Quantitation of glandular gastric changes in rats given a proton pump inhibitor for 3 months with emphasis on sampling scheme selection.

Proton pump inhibitors and H2-receptor antagonists suppress gastric acid secretion and secondarily induce hypergastrinemia. Sustained hypergastrinemia has a trophic effect on stomach fundic mucosa, including enterochromaffin-like (ECL) cell hypertrophy and hyperplasia. Histomorphometric quantitation of the pharmacologic gastric effects was conducted on 10 male and 10 female rats treated orally with LY307640 sodium, a proton pump inhibitor, at daily doses of 25, 60, 130, or 300 mg/kg for 3 mo. Histologic sections of glandular stomach, stained for chromogranin A, were evaluated by image analysis to determine stomach mucosal thickness, mucosal and nonmucosal (submucosa and muscularis) area, gastric glandular area, ECL cell number/area and cross-sectional area. Total mucosal and nonmucosal tissue volumes per animal were derived from glandular stomach volumetric and area data. Daily oral doses of compound LY307640 sodium caused slight to moderate dose-related mucosal hypertrophy and ECL cell hypertrophy and hyperplasia in all treatment groups as compared with controls. All observed effects were prominent in both sexes but were generally greater in females. The morphometric sampling schemes were explored to optimize the data collection efficiency for future studies. A comparison between the sampling schemes used in this study and alternative schemes was conducted by estimating the probability of detecting a specific percentage of change between the male control and high-dose groups based on Tukey's trend test. The sampling scheme analysis indicated that mucosal thickness and mass had been oversampled. ECL cell density quantitation efficiency would have been increased by sampling the basal mucosa only for short-term studies. The ECL cell size sampling scheme was deemed appropriate for this type of study.

Chronic toxicity of zatosetron, a 5-HT3 receptor antagonist, in rhesus monkeys.

A 1-year chronic toxicity study was conducted in which rhesus monkeys (4/sex/dose) were given daily doses of 0, 3, 10, or 25 mg zatosetron/kg by nasogastric intubation. Clinical signs of toxicity characterized by salivation, diarrhea or soft stools, and/or emesis occurred in animals that received 10 or 25 mg/kg of zatosetron. One monkey in the high-dose group and one in the middle-dose group died as a result of intratracheal administration of the compound. The death of another monkey in the high-dose group was associated with an unexpectedly high (3-fold the mean plasma Cmax value in surviving females in this group) plasma level of zatosetron as indicated by postmortem analysis of heart blood. Animals of both sexes in all treatment groups gained weight at a slightly reduced rate when compared to control monkeys. Depressed appetite occurred in some monkeys in all treatment groups but was most evident in those receiving 25 mg/kg. Evaluation of ECG's indicated that treatment with zatosetron did not produce any rhythm or conduction disturbances. However, there was a mild increase in the Q-Tc interval throughout the treatment period at 4 hours postdosing in monkeys in the middle- and high-dose groups and a slight increase prior to dosing in animals in the high-dose group. Mean plasma Cmax and AUC(0-24 hr) values on Day 360 were dose proportional for zatosetron and for the N-demethylated metabolite in both sexes over the dose range tested. The mean t1/2 (elimination phase) for the plasma disappearance of zatosetron ranged from 3.4 to 7.2 hr in males and from 2.3 to 6.8 hr in females. Hematology, urinalysis, and clinical chemistry parameters were unaffected by treatment. There were no treatment-related gross or microscopic alterations or changes in organ weights. With the exception of mild effects on body weight gain, there was no evidence of chronic toxicity in monkeys given 3 mg/kg zatosetron daily for 1 year.

Chronic toxicity, metabolism, and pharmacokinetics of the 5-HT3 receptor antagonist zatosetron (LY277359) in Fischer 344 rats.

Studies were done to characterize the chronic toxicity, metabolism, and pharmacokinetics of a 5-HT3 receptor antagonist in Fischer 344 rats. Animal were given daily gavage doses of 10, 30, or 90 (females only were increased from 90 to 120 mg/kg for months 7-12) mg/kg of zatosetron for 1 year. Treatment-related histologic changes occurred primarily in the liver and kidney of rats given 30 or 90/120 mg/kg and consisted of hepatocellular fatty change (males only), hepatic granuloma formation, and histiocytosis (females only), and renal pigment deposition (both sexes), lesions not previously described in animals treated with 5-HT3 receptor antagonists. Decreased erythrocyte parameters, increased total leukocyte, lymphocyte, and neutrophil counts, and increased serum alkaline phosphatase, gamma glutamyltransferase, alanine transaminase, and liver weights in females were most likely related to the chronic inflammatory process in the liver. Increased alanine transaminase and transiently increased alkaline phosphatase with increased liver weights in males were likely related to the hepatocellular fatty change. Increased renal tubular epithelial pigment deposition (lipofuscin and hemosiderin) was observed in males and females in the high-dose group and in females in the middle-dose group. Both had increased kidney weights and increased serum inorganic phosphorus. Females in the high-dose group had increased urine volume, decreased pH, and increased total excretion of sodium, potassium, chloride, and creatinine. These changes may have been a reflection of tubular dysfunction associated with excessive pigment deposition. No toxicologically significant effects occurred in rats treated with 10 mg/kg/day for 1 year. Plasma concentrations of zatosetron and its 3-hydroxy metabolite increased with increasing dose and duration of dosing in both males and females during the first 6 months of dosing. Subsequent values measured at 12 months showed no substantive increases except in males given the highest dose. At comparable doses, consistent sex differences (F > M) in mean 1-hr plasma content of parent compound were evident across dose and time. Zatosetron-induced hepato- and nephrotoxicity seems to be peculiar to the rat and is observed only at very high doses relative to the proposed human clinical dose.

A review of the toxicology of the antibiotic MICOTIL 300.

MICOTIL 300 is a new macrolide antibiotic for the treatment of Bovine Respiratory Disease complex. As with other macrolides used in human and veterinary medicine, overdoses of MICOTIL do not produce pathognomonic lesions. The toxicity dose response varies among laboratory animal and domestic livestock species. However, clinical evidence of MICOTIL toxicity due to large doses is generally a manifestation of the positive chronotropic and negative inotropic cardiovascular effects. No adverse environmental effects are expected from the use of MICOTIL in cattle.

Comparative toxicity of oral cephalosporin antibiotics in the rabbit and in a rabbit kidney cell line (LLC-RK(1)).

The nephrotoxic potential of four oral cephalosporin antibiotics, cephalexin, cefaclor, LY195885 and LY171217, was determined in rabbits given single oral doses of 250-500 mg/kg body weight. Histopathological changes, blood chemistry, and ex vivo renal slice function were evaluated 48 hr after dosing. Additionally, the viability of rabbit renal cells in culture (LLC-RK(1)) was evaluated by nigrosin dye exclusion after 48 hr exposure to each antibiotic at concentrations of 0.5-2.0 mg/ml. Only LY171217 was significantly nephrotoxic in vivo. Prominent lesions were observed at 500 mg/kg body weight and were accompanied by marked increases in blood urea nitrogen and serum creatinine, and decreases in ex vivo renal slice gluconeogenesis and p-aminohippurate and tetraethylammonium uptake. In vitro toxicity to renal cells correlated well with the in vivo results yielding TC(50) values (TC(50) = concentration producing 50% lethality) > 1.0 mg/ml for cephalexin, LY195885 and cefaclor. LY171217, on the other hand, was significantly toxic in vitro (TC(50) = < 0.5). These results suggest that renal cells in culture may provide a useful method for examining the nephrotoxic potential of oral cephalosporins before in vivo studies.

A species comparison of the toxicity of nabilone, a new synthetic cannabinoid.

Acute, subchronic, and chronic studies were conducted in various species to evaluate and compare the toxicity of nabilone, a new synthetic 9-ketocannabinoid that is orally effective for the treatment of nausea and vomiting induced by cancer chemotherapy agents. The oral LD50 in mice and rats for nabilone formulated as a polyvinylpyrrolidone (PVP) codispersion was in excess of 1000 mg/kg. Among nonrodents, rhesus monkeys had a higher tolerance to the CNS depression induced by single oral doses of nabilone-PVP than did dogs. Rats fed dietary mixtures of nabilone-PVP which provided approximate daily nabilone doses of 1 to 93 mg/kg tolerated treatment for 3 months with no deaths. Treatment-related changes (at doses greater than or equal to 5 mg/kg) were limited to reduced body temperature, slight-to-moderate decreases in weight gain, and behavioral changes (e.g., hyperactivity, hyperirritability to touch, and hypoactivity). All dogs treated for 3 months with daily oral doses of up to 1.0 mg/kg survived; treatment-related effects were limited to transient episodes of ataxia and anorexia. Nabilone treatment of rats and dogs for 3 months produced no evidence of systemic toxicity in the clinical chemistry, hematology, or pathology parameters examined. Chronic treatment of dogs with daily oral doses of nabilone-PVP equal to 0.5, 1.0, or 2.0 mg of nabilone/kg produced cumulative toxicity; by the end of 7 months, 2, 6, and 7 dogs in the respective dose groups had died. In a number of instances, death was preceded by one or more convulsive episodes. In contrast to the dog, the toxic potential of nabilone was minimal in rhesus monkeys treated with nabilone-PVP for 1 year at daily oral nabilone doses of up to 2.0 mg/kg. The enzymatic reduction of the 9-keto group of nabilone to form carbinol metabolites was a major metabolic pathway for nabilone in dogs but not in rhesus monkeys. The carbinols were long-lived metabolites in the plasma of dogs and accumulated in the plasma compartment with time. Furthermore, the carbinol metabolites were found to concentrate in the brain tissues of treated dogs. Although the precise mechanism for this marked species difference in chronic toxicity is not known, the metabolic differences responsible for the presence of the carbinol metabolites at high concentrations in the plasma and brain over time may play a role in the toxicity observed in the dog.

Species specificity in the metabolism of nabilone. Relationship between toxicity and metabolic routes.

The disposition of 14C-nabilone has been examined in dogs and monkeys; it is rapidly absorbed and extensively metabolized in both species. A metabolic pathway initiated by the stereoselective enzymic reduction of the 9-keto moiety of nabilone was of major importance in the biotransformation of nabilone in the dog but was only a minor pathway in the monkey. The resulting long half-lived carbinol metabolites accounted for 77% of the circulating 14C in dog but for only 19% in monkey, 48 h after drug administration. Accumulation of carbinol metabolites in brain tissue was observed in dogs, with 24 h brain concentrations being five to six times higher than the plasma concentrations. No accumulation of carbinol metabolites, nabilone or of 14C occurred in the brain of monkeys. Accumulation of the carbinol metabolites in brain tissue has been implicated as the causative factor for the CNS toxicity observed in dogs treated chronically with nabilone. Comparison of nabilone pharmacokinetics in dog and humans showed the dog to be unique in producing high levels of carbinol metabolites, so that monkey appeared to be a more appropriate model than dog for pre-clinical toxicological and safety studies. Studies with liver 15,000 g supernatants indicated that the enzymic reduction of nabilone to its carbinol metabolite was a viable metabolic pathway in rat, dog and monkey, so that a distinct species difference exists between dog and monkey in the subsequent metabolism and/or elimination of these carbinol metabolites.

An evaluation of the toxicity of cefaclor in laboratory animals.

The toxicity of cefaclor, a new orally-administered cephalosporin, was evaluated in laboratory animals given single or multiple doses of the antibiotic. The acute toxicity data for cefaclor in mice, rats, dogs and monkeys were comparable to that previously reported for cephalexin. Rats were maintained on dietary mixtures of cefaclor which provided average daily doses of approximately 230 to 950 mg/kg for 28 days in subacute toxicity tests, and 160 to 675 mg/kg for 1 year in chronic toxicity tests. Treatment-related effects in the above studies were limited to soft stool excretion and caecal dilatation in the subacute test. Effects in dogs given daily oral doses of 50 to 200 mg/kg for 30 days were limited to a transient moderate fall in haemoglobin concentration in the two males at the highest dose. Soft stool excretion and occasional episodes of emesis were observed in dogs given cefaclor for 1 year at oral doses of 100 to 400 mg/kg/day. A reversible thrombocytopenia occurred in one animal at the highest dose. Analysis of various tissue fluids taken 2 hours after the last dose revealed that the concentration of cefaclor in the synovial fluid was approximatley one-half of that in serum. The results of these studies indicate that cefaclor has a low toxic potential in the species tested.

The influence of acute diazepam pretreatment on the action and disposition of [14C]pentobarbital in rats.

Diazepam (DZP) pretreatment (100mg/kg, ip) of rats 6 h before pentobarbital administration (45 mg/kg, ip) prolonged the barbiturate-induced narcosis. The concentrations of [14C]pentobarbital and total pentobarbital derivatives in blood or brain showed no differences between control and DZP-pretreated animals. The brain and blood concentrations of pentobarbital, when measured at a time corresponding to the respective arousal times from pentobarbital narcosis, were lower in the DZP-pretreated group. These results indicate that acute DZP pretreatment increases the sensitivity of the rat brain to pentobarbital rather than inducing changes in the disposition of the barbiturate.

The effects of oral diazepam pretreatment on the biliary excretion of sulfobromophthalein in rats.

Studies were performed with rats to examine the effects of single, as well as repetitive oral diazepam (DZP) pretreatment on biliary sulfobromophthalein (BSP) excretion rates and on bile flow parameters. One-hour pretreatment of male rats with 150 mg/kg of DZP resulted in about a one-third reduction in the peak biliary excretion rate of BSP (60 mg/kg, iv) and this was associated with a decrease in relative proportions of conjugated BSP in bile. The biliary excretion of preconjugated BSP was unaffected. BSP hepatic uptake and storage were apparently unaffected. In vitro DZP markedly inhibited BSP conjugating activity. In contrast to the above results, when BSP excretion was examined 1 h after the last of five daily oral doses of DZP (150 mg kg-1 day-1), no change in the peak elimination rate of this dye was evident. However, bile flow rates were higher in DZP-treated rats than in controls. When rats were examined 24 h after the last of five daily oral doses of DZP (150 mg/kg), the choleretic response persisted. Further studies showed that the repetitive DZP pretreatment enhanced the bile salt-independent mechanisms of bile formation.

Potentiation of the hepatotoxic responses to chemicals in alloxan-diabetic rats.

Alloxan diabetes enhances the hepatotoxic response of male rats to chloroform and 1, 1, 2-trichloroethane, but not to trichloroethylene nor 1, 1, 1-trichloroethane. Insulin treatment partially protects the animals against the alloxan-induced enhancement of chloroform hepatotoxicity. Alloxan diabetes also enhances the hepatotoxic response to galactosamine but not to beryllium nor alpha-naphthylisothiocyanate.

Potentiation of carbon tetrachloride-induced hepatotoxicity in alloxan- or strepto- zotocin-diabetic rats.

Studies were performed to examine the effects of alloxan- or streptozotocin-induced diabetes on carbon tetrachloride (CCl4) liver injury. Male rats were pretreated with single i.v. injections of alloxan monohydrate (40 or 80 mg/kg) or streptozotocin (65 mg/kg). A challenging dose of CCl4 (0.1 ml/kg i.p.) was given to rats 4 days after alloxan pretreatment or 5 days after streptozotocin pretreatment, and the animals were sacrificed 24 hours later. Biochemical and morphologic evidence was obtained to show that pretreatment with the diabetogenic agents markedly enhanced CCl4-induced hepatotoxity. The challenging dose of CCl4 had no effect on the serum glutamic pyruvic transaminase (SGPT) activity in control rats. However, the administration of this dose of CCl4 to rats pretreated with 40 and 80 mg/kg of alloxan as well as to rats pretreated with streptozotocin resulted in 11-, 68-, and 32-fold increases, respectively, in SGPT activity. Hepatic triglyceride concentrations in the diabetic rats were also markedly elevated above control values after CCl4 challenge. Alloxan- or streptozotocin-pretreatment alone did not enhance these biochemical parameters of liver injury. Hepatic glucose-6-phosphatase activity, which increased in the rats given a diabetogenic agent, was lowered as a result of CCl4 injection. Insulin treatment of rats given alloxan (80 mg/kg) markedly protected against CCl4-induced hepatotoxicity. The severity of the morphologic changes in diabetic rats given CCl4 correlated with the biochemical findings.