Oncological outcomes in rats given nephrocarcinogenic exposure to dietary ochratoxin a, followed by the tumour promoter sodium barbital for life: a pilot study.

The potent experimental renal carcinogenesis of ochratoxin A (OTA) in male rats makes the dietary contaminant a potential factor in human oncology. We explored whether the tumour promoter sodium barbitate could shorten the otherwise long latency between exposure to toxin and tumourigenesis. Young rats, of a hybrid in which mononuclear leukaemia was rare, were given feed contaminated (5 ppm) with OTA for 36 weeks to initiate renal tumourigenesis. Some individuals were thereafter given sodium barbitate (500 ppm in drinking water) for life. Pathological outcomes were studied at or near the end of natural life. Renal tumours in males given barbitate became evident after latency of one year, but only slightly before those without barbitate. In contrast, female mammary tumourigenesis was advanced by at least 6 months synchronously in all rats given the OTA-barbitate regimen compared to tumourigenesis in controls. Diagnosis of malignant mammary angiosarcoma in a female given the OTA-barbitate regimen is a new finding in the rat. The long latency of OTA-induced renal tumourigenesis was not notably susceptible to accelerated promotion by barbitate, contrasting with an apparently marked effect of barbitate on development of mammary tumours.

Promotion by sodium barbital induces early development but does not increase the multiplicity of hereditary renal tumors in Eker rats.

Induced cell proliferation is important in the mode of action of many non-genotoxic renal carcinogens. Since Tsc2 mutant (Eker) rats are genetically predisposed to the development of renal cell tumors, they provide a useful animal model in which to study the action of renal carcinogens. Sodium barbital was used as a model non-genotoxic renal carcinogen to test whether a concentration that increased renal tubular proliferation without severe nephrotoxicity would enhance tumor induction in a hereditary tumor model. First, a subchronic concentration-response study was conducted in wild-type male Long-Evans rats to determine increased cell proliferation without severe nephrotoxicity. Rats were dosed with sodium barbital in the feed at 0, 50, 250, 500, 1000, 2000 or 4000 p.p.m. for 3 or 8 weeks. Cell proliferation within the cortex and nephrotoxicity were quantitated. Enhanced proliferation with minimal nephrotoxicity occurred at 500 p.p.m. A second study was conducted in male Tsc2 mutant rats given sodium barbital in the feed at 0, 100 or 500 p.p.m. from 9 weeks of age to either 6 or 12 months of age. An additional group of rats was treated with sodium barbital for 6 months and then provided control feed until 12 months of age. Rats necropsied at 6 months of age had a concentration-dependent increase in preneoplastic and total renal lesions. Sodium barbital-treated rats necropsied at 12 months of age had numbers of lesions that were not different from controls. Total combined preneoplastic and neoplastic lesions in the 6 month, high dose group was the same as the 12 month control group. These data show that sodium barbital caused progression to the stage of spontaneous renal lesions in Tsc2 mutant rats but did not increase their overall number. These data suggest that enhanced cell proliferation without significant cytotoxicity exerted a promotional influence in this hereditary model.

Mouse popliteal lymph node assay for assessment of allergic and autoimmunity-inducing potentials of low-molecular-weight drugs.

In the present collaborative study, popliteal lymph node (PLN) responses to penicillin G (an allergenic chemical), D-penicillamine (an autoimmunity-inducing chemical), and barbital (a negative reference chemical) were investigated in three different mouse strains by ten pharmaceutical companies. Two inbred mouse strains (BALB/c and A/J) and one outbred strain (ICR) were subcutaneously injected with saline solutions containing penicillin G (1.25, 2.5 and 5 mg/mouse), D-penicillamine (0.5, 1 and 2 mg/mouse), or barbital (2 mg/mouse) into one hind footpad and saline only was injected into the contralateral footpad. PLN cellularity indices were determined on day 7. In the three strains tested, the penicillin G and D-penicillamine injections resulted in approximately dose-dependent responses. In contrast, barbital failed to generate a significant PLN reaction. In the typical data from one of the participating laboratories, the PLN responses of A/J, BALB/c, and ICR to penicillin G were high, intermediate and low, respectively, while their PLN responses to D-penicillamine were all high. Some variation in PLN cellularity indices was observed among the participating laboratories, but reproducibility of the popliteal lymph node assay (PLNA) evaluation was partly confirmed. Although the appropriate selection of mouse strains and drug dosage levels has to be considered, these results suggest that the PLNA may be an appropriate screening system for prediction of the allergic or autoimmunity-inducing potentials of low-molecular-weight drugs.

The popliteal lymph node assay: results of a preliminary interlaboratory validation study.

