Different effects of fibrates on the microsomal fatty acid chain elongation and the acyl composition of phospholipids in guinea-pigs.

1. The effects in vitro and in vivo of three fibric acid derivatives, clofibrate (CFB), bezafibrate (BFB) and gemfibrozil (GFB) on some enzyme activities related to fatty acid biosynthesis, namely palmitoyl-CoA synthetase and hydrolases (microsomal and cytosolic), NADH and NADPH cytochrome c reductases and acyl-CoA elongases were investigated in guinea-pigs. 2. The three fibrates inhibited acyl-CoA elongation in vitro, irrespective of the substrate of elongation used (saturated, monounsaturated, polyunsaturated) and with an order of potency GFB > BFB > CFB. In the case of GFB, inhibition occurred at concentrations that can be reached in vivo. 3. Microsomal palmitoyl-CoA hydrolase and synthetase were also inhibited in vitro (GFB > or = BFB > CFB), whereas NADH cytochrome c reductase activity was increased by GFB. Nevertheless, the magnitude of changes were lower than those observed in elongation activities. 4. Treatment with fibrates did not produce peroxisomal proliferation in guinea-pigs, as measured by peroxisomal beta-oxidation activity and liver weight/body weight ratio. Nevertheless, fibrates provoked a reduction in plasma cholesterol and triglycerides, at least in GFB- and BFB-treated animals. 5. Fatty acid elongation was significantly modified by GFB treatment in vivo. The remaining enzyme activities studied were only slightly changed by fibrate treatment. 6. Treatment with BFB and to a lesser extent with CFB, increased the relative proportion of MUFA (palmitoleic and oleic acids) in microsomal phospholipids, whereas PUFA (mainly linoleic acid) decreased. GFB behaved differently, increasing palmitic and linoleic acids and decreasing stearic and oleic acids. The latter changes are attributable to an inhibition of elongation activity by GFB. 7. The changes observed after fibrate treatment in both rats and guinea-pigs, as they are not directly related to peroxisome proliferation, could be more reliably extrapolated to man than those observed only in rats.     

Reduction by clofibric acid of serum arachidonic acid in rats. Effect on the acyl composition of renal phospholipids.

Alterations induced by p-chlorophenoxyisobutyric acid (clofibric acid) in the composition of phosphatidylcholine and cholesterol esters in serum and their influence on the composition of phosphatidylcholine in the kidney were studied. Rats of different ages responded differently to the drug in terms of the levels of arachidonic acid (20:4) and linoleic acid (18:2) in the phosphatidylcholine and cholesterol esters in the serum. Administration of clofibric acid to 26-week-old rats for 2 weeks caused a marked decreased in the relative level of 20:4 in phosphatidylcholine and cholesterol esters in serum, whereas similar treatment of 6-week-old rats resulted in a reduction of 18:2 and, to a lesser extent, of 20:4 in serum lipids. The decrease in phosphatidylcholine that contained 20:4 in the serum of old rats was mainly due to a decrease in the concentration of stearoyl-arachidonoyl (18:0-20:4) species. The decrease in cholesterol arachidonate in serum caused by the treatment of old rats with clofibric acid seemed to be due to a reduction in the relative level of serum phosphatidylcholine containing 20:4. The marked reduction in serum lipids that contained 20:4 caused a decrease in the relative level of 20:4 in renal phospholipids, in particular, a decrease in the proportion of palmitoyl-arachidonoyl (16:0-20:4) and 18:0-20:4 phosphatidylcholine.     

Differential effects of cadmium on the hepatic microsomal cytochrome p-450 system in male and female rats

Cadmium (Cd) produced a marked sex-related difference with respect to inhibition of the hepatic microsomal monooxygenase enzyme system in the rat. Following in vivo cadmium (2 $\text{mg}\text{kg}$ i.p.) treatment, significant decreases in the levels of cytochrome P-450, significant reductions in the magnitudes of spectral binding (aniline or ethylmorphine), and significant inhibitions of microsomal metabolism (aniline and ethylmorphine) were observed with microsomes isolated from male but not female rats. Of these parameters only aniline metabolism was significantly altered in females. Following the in vitro addition of Cd (10^?6 M to 10^?3 M) to hepatic microsomes isolated from untreated male or female rats, sex-related changes were also observed in these parameters. Significant, concentration-dependent reductions were observed in cytochrome P-450 levels of both sexes but the males showed greater sensitivitiy to the cadmium effect. With respect to binding spectra, cadmium addition produced a concentration dependent inhibition of aniline only in the male rat. Ethylmorphine binding was inhibited only at the higher cadmium concentrations in both sexes. With respect to drug metabolism, cadmium addition inhibited both aniline and ethylmorphine metabolism in male rats and only aniline metabolism in female rats. These results showed that there are sex-related differences in the interaction of the hepatic microsomal monooxygenase enzyme system with cadmium both after in vitro addition as well as in vivo treatment with the metal.     

Hepatic microsomal phospholipids in rats exposed intratracheally to coal fly ash

The effects of intratracheal administration of fly ash (50 mg/kg body weight, daily for 7 days) on hepatic microsomal phospholipid metabolism has been studied in rats using various phospholipid precursors, viz NaH₂ ³²PO₄, (methyl-¹⁴C)-choline, and (methyl-¹⁴C)-methionine. Fly ash administration significantly increased microsomal phosphatidylcholine (PC), and lysophosphatidylcholine (LPC). The incorporation of NaH₂ ³²PO₄ into total liver phospholipids, PC and Phosphatidyl ethanolamine (PE) was significantly increased in fly ash-treated rats as compared to the control. Fly ash administration also increased the incorporation of (methyl-¹⁴C)-choline into microsomal PC. Incorporation of (methyl-¹⁴C)-methionine into microsomal PC was not affected. Fly ash administration decreased the per cent distribution of arachidonic acid in PC and PE and increased that of oleic acid in PC and of linoleic acid in PE.     

Difference between effects of chlorpromazine and perphenazine on microsomal phospholipids and enzyme activities in rat liver.

The effects of acute administration of chlorpromazine (CPZ) and perphenazine (PPZ) on hepatic microsomal phospholipids (PLs) and enzyme activities in the male rat were examined in order to elucidate the relationship between individual PLs and drug-metabolizing activity. Cytochrome P-450 and aniline (AN) hydroxylation activity were initially decreased in CPZ-treated rats, but cytochrome P-450 subsequently recovered to a level not significantly different from the control and AN hydroxylation was markedly increased, while in PPZ-treated rats, they remained depressed. CPZ increased the activities of glycerophosphate acyltransferase (GPA) and choline phosphotransferase (CPT), while PPZ increased the activities of phosphatidate cytidylyltransferase (PCT), phosphatidate phosphohydrolase (PPH) and CPT. Concurrently, CPZ raised microsomal phosphatidylcholine (PC), phosphatidylethanolamine (PE), phosphatidylserine-inositol (PSI) and sphingomyelin (SM), while PPZ increased PC and PE, but did not affect the levels of PSI and SM. Acyl components of phospholipids were also modified. CPZ significantly decreased the ratio of saturated to unsaturated fatty acids, particularly in the PC and PE fractions, while the effect of PPZ was only slight. The results imply that an increase of AN hydroxylation activity may involve the incorporation of unsaturated fatty acids into enzyme-associated PC and PE.     

Differential effects of 3 dipyridyl isomers on hepatic microsomal cytochrome P450 and heme oxygenase in rats

We compared the effects of 3 dipyridyl isomers, 2,2′-dipyridyl, 2,4′-dipyridyl and 4,4′-dipyridyl, on hepatic microsomal heme oxygenase and drug-metabolizing enzyme activities in male rats. 2,2′-Dipyridyl increased cytochrome P450 (P450) content at lower doses, but decreased with increasing dose levels. Immunoblot analysis revealed that 2,2′-dipyridyl did not induce both P450 1A1/2 and P450 2B1/2, in contrast to 2,4′- and 4,4′-dipyridyls, both of which were inducers of either P450 1A1/2 and/or P450 2B1/2. Some drug-metabolizing enzyme activities gradually declined with the increasing dose level of 2,2′-dipyridyl. 2,2′-Dipyridyl was able to induce hepatic microsomal heme oxygenase in a dose-dependent manner, but 2,4′- and 4,4′-dipyridyls did not, even at the highest dose (0.80 mmol/kg) examined. Treatment of rats with 2,2′-dipyridyl resulted in the increase of glutathione (GSH) content in a dose-dependent manner, but not 4-substituted isomers. A time course study with 2,2′-dipyridyl revealed that it produced a significant decrease in hepatic GSH content at early time periods (2–6 h) after its administration with an inverse increase in heme oxygenase activity. The present investigation has revealed that in contrast to the induction of P450 by 4-substituted dipyridyl compounds, 2,2′-dipyridyl is a novel inducer of hepatic microsomal heme oxygenase, together with the change in hepatic GSH content. This study would provide information on the differential effects of simple dipyridyl isomers on hepatic enzymes involved in heme and drug metabolism.     

The role of microsomal phospholipids and their fatty acid composition in the control of hepatic metabolism of lignocaine

1. Sex differences exist in the metabolism of lignocaine by the rat liver. Microsomal phospholipids have been implicated in the control of these sex differences. Induction of diabetes in the male rat abolishes these sex differences. The difference in drug metabolism between the male and female rat is, thus, the same as that between the control and diabetic male rat. 2. By using reconstitution of delipidated male microsomal proteins with male-, female- and diabetic-derived phospholipids as well as synthetic phospholipids, it should be possible to delineate the role of phospholipids in the control of drug metabolism. 3. Female- and diabetes-derived phospholipids decrease the activity of the male-specific lignocaine N-deethylase specifically by between 35 and 52%. 4. Analysis of the phospholipid classes and fatty acid content of the various fractions indicated that stearic acid content was increased and arachidonic acid content decreased in both female- and diabetic-derived lipids as compared to control males. Linoleic acid content was decreased in female- but increased in diabetic-derived lipids as compared to control males. Subsequent correlation to N-deethylase activity, however, rules out all but the arachidonic acid content of the phospholipids as a controlling factor of lignocaine metabolism. 5. Use of synthetic phosphatidylcholine (PC) species for reconstitution indicates that diarachidonyl-PC is the most efficient at activating the N-deethylase and indicates that the degree of unsaturation of the fatty acyl side-chains of PC is of major importance in the regulation of this enzyme activity.(ABSTRACT TRUNCATED AT 250 WORDS)     

Differential action of progesterones on hepatic microsomal activities in the rat.

A significant reduction was found in the activity of drug-metabolizing enzymes (aminopyrine N-demethylase and coumarin 3-hydroxylase) and glucose 6-phosphatase in hepatic microsomes after the administration of reduced derivatives of progesterone (5α-pregnane-3β,-ol-20-one, 5β-pregnane-3α-ol-20-one, 5α-pregnane-3β,20β-diol and 5β-pregnane-3α,20α-diol) to rats. These steroids slightly raised inosine diphosphatase activity. On the other hand, 16α-hydroxyprogesterone and pregnenolone-16α-carbonitrile significantly increased drug metabolism and slightly elevated glucose 6-phosphatase. The contrasting action of the different progesterone derivatives was associated with changes in microsomal phospholipid synthesis. Pregnanolone and pregnanediol significantly decreased the de novo incorporation of [¹⁴C-Me]-l-methionine into microsomal phospholipids, mainly manifesting in phosphatidylcholine, phosphatidylethanolamine and lysophosphatidylcholine fractions; reduced the activity of S-adenosyl-l-methionine:microsomal-phosphatidylethanolamine methyl transferase; and caused a reduction of total microsomal phosphatidylcholine:phosphatidylethanolamine ratio. In contrast, 16α-hydroxy-progesterone and pregnenolone-16α-carbonitrile increased the de novo synthesis of microsomal phospholipids, methyl transferase activity and the ratio of total microsomal phosphatidylcholine: phosphatidylethanolamine. Treatment of rats with reduced progesterone derivatives diminished microsomal progesterone hydroxylation in the 16α- and 6β-position and raised progesterone Δ⁴-5α-dehydrogenase activity measured in vitro. On the other hand, 16α-hydroxyprogesterone and pregnenolone-16α-carbonitrile elevated progesterone hydroxylation. Considering these opposite effects it can be postulated that in the rat the induction of drug-metabolizing activity of the hepatic endoplasmic reticulum might be controlled by a balance displayed in the synthesis and metabolism of various progesterone derivatives.     

Differential effects of diabetes on microsomal metabolism of various substrates: Comparison of streptozotocin and spontaneously diabetic wistar rats

We have examined the effect of recent onset diabetes on several aspects of hepatic microsomal metabolism in both streptozotocin (STZ)-induced and spontaneously diabetic BioBreeding (BB) male and female Wistar rats. Differential alterations of the diabetic state on hepatic microsomal enzyme activities were observed. Female diabetic rats exhibited no change in benzo[a]pyrene (BP) hydroxylase activity, a decrease in testosterone Δ⁴-hydrogenase, and an increase in aniline hydroxylase. On the other hand, male diabetic rats demonstrated a decrease in hepatic BP hydroxylase activity, no change in testosterone Δ⁴-hydrogenase, and an increase in aniline hydroxylase. Insulin treatment corrected these effects. No change in kidney BP hydroxylase activity was apparent in either female or male diabetics. There were no marked differences between the chemically induced and genetic models of diabetes with respect to the metabolism studies. Serum testosterone levels were significantly lower than control in male BP diabetics, whereas no change was apparent in female diabetics. Light microscopy and serum insulin determinations indicated that the spontaneously diabetic animals we examined were not severely diabetic. From electrophoresis of hepatic microsomal proteins we determined that spontaneous diabetes of short duration does alter the protein distribution in the cytochrome P-450 region. We conclide that the acute effects of STZ-induced and spontaneous diabetes on hepatic microsomal metabolism are quantitatively and qualitatively similar, despite probable differences in etiology of the diabetic state.     

Differential effects of d- and l-amphetamine on mouse-killing behavior in rats.

Muricidal behavior in rats was selectively antagonized by both the d- and the l-isomer of amphetamine. However, d-amphetamine was approximately 8 times as potent as l-amphetamine as an inhibitor of mouse killing. The results of this study suggest that amphetamine antagonizes muricidal behavior in rats primarily via noradrenergic mechanisms. In addition, these results, as well as those in previous reports, imply that agents which modify the level of activity at central noradrenergic receptors may significantly alter the mouse-killing response of rats.     

Effects of DI-(2-ethylhexyl) phthalate on the content and composition of hepatic mitochondrial and microsomal phospholipids in the rat

Feeding a 20% casein diet containing di-(2-ethylhexyl) phthalate (DEHP) at a 0.5% level to young male rats for 7 days resulted in a significant increase in hepatic phospholipids (PL), based either on per unit weight of the liver or on protein. Although the concentration of PL increased in both the mitochondrial and the microsomal fractions, the magnitude of the increase was much more marked in the former. The increase of PL in hepatic microsomal and mitochondrial fractions was attributed to increases in phosphatidylethanolamine (PE) and phosphatidylcholine (PC). In terms of percentage composition, PE increased significantly, whereas PC remained unchanged, leading to an elevation in the PE/PC ratio in both fractions. A similar response was observed in rats fed 0.1 to 0.5% DEHP for 30 days. Although hepatic PL accumulation was observed in rats fed a diet containing different dietary levels of casein and corn oil, the extent of the increase was much greater on a low protein diet. DEHP caused a decrease in the concentration of hepatic triglycerides (TG), and the magnitude of the reduction appeared to be greater in rats fed diets containing zero or low levels of essential fatty acids. The fatty acid profiles of PE and PC were modified differently by DEHP. Of interest was a significant increase in arachidonic acid in PE and a decrease in PC in two subcellular fractions examined. The rate of swelling of isolated mitochondria from the livers of rats fed DEHP was markedly slower than that of the controls. Some structural changes were also observed by electron microscopy.     

Differential effects of hepatic microsomal enzyme inducing agents on liver blood flow

Male and female rats were injected i.p. for 5 days with phenobarbitone (80 mg/kg/day), antipyrine (80 mg/kg/day), phenytoin (65 mg/kg/day) or chlordiazepoxide (40 mg/kg/day). Seven days after start of treatment, radioactive microspheres were used to determine liver blood flow in some animals from treatment groups. Other animals were used to measure liver microsomal protein, cytochrome c reductase and cytochrome P450. Pentobarbitone sleeping time was also determined. In males, all the drugs significantly increased cytochrome P450 content and liver weight/100 g body weight (bw) relative to salinetreated controls. Also, pentobarbitone sleeping time was significantly decreased by all 4 drugs. However, only phenobarbitone changed liver blood flow; liver weight was increased by 23 per cent and this was paralleled by a 32 per cent increase in liver blood flow/100 g bw. Female rats receiving saline had lower liver cytochrome P450 contents and longer pentobarbitone sleeping times than the control males. In addition, the drugs were less potent in the females; all significantly reduced sleeping time but liver weight/100 g bw and cytochrome P450 content were only significantly increased by phenobarbitone and phenytoin. After phenytoin, antipyrine and chlordiazepoxide, liver blood flows/100 g bw were within 4 per cent of the control value whereas with phenobarbitone there was a 9 per cent increase which accompanied an 11 per cent increase in liver weight/100 g bw. The dose-effect relations of phenobarbitone were determined in male rats using doses of 5, 10 and 80 mg/kg/day and some animals were given amylobarbitone (80 mg/kg/day) in order to see if it also changed liver blood flow. Phenobarbitone gave dose-dependent effects on the biochemical parameters, liver weight; 100 g bw and liver blood flow/100 g bw. Amylobarbitone was much less potent than phenobarbitone but did cause parallel changes in liver blood flow and liver weight/100 g bw. It is concluded that, of all the hepatic microsomal enzyme inducing agents which have been studied, only-barbiturates increase both liver blood flow and liver weight.     

Differential effects of octreotide on motor responses to nicotine in rats.

We tested the hypothesis that brain somatostatin levels modify two motor behaviors evoked by ICV infusions of nicotine. Unrestrained, awake rats were given fixed-concentration infusions of nicotine until the prostration/immobility (PI) syndrome and convulsions were produced. Infusion duration ranged from 0.9 to 1.2 min for the PI syndrome and 2.5 to 4.9 min for the convulsions. Octreotide, a stable somatostatin analog (4.5 micrograms, ICV), significantly raised the threshold for nicotine convulsions 1.0 and 5.5 h after pretreatment but not at 24 or 48 h. Cysteamine, a somatostatin releaser and depletor (0.35-0.75 mg/rat, ICV), also caused a dose-dependent increase in seizure threshold. Similarities in the response to octreotide and cysteamine suggest that depression of nicotine convulsions by cysteamine may be mediated by release of endogenous somatostatin. Neither octreotide nor cysteamine altered the threshold for the PI syndrome. These results support the view that one motor behavior evoked by nicotine is subject to control by somatostatin whereas another is not.     

Differential effects of intrastriatal estradiol on the dorsal immobility response in male rats.

The effects of implants of 17 beta-estradiol and cholesterol in four regions of the dorsal striatum were tested on the duration of the dorsal immobility response in gonadectomized male rats. The dorsal immobility response was significantly potentiated by 4-h implants of 17 beta-estradiol in the dorsomedial and dorsolateral regions of the dorsal striatum but not in the ventromedial and ventrolateral regions. These data further support the growing evidence that estradiol acts directly but differentially on the striatum to affect behaviors in the rat.     

Differential effects of CCK-JMV-180 on food intake in rats and mice.

Boc-Tyr(SO3)-Nle-Gly-Trp-Nle-Asp-2-phenylether ester (CCK-JMV-180) has been reported to be a CCK-based heptipeptide with novel in vitro properties. Based on studies conducted in rat and mouse pancreatic acini, it has been proposed that the compound acts as an agonist at the high-affinity site and an antagonist at the low-affinity site in the rat, but as an agonist at both sites in the mouse. In the present study, we examined the effects of CCK-JMV-180 on locomotor activity in the rat and on intake of a liquid diet in the rat and mouse. Although CCK-JMV-180 slightly reduced activity on its own in the rat, it completely reversed the suppression produced by coadministration of CCK-8. In rat feeding studies, CCK-JMV-180 failed to suppress intakes of a liquid diet, but was able to antagonize the anorectic effects of CCK-8. In contrast, in the mouse CCK-JMV-180 potently suppressed intakes on its own, and this effect was blocked by pretreatment with the selective CCK-A receptor antagonist, A-70104. The results of these studies suggest that similar receptor mechanisms are involved in CCK's ability to inhibit food intake in vivo and its effects on pancreatic function in vitro.     

Effect of phenobarbital on methyl transfer between methylated drugs and hepatic microsomal phospholipids.

The effect of phenobarbital on the incorporation of the label from N-[¹⁴C-Me]nicotine and [¹⁴C]formaldehyde into hepatic phospholipids of the rat has been studied. ¹⁴C was utilized for the formation of methylated phospholipids from both precursors. Phenobarbital elicited no significant action either on the synthesis of total hepatic phospholipids or on the incorporation of radioactivity into the total or individual liver phospholipid fractions. However, this treatment increased phospholipid content and the uptake of the label from nicotine into microsomal phospholipids. Phenobarbital raised microsomal phosphatidylethanolamine, -choline (PC), -serine (PS), and lysophosphatidylcholine contents and the incorporation of ¹⁴C-labeled methyl groups from nicotine into PC and PS fractions. Radioactivity from [¹⁴C]formaldehyde was also incorporated into hepatic phospholipids. Phenobarbital however, had no significant effect on the incorporation either into total or microsomal phospholipids. Comparing the utilization of ¹⁴C for synthesis of liver microsomal phospholipids from N-[¹⁴C-Me]nicotine or [¹⁴C]formaldehyde with the natural methyl donor, l-[¹⁴C]-Me]methionine, greater amounts were taken up from methionine than nicotine or formaldehyde. The methyl group of nicotine was probably incorporated into phospholipids via the metabolic pool; the enhancing effect of phenobarbital on this process was associated with increased metabolism and with increased methyl transfer into methyl group containing microsomal phospholipids.