Acid Levels In Rats Research Articles

Uric acid and chronic kidney disease: which is chasing which?

Serum uric acid is commonly elevated in subjects with chronic kidney disease (CKD), but was historically viewed as an issue of limited interest. Recently, uric acid has been resurrected as a potential contributory risk factor in the development and progression of CKD. Most studies documented that an elevated serum uric acid level independently predicts the development of CKD. Raising the uric acid level in rats can induce glomerular hypertension and renal disease as noted by the development of arteriolosclerosis, glomerular injury and tubulointerstitial fibrosis. Pilot studies suggest that lowering plasma uric acid concentrations may slow the progression of renal disease in subjects with CKD. While further clinical trials are necessary, uric acid is emerging as a potentially modifiable risk factor for CKD. Gout was considered a cause of CKD in the mid-nineteenth century, and, prior to the availability of therapies to lower the uric acid level, the development of end-stage renal disease was common in gouty patients. In their large series of gouty subjects Talbott and Terplan found that nearly 100% had variable degrees of CKD at autopsy (arteriolosclerosis, glomerulosclerosis and interstitial fibrosis). Additional studies showed that during life impaired renal function occurred in half of these subjects. As many of these subjects had urate crystals in their tubules and interstitium, especially in the outer renal medulla, the disease became known as gouty nephropathy. The identity of this condition fell in question as the presence of these crystals may occur in subjects without renal disease; furthermore, the focal location of the crystals could not explain the diffuse renal scarring present. In addition, many subjects with gout also had coexistent conditions such as hypertension and vascular disease, leading some experts to suggest that the renal injury in gout was secondary to these latter conditions rather than to uric acid per se. Indeed, gout was removed from the textbooks as a cause of CKD, and the common association of hyperuricemia with CKD was solely attributed to the retention of serum uric acid that is known to occur as the glomerular filtration rate falls. Renewed interest in uric acid as a cause of CKD occurred when it was realized that invalid assumptions had been made in the arguments to dismiss uric acid as a risk factor for CKD. The greatest assumption was that the mechanism by which uric acid would cause kidney disease would be via the precipitation as crystals in the kidney, similar to the way it causes gout. However, when laboratory animals with CKD were made hyperuricemic, the renal disease progressed rapidly despite an absence of crystals in the kidney. Since this seminal study, there has been a renewed interest in the potential role uric acid may have in both acute and CKD. We briefly review some of the major advances that have occurred in this field in the last 15 years.

The Effect of the Serum Amino Acid Levels Thiosemicarbazone Derivatives and its Metal Complexes on Rats

Advers biological activities of Thiosemicarbazone (TSC) and Schiff base (SB) derivatives have been widely studied in rats and in other animal species using different doses, times and routes of administration. To date, no attempt has been made to study alterations occurring in the amino acid profile in the effects of the thiosemicarbazone derivative and its metal complexes on the rats. At this study, the rats were injected subcutaneously with a new thiosemicarbazone and its LH-Zn and LH-Cu complexes (25 mg/kg body weight) and then sacrified after 15th days. The aim of this study was to determine the effect of new compounds on the serum amino acid levels in rats. In the comparision done among groups, it was observed that serum amino acid concentrations were statistically changed (p < 0.05).

Dose-dependent effects of diallyl disulfide on plasma glucose and free fatty acid levels in rats

The acute effects of oral administration of diallyl disulfide (DADS), the major organosulfur compound of garlic, on plasma glucose and free fatty acid (FFA) concentrations were examined in rats. Male, 10-week-old Sprague-Dawley rats were divided into DADS-free and DADS-administered (dose = 10, 20, and 40 mg/kg body weight [BW]) groups. Plasma samples were prepared from whole blood drawn from the tail vein 0, 1, 2, 4, and 6 hr after administration. The stomachs were isolated, and the contents were measured 8 hr after administration. In DADS-administered groups, plasma glucose concentrations were increased in a dose-dependent manner 1 hr after the administration. The increase was transient, except in groups administered 40 mg/kg BW of DADS, in which plasma glucose levels remained significantly higher than the DADS-free levels throughout the experimental period. Similar patterns were observed in the plasma FFA concentrations, although the significant differences were lower than those observed in the plasma glucose concentrations. The gastric contents were dose-dependently elevated after DADS administration. The increase was significant when 20 or 40 mg/kg BW of DADS was administered. These results suggest that oral administration of DADS can mobilize energy substrates into the blood, although a higher dose of DADS slows gastric emptying.

The effect of barbiturate anaesthesia on the blood alpha-keto acid levels in rats and rabbits.

The effect of barbiturate anaesthesia on the blood alpha-keto acid levels in rats and rabbits.

Profile of Plasma Amino Acid Levels in Rats Exposed to Acute Hypoxic Hypoxia

The effect of acute hypoxic hypoxia on the profile of plasma amino acids in rats was studied and compared to that resulting from acute liver injury induced by giving carbon tetrachloride. In hypoxic rats exposed to 45% air in N(2) for 5h, the concentrations of branched chain amino acids, including valine, leucine and isoleucine, and aromatic amino acids such as phenylalanine and tyrosine were significantly increased as compared to those in normoxic rats. The ratio of branched-chain to aromatic amino acids (Fischer's ratio) was significantly decreased. The levels of arginine and citrulline, which are related to the urea cycle, were also depressed. Furthermore, plasma proline level was reduced in hypoxic rats. The activities of plasma marker enzymes for tissue damage remained unchanged during hypoxia, indicating that tissue injury was not induced by exposure to hypoxic conditions. We suggest that the characteristic profile of plasma amino acids and the Fischer ratio are valuable tools for understanding the pathology of acute hypoxia in the absence of systemic tissue damage.

Hydroxyl containing seleno-imine compound exhibits improved anti-oxidant potential and does not inhibit thiol-containing enzymes

Design and synthesis of organoselenium compounds with high thiol peroxidase (TPx) and low thiol oxidase (TOx) activities have been a difficult task and remains a synthetic-activity relationship dilemma. In this regard we are reporting for the first time a detail experimental data (both in vitro and in vivo) about the anti-oxidant and toxicological profile of an Imine (–N) containing organoselenium compound (Compound A). The TPx activity of Compound A was significantly higher than diphenyl diselenide (DPDS). Both Compound A and DPDS protected sodium nitropruside (SNP) induced thiobarbituric acid reactive species (TBARS) production in rats tissue homogenate with significantly higher activity observed for Compound A than DPDS (p<0.05). The Compound A also exhibited strong antioxidant activity in the DPPH and ABTS radical scavenging assays. This study reveals that an imine group close to selenium atom drastically enhances the catalytic activities in the aromatic thiol (PhSH) assay systems. The oxidation of biologically significant thiols reflects the toxicity of the compounds. However, the present data showed that treatment with Compound A at 0, 10, 25 or 50mg/kg was not associated with mortality or body weight loss. Similarly it did not inhibit α-ALA-D and Na+1/K+1 ATPase (sulfhydryl group containing enzymes) activities after acute oral treatment; rather it enhanced non-protein thiols (NPSH) concentration. The Compound A did not cause any oxidative stress as measured by TBARS production in rat's tissue preparation. Our data also indicate that exposure to Compound A did not affect plasma transaminase activities or levels of urea and creatinine in rats. Ascorbic acid is always considered a marker of oxidative stress and the reduction of its content may indicate an increase in oxidative stress. Treatment with Compound A did not alter Ascorbic acid levels in rats. The conducted in vitro and in vivo tests show the versatile therapeutic potential of this compound in the area of free radical induced damages, will undoubtedly enhance our understanding of the mechanism of model compounds and may ultimately yield insights that result in improved GPx mimics.

Could Uric acid have a Pathogenic Role in Chronic Allograft Dysfunction?

Introduction: Chronic allograft dysfunction (CAD) is the primary cause of chronic graft failure after kidney transplantation. The pathogenesis of CAD involves both antigen-dependent and antigen-independent mechanisms. Serum uric acid could have a role in both mechanisms. Review: Hyperuricemia in subjects with renal transplantation is not usually viewed as clinically significant unless the subject develops gout. Drugs used in the treatment of hyperuricemia and gout are more likely to cause side effects in the kidney transplant reciepient. However, there are recent studies that raise the possibility that uric acid could have a role in CAD. Soluble uric acid has been shown to stimulate the proliferation of vascular smooth muscle, to inhibit endothelial cell proliferation and to reduce bioactive levels of endothelial NO. Studies in experimental models have found that hyperuricemia can cause hypertension associated with renal injury characterized by microvascular disease, tubulointerstitial disease, glomerular hypertrophy and glomerulosclerosis. Hyperuricemia was also found to worsen preexistent renal disease, and to be associated with the development of severe vascular lesions that are reminiscent of those observed in CAD. Furthermore, chronic cyclosporine nephropathy is very similar to what is observed in normal rats simply by raising uric acid levels. In addition, raising uric acid levels in rats receiving cyclosporine accelerates the nephropathy whereas lowering uric acid ameliorates the renal lesions. Conclusion: The pathogenic role of uric acid in CAD is still controversial. A controlled clinical trial would be the best approach to determine the effect of uric acid lowering treatment on the development of CAD and long term graft function. Keywords: chronic allograft dysfunction, cyclosporine, uric acid, hyperuricemia, vascular disease

Sutherlandia frutescens limits the development of insulin resistance by decreasing plasma free fatty acid levels

Intake of high caloric food induces raised plasma free fatty acids, culminating in insulin resistance (IR) and Diabetes mellitus type 2 (DMT2). The present study has shown for the first time that Sutherlandia frutescens reduces plasma free fatty acid levels in rats fed a high fat diet, thereby preventing the development of insulin resistance. A commercially available S. frutescens extract was administered to rats to examine its effects on the progression of high fat diet induced IR. In comparison to rats fed high fat diet only (positive control for IR), levels of plasma free fatty acids (FFA) were significantly reduced after one week (p < 0.025). Twelve weeks of treatment with S. frutescens reduced the level of plasma free fatty acids below that of rats fed a normal diet (negative control) (p < 0.025). QUICKI and HOMA-IR index confirmed that S. frutescens treated rats did not develop IR when fed a high fat diet for twelve weeks. In addition to preventing IR and reducing plasma FFA, chronic medication over twelve weeks decreased total cholesterol levels and the LDL/HDL ratio. We propose that S. frutescens is an effective medicinal remedy to prevent elevated plasma free fatty acids and IR, and therefore DMT2.

Discovery of 3-(3-cyano-4-pyridyl)-5-(4-pyridyl)-1,2,4-triazole, FYX-051-a xanthine oxidoreductase inhibitor for the treatment of hyperuricemia

Our previous study identified 2-[2-(2-methoxy-ethoxy)-ethoxy]-5-[5-(2-methyl-4-pyridyl)-1 H-[1,2,4]triazol-3-yl]-benzonitrile ( 2) as a safe and potent xanthine oxidoreductase (XOR) inhibitor for the treatment of hyperuricemia. Here, we synthesized a series of 3,5-dipyridyl-1,2,4-triazole derivatives and, in particular, examined their in vivo activity in lowering the serum uric acid levels in rats. As a result, we identified 3-(3-cyano-4-pyridyl)-5-(4-pyridyl)-1,2,4-triazole (FYX-051, compound 39) to be one of the most potent XOR inhibitors; it exhibited an extremely potent in vivo activity, weak CYP3A4-inhibitory activity and a better pharmacokinetic profile than compound 2. Compound 39 is currently being evaluated in a phase 2 clinical trial.

Vitamin E supplementation does not prevent ethanol-reduced hepatic retinoic acid levels in rats

Chronic, excessive ethanol intake can increase retinoic acid (RA) catabolism by inducing cytochrome P450 2E1 (CYP2E1). Vitamin E (VE) is an antioxidant implicated in CYP2E1 inhibition. In the current study, we hypothesized that VE supplementation inhibits CYP2E1 and decreases RA catabolism, thereby preventing ethanol-induced hepatocyte hyperproliferation. For 1 month, 4 groups of Sprague-Dawley rats were fed a Lieber-DeCarli liquid ethanol (36% of the total energy) diet as follows: either ethanol alone (Alc group) or ethanol in combination with 0.1 mg/kg body weight of all-trans-RA (Alc + RA group), 2 mg/kg body weight of VE (Alc + VE group), or both together (Alc + RA + VE group). Control rats were pair-fed a liquid diet with an isocaloric amount of maltodextrin instead of ethanol. The ethanol-fed groups had 3-fold higher hepatic CYP2E1 levels, 50% lower hepatic RA levels, and significantly increased hepatocyte proliferation when compared with the controls. The ethanol-fed rats given VE had more than 4-fold higher hepatic VE concentrations than the ethanol-fed rats without VE, but this did not prevent ethanol induction of CYP2E1, lower hepatic retinoid levels, or hepatocellular hyperproliferation. Furthermore, VE supplementation could not prevent RA catabolism in liver microsomal fractions of the ethanol-fed rats in vitro. These results show that VE supplementation can neither inhibit ethanol-induced changes in RA catabolism nor prevent ethanol-induced hepatocyte hyperproliferation in the rat liver.

Liver glucose-6-phosphatase activity and blood fatty acid level in rats with insulin-induced hypoglycemia

Liver glucose-6-phosphatase activity and blood fatty acid level in rats with insulin-induced hypoglycemia

The prooxidant effect of sodium metabisulfite in rat liver and kidney

Sodium metabisulfite (Na 2S 2O 5) is used as an antioxidant and antimicrobial agent in a variety of drugs and functions as a preservative in many food preparations. In addition to their antioxidant activity, sulfites oxidize to sulfite radicals ( SO 3 - ) initiating lipid peroxidation. This study was performed to elucidate the effect of subchronic Na 2S 2O 5 (520 mg/kg/day) ingestion on hepatic and renal antioxidant enzyme activities and lipid peroxidation in albino rats. The antioxidant effect of l-carnitine was also tested in rats treated with Na 2S 2O 5. Plasma uric acid levels were monitored in all rats included in the study. Malondialdehyde (MDA) levels significantly increased in Na 2S 2O 5 treated rats vs. controls, with kidney values of 2.21 ± 0.21 vs. 1.22 ± 0.35 and liver values of 79.85 ± 19.5 vs. 31.36 ± 5.0 nmol/mg protein, respectively. Selenium-glutathione peroxidase (GPx) activity was significantly increased in Na 2S 2O 5 treated rats vs. controls, with kidney values of 38.22 ± 2.21 vs. 8.09 ± 0.76 and liver values of 31.11 ± 6.37 vs. 11.70 ± 1.02 U/g protein, respectively. Sodium metabisulfite treatment increased plasma uric acid levels in rats that were included in the study. No protective effect of l-carnitine was observed against lipid peroxidation in both liver and kidneys of rats treated with Na 2S 2O 5. The presented data confirm the prooxidant activity of sulfites and suggest that increased GPx activity and plasma uric acid levels may partially reduce the observed renal and hepatocellular oxidative damage caused via the ingestion of sulfites.

Vitamin A deficiency reduces liver and colon docosahexaenoic acid levels in rats fed high linoleic and low alpha-linolenic acid diet

Studies indicate that the transcription factor peroxisome proliferator-activated receptors (PPARs) regulate the activity of delta-6 and -5 desaturases and several key enzymes of peroxisomal β-oxidation, including acyl-CoA oxidase. These enzymes are vital for the synthesis of docosahexaenoic (22:6ω3; DHA) and osbond (22:5ω6, OA) acids. An activated PPAR must form a hetrodimer with the obligate cofactor retinoid X receptor (RXR) to interact with a peroxisome proliferator responsive element (PPRE) of a target gene and to regulate transcriptional expression. The vitamin A metabolite, 9- cis retinoic acid, is the most potent ligand of RXR. We have tested the possibility that deficiency of vitamin A would compromise tissue levels of both DHA and OA in rats. Two groups of male Wistar rats were randomly distributed to receive vitamin A deficient (VAD) or sufficient (VAS) diet. After seven weeks of feeding, the rats were killed and colon and liver tissues removed for the analysis of fatty acids and antioxidant status. The VAD compared to the VAS rats had elevated levels of arachidonic (AA, P<0.001), adrenic acid (22:4ω6, P<0.005) and OA ( P<0.0001) and reduced proportions of eicosapentaenoic (EPA, docosapentaenoic (DPA), DHA and total ω3 fatty ( P<0.0001) in colon choline phosphoglycerides (CPG). Similarly, liver CPG of the VAD rats had higher AA and adrenic acid and OA ( P<0.0001), and lower EPA, DPA and DHA ( P<0.0001) than the VAS rats. There was a similar fatty acid pattern in ethanolamine phosphoglycerides of the colon and liver tissues. These differences could not be explained by the conventional microsomal-peroxisomal pathway of the synthesis of the long-chain ω6 and ω3 polyunsaturated fatty acids. We postulate that deficiency of dietary vitamin A and the consequential depletion of retinoids inhibits DHA, and enhances OA, synthesis by differential effects on the independent synthetic pathways of the two fatty acids in the mitochondria. Various studies have documented that both DHA and vitamin A are vital for optimal visual and neural development and function. There is a need for further investigations to elucidate how vitamin A deficiency reduces membrane DHA level, and to delineate the synergistic effect of the two nutrients on vision, learning and memory.

Dietary linoleic acid suppresses gene expression of rat liver alpha-amino-beta-carboxymuconate-epsilon-semialdehyde decarboxylase (ACMSD) and increases quinolinic acid in serum.

Hepatic ACMSD [EC4.1.1.45] plays a key role in regulating NAD biosynthesis from tryptophan. We previously reported that ingestion of polyunsaturated fatty acids by rats leads to a decrease in their hepatic ACMSD activity. We purified ACMSD and cloned cDNA encoding rat ACMSD. Therefore, in this study, we examined whether dietary linoleic acid altered ACMSD gene expression and its protein level. Moreover we measured the tryptophan catabolite quinolinic acid level in rats. In the rats fed with linoleic acid, ACMSD mRNA and its protein levels in the liver were strongly suppressed and serum quinolinic acid was significantly increased as compared with the rats fed on a fat-free diet. These results suggest that the transcription level of ACMSD is modulated by linoleic acids or their metabolites and probably there is an inverse relationship between ACMSD activity and the production of quinolinic acid converted from tryptophan.

Modulation of rat liver apolipoprotein gene expression and serum lipid levels by tetradecylthioacetic acid (TTA) via PPARα activation

3-Thia fatty acids are modified fatty acids that promote hepatic peroxisome proliferation and decrease serum triacylglycerol, cholesterol and free fatty acid levels in rats. In vivo administration of tetradecylthioacetic acid (TTA) to rats led to a significant decrease in liver apolipoproteins apoA-I, A-II, A-IV, and C-III mRNA levels, and to an increase of liver acyl-CoA oxidase (ACO), carnitine palmitoyltransferase-II, and 3-hydroxy-3-methylglutaryl coenzyme A synthase (HMG-CoA synthase) mRNA levels and activities. By contrast, no significant changes of lipoprotein lipase (LPL) mRNA levels were detected in rat epididymal adipose tissue. Liver carnitine palmitoyltransferase-I, apoB, apoE, and LDL receptor mRNA levels were not significantly affected. When tested in vitro, TTA increased rat ACO and carnitine palmitoyltransferase-I mRNA levels in primary rat hepatocytes and also LPL mRNA levels in 3T3-L1 preadipocytes. TTA also enhanced the transcriptional activity of chimeras containing the DNA binding domain of the yeast transcription factor Gal4 fused to the ligand binding domain of either human PPARalpha or human PPARgamma. The effect depended on the concentration tested and the cell type. In conclusion, our data suggest that in vitro, TTA activates both PPARalpha and PPARgamma, but the latter with much lower affinity. TTA affects serum lipid levels in vivo in rats by acting mainly on the liver via PPARalpha where it decreases the liver expression of genes involved in vascular lipid transport and increases the expression of genes involved in intracellular fatty acid metabolism. -Raspé, E., L. Madsen, A-M. Lefebvre, I. Leitersdorf, L. Gelman, J. Peinado-Onsurbe, J. Dallongeville, J-C. Fruchart, R. Berge, and B. Staels. Modulation of rat liver apolipoprotein gene expression and serum lipid levels by tetradecylthioacetic acid (TTA) via PPARalpha activation.

The influence of nimodipine, nicardipine and amlodipine on the brain free fatty acid level in rats with penicillin-induced seizures

1. 1. The aim of this study was to investigate the effects of the calcium channel blockers, nimodipine, nicardipine and amlodipine, on the brain free fatty acid (FFA) level in rats with chemically-induced seizures. 2. 2. The study was carried out on Hannover-Wistar rats. Animals were anesthetized and placed in a stereotaxic apparatus. Each of them received an injection of penicillin (5000 IU/5 μl) into the left lateral ventricle (i.c.v.). Various doses (1,3,10 or 30 mg/kg) of niniodipine, nicardipine or amlodipine had been injected i.p, 30 min before the penicillin application. The rats were decapitated 5 min after the occurrence of epileptic seizures. FFAs were quantified by gas chromatography using the internal standard method. 3. 3. The results demonstrate that i.c.v. injection of penicillin was associated with significant increase in the brain FFA concentration. Tested doses of nicardipine and amlodipine did not influence the increase of the brain free palmitic, stearic, oleic and arachidonic acid level while nimodipine prevented the accumulation of free palmitic, oleic and arachidonic acid in rats with penicillin-induced seizures. Statistically insignificant decrease of steric acid was observed in animals pretreated with nimodipine. 4. 4. It maybe assumed that the brain FFA accumulation caused by i.c.v. penicillin administration is not predominantly associated with a disturbance in calcium homeostasis via L-type voltage-sensitive calcium channels, but by some other membrane and/or intracellular mechanisms .

Quick responses of plasma amino acid concentration to changes in dietary amino acid levels in rats.

In order to evaluate the availability of plasma amino acid (AA) concentration as a parameter to determine AA requirements of rats within a short period, the response of the plasma AA concentration to changes in dietary AA levels was studied. In Experiment 1, to clarify whether the plasma AA concentration responded to changes in dietary AA levels and how long the response of the plasma AA concentration was maintained, 10 male rats were fed a casein diet for a 7 day period, and then the food was changed to AA diets. The serine (Ser), glycine (Gly) and threonine (Thr) levels were decreased from 120, 90 to 60% of the casein diet at 7-day intervals successively. Blood samples were taken at 2, 4 and 7 days after changing the diet, from the vena caudalis by vein puncture. In all cases, the plasma AA concentration responded to changes in dietary AA within 2 days, and the response was maintained until the 7th day. Experiment 2 was conducted to confirm that the plasma concentration of each AA responded to the changes in dietary AA levels at the first day and the responses of the plasma concentration of AA were maintained until the 4th day after changing the dietary AA levels, when the combination of metabolically unrelated AA was added. Five rats in A groups were fed diets with added arginine + Thr, histidine + methionine, lysine or isoleucine + Ser, and five rats in B groups were fed diets with added Gly + valine, leucine + phenylalanine or Thr + tyrosine + valine for a 4-day period successively. Blood samples were taken at 1 and 4 days after changing diets. Although the plasma concentrations of histidine, leucine, methionine, phenylalanine and Ser did not respond to changes in dietary AA levels, those of other AA responded at 1 day and the responses were maintained until the 4th day.

Effects of epidermal growth factor on maternal and fetal serum amino acid levels in rats.

Pregnant rats were subcutaneously administered with mouse epidermal growth factor (EGF) at the concentration of 0, 100, or 200 micrograms/kg body weight/day from day 18 to 21 of gestation. The amino acid analysis by high-performance liquid chromatography demonstrated that the umbilical venous/maternal and fetal/maternal ratio of serum proline concentration increased in EGF dose-dependent manner accompanied by the increase in the ratios of total fetal weight and placental weight to maternal body weight gain. These results suggested that EGF regulates fetal growth by, as one of its possible mechanism, promoting placental proline supply from mother to fetus.

( R, S)-3,4-dichlorobenzoylalanine (FCE 28833A) causes a large and persistent increase in brain kynurenic acid levels in rats

Kynurenic acid is an endogenous excitatory amino-acid receptor antagonist with neuroprotective and anticonvulsant properties. We demonstrate here that systemic administration of the new and potent kynurenine 3-hydroxylase inhibitor ( R, S)-3,4-dichlorobenzoylalanine (FCE 28833A) causes a dose-dependent elevation in endogenous kynurenine and kynurenic acid levels in rat brain tissue. In hippocampal microdialysates, peak increases of 10- and 80-fold above basal kynurenic acid concentrations, respectively, were obtained after a single oral or intraperitoneal administration of 400 mg/kg FCE 28833A. After intraperitoneal treatment with FCE 28833A, extracellular brain kynurenic acid levels remained significantly elevated for at least 22 h, rendering this compound a far more effective enhancer of kynurenic acid levels than the previously described kynurenine 3-hydroxylase blocker m-nitrobenzoylalanine. FCE 28833A and similar molecules may have therapeutic value in diseases which are linked to a hyperfunction of excitatory amino-acid receptors.

Chronic administration of the luteinizing hormone-releasing hormone (LHRH) antagonist cetrorelix decreases gonadotrope responsiveness and pituitary LHRH receptor messenger ribonucleic acid levels in rats

Continuous exposure to LHRH or its agonistic analogs results in a reduction of LHRH receptor sites and messenger RNA (mRNA) transcripts as well as in desensitization of the pituitary gonadotropes. To determine, whether LHRH antagonists might be similar in this respect to the agonists, we treated male rats for 4 weeks with daily sc injections of LHRH antagonist [Ac-D-Nal2,Phe(4Cl)2,D-Pal(3)3, D-Cit6,D-Ala10]LHRH (Cetrorelix acetate) or LHRH agonist, [D-Trp6]LHRH, in doses of 100 micrograms/animal-day. Another group of rats received a single im injection of 4.5 mg Cetrorelix pamoate depot, a sustained delivery formulation of the LHRH antagonist. An iv stimulation test with LHRH (200 ng/rat) was performed after 4 weeks of treatment. The rats were killed, and pituitary LHRH receptor characteristics were measured by RRA. To examine the effect of LHRH antagonist treatment on the expression of the pituitary LHRH receptor gene, some of the rats injected with Cetrorelix pamoate depot were killed after 2 weeks, and levels of LHRH receptor mRNA were determined by Northern blot and dot blot hybridization to a 32P-labeled rat complementary DNA probe. Our data show that LHRH-stimulated LH secretion at 30 min was suppressed by approximately 33% (P < 0.01) in rats pretreated with [D-Trp6]LHRH compared to that in animals injected with LHRH alone. Pretreatment of the rats with the LHRH antagonist suppressed the LH response to LHRH more markedly, the LH levels at 30 min were decreased by 89.8% and 96% in groups treated with Cetrorelix acetate and Cetrorelix pamoate depot, respectively. The testosterone response was virtually abolished in groups receiving Cetrorelix. The concentration of pituitary receptors for LHRH fell by 69% in the [D-Trp6]LHRH group, whereas the reductions in the Cetrorelix acetate group and in the group that received Cetrorelix pamoate depot were 77% and 82%, respectively. Treatment with Cetrorelix pamoate depot led to a 75-80% decrease in the levels of 5.0- and 4.5-kilobase forms of LHRH receptor mRNA compared to those in the control group. Dot blot analysis also showed 83% reduction in the mRNA for LHRH receptor. In conclusion, these data demonstrate that prolonged administration of LHRH antagonists such as Cetrorelix causes an impairment of gonadotropin secretion and a marked decrease in the levels of LHRH receptors as well as in the expression of the LHRH receptor gene. Thus, the down-regulation of pituitary LHRH receptors produced by LHRH antagonists appears to be similar to that resulting from the agonists.