The popliteal lymph node (PLN) assay was proposed to predict possible autoimmune effects of xenobiotics. A preliminary interlaboratory validation study of the PLN assay was conducted in Wistar rats. Three laboratories tested in blind fashion four compounds, namely chlorpromazine, zimeldine, hydrazine and streptozotocin, which were reported to cause autoimmune-like reactions in humans, and one compound, i.e. barbital, which was not, using strictly the same experimental procedure. All tested substances were injected into the hind footpad of rats on day 1, and PLN weight and cellularity were measured on day 8. Comparison of the controlateral PLN was used to calculate weight and cellularity indices. The results were independently analyzed in a fourth laboratory. All four positive compounds were detected by the three laboratories using both weight and cellularity indices, and the negative compound consistently proved negative. Despite variations in absolute values between laboratories, although not significant, these results provide further evidence of the potential predictive value of the PLN assay.

Renal tubular tumors and atypical hyperplasias in B6C3F1 mice exposed to lead acetate during gestation and lactation occur with minimal chronic nephropathy.

Lead is a high-priority hazardous substance in humans and a renal carcinogen in adult rodents. This study assessed the carcinogenic potential and toxicity of gestational and lactational lead exposure in (C57BL/6NCr x C3H/HeN)F1 (hereafter called B6C3F1) mice. Effects of a renal tumor promoter [barbital sodium (BB)] on lead-initiated lesions were also studied. Pregnant female C57BL/6NCr mice (10-15/group) previously bred with C3H/HeN males were given lead acetate (0, 500, 750 and 1000 ppm lead) ad libitum in their drinking water, starting on gestation day 12 and continuing to 4 weeks postpartum. Offspring were then weaned and divided into same-sex groups of 23-25 and observed for a maximum of 112 weeks. Other groups received lead and then continuous BB (500 ppm) ad libitum in their drinking water from weaning onward. In control male offspring (0 lead/0 BB), renal proliferative lesions [(RPLs); defined as atypical tubular hyperplasia or tumor] occurred rarely (1 lesion-bearing mouse/23 mice examined, 4%) and did not include tumors. RPLs increased in a dose-related fashion with lead exposure (500 lead/0 BB, 4/25, 16%; 750 lead/0 BB, 6/25, 24%; 1000 lead/0 BB, 12/25, 48%) in male offspring and were often multiple. All lead-treated groups had renal tumors, including carcinoma, but these were most common at the highest dose (1000 lead/0 BB, 5/25). Lead-induced renal tumors arose in the absence of the extensive chronic nephropathy and lead inclusion bodies typically seen with lead carcinogenesis in rodents exposed chronically as adults. Postnatal BB exposure had no effect on RPL incidence (e.g., 1000 lead/500 BB, 8/25, 32%). Lead-treated female offspring also developed RPLs, including adenoma and carcinoma, but at a much lower rate than males. Thus, short-term lead exposure during the gestational/lactational period has carcinogenic potential in the mouse kidney.

Transplacental carcinogenesis by cisplatin in F344/NCr rats: promotion of kidney tumors by postnatal administration of sodium barbital.

Transplacental carcinogenic effects of cis-dichlorodiammineplatinum (cis-DDP) in F344 rats were investigated. Pregnant F344 rats were given a single i.p. administration of either cis-DDP (5 mg/kg body wt; group 1) or saline only (group 2) on Day 18 of gestation. Offspring of groups 1 and 2 were randomly and equally divided into two subgroups: 1a, 1b and 2a, 2b, respectively. Beginning at 4 weeks of age, offspring in groups 1b and 2b received 500 ppm of sodium barbital (NaBB) in diet, while those in groups 1a and 2a received normal diet. The experiment was terminated at 79 weeks of age. A low incidence (2/19; 10.5%) of male offspring exposed to transplacental cis-DDP (group 1a) developed renal cell adenomas. Postnatal administration of NaBB significantly enhanced this incidence (10/22; p < 0.01) in cis-DDP-initiated offspring. Also, multiple kidney tumors were more common in group 1b than any other group and three animals in this group developed frank renal cell carcinomas. cis-DDP, administered transplacentally, was a complete carcinogen for rat liver as the incidence of hepatocellular adenomas was significantly higher in offspring exposed transplacentally to cis-DDP than in those exposed to saline (20/82 vs 3/75; p < 0.05). NaBB, a known promoter of hepatocellular carcinogenesis in adult rats initiated with N-nitrosodiethylamine, failed to promote liver carcinogenesis initiated by transplacental cis-DDP. Tumors of the central nervous system (3/82; gliomas) and peripheral nervous system (2/82; schwannomas) were found only in offspring exposed transplacentally to cis-DDP. Thus, cis-DDP, administered transplacentally, was a strong initiator of renal cell tumors and a complete carcinogen for multiple organs in rat offspring.

Induction of CYP2B1 mediated pentoxyresorufin O-dealkylase activity in different species, sex and tissue by prototype 2B1-inducers.

The induction of CYP2B1 mediated pentoxyresorufin O-dealkylase (PROD) activity by various xenobiotics was explored in liver, kidney and lung from a variety of animal species of both sexes, in order to gain insights into the substrate specificity of induced CYPs. Marked species- and sex-related differences in the inducibility of PROD activity by tested chemicals were observed, the mouse being always more responsive when compared to hamster or rat. Induction by sodