup's own characteristics, they became a less attractive stimulus in eliciting the display of maternal behaviors from their mothers.     

Differential effects of swimming and running on microsomal metabolism in middle-aged and aged Fischer 344 rats.

Cytochrome P-450 (P-450) content as well as p-nitroanisole (pNA) O-demethylase and UDP-glucuronyltransferase (UDPGT) activities were determined in livers of middle-aged (MA; 12 or 18 months) and aged (24-26 months) rats exercised by either treadmill running or swimming. In addition, aniline hydroxylase activity was measured in MA runners and aged swimmers and compared to respective sham and non-handled controls. Treadmill exercise consisted of running aged and MA rats on a motorized treadmill for 16 and 20 m/min respectively, 60 min/day and 4 times per week, for 8 weeks. Sham rats were placed on the treadmill twice per week for 5 min at 8 m/min. No differences were found in any parameter comparing sham rats to non-handled controls. Running did not affect body weight or hepatic microsomal protein during the 8-week study. A 33-35% decline in microsomal P-450 content in treadmill exercised MA and aged rats was found. PNA O-demethylase activity was decreased 30% in MA and 45% in aged runners and aniline metabolism was inhibited 21% in MA rats. UDPGT activity was not affected by running in MA or aged rats. Swimming exercise was accomplished by placing the rats in a tank of water (32-33 degrees C) filled to a depth of 2 ft. Swim time was 60 min twice daily, 5 times per week. The aged and MA rats were trained for 6 months and 1 year, respectively. Two control groups, non-swimming sedentary (dry control) and 1 min swim/day sham (wet control), were utilized. MA and aged wet controls and swimmers weighted 8% and 15% less respectively, than MA and aged sedentary rats. Microsomal protein was significantly increased in MA swimmers compared to sedentary (20%) and wet control (35%) but no change was found with swimming in the aged rats. The results of the enzymatic studies were variable in the MA rats. Increases in P-450 content were found in wet controls (16%) and swimmers (27%) of the MA group, but only the swimming change was significant. No significant change was determined for pNA metabolism between swimmers and wet (22%) or dry (17%) controls. Aged swimmers and wet controls were more consistent, with no change in any of the parameters except aniline metabolism which was significantly increased in wet controls (25%) and swimmers (28%) as compared to dry controls. No significant change in UDPGT activity was measured in either age group of swimmers.(ABSTRACT TRUNCATED AT 400 WORDS)     

Effects of dietary fat on drug metabolism

The fact that nutriture affects drug metabolism and drug action in laboratory animals is undisputable. Activation or detoxification of drugs and potential carcinogens can also be modified by diet. The quantity and quality of dietary fat affects lipid composition and physical characteristics of biological membranes and enzymatic activity of several components of the drug metabolizing enzyme system. These changes have been associated with alterations in the physiological response to drugs and to the resulting mutagenicity and carcinogenicity of procarcinogens. It is suggested by these data that dietary fat, by altering fatty acid composition of biological membranes, alters the physical and biochemical characteristics of these membranes, thereby directly affecting drug entrance into the membrane; the stability of the membrane; the potential for lipid peroxidation; and the activity of the phospholipid dependent enzymes associated with these membranes. The fact that these membrane associated changes can occur rapidly and that brief periods of fatty acid deprivation can profoundly affect the inducibility of these enzymes by xenobiotics suggests that the potential for drug-nutrient interactions exists in the absence of frank nutrient deficiency states.     

Effects of a diphophonate on calcium metabolism in calcium-deprived rats.

Calcium metabolism was studied in growing rats, submitted to calcium deprivation ofvarious intensity. A decreased intake resulted in decreased net absorption of calcium(V'na), no change in bone formation (V'o+), and an increase in bone resorption (V'o -). In animals given dichloromethylene disphosphate (Cl'2MDP), a compoundknown to inhibit bone resorption, V'o+ was less than in the controls but again the same at all calcium intakes; V'na was below V'o+, V'o- still increased as the calcium intake was reduced. The various kinetic parameters in rats receiving Cl'2MDPwere indistinguishable from published data in parathyroidectomized (PTX) animals, yetblood calcium was low in PTX rats but normal in Cl'2MDP-atreated rats. It appears that the rat has an efficient mechanism for increasing bone resorption which is not inhibited by Cl'2MDP and does not require parathyroid hormone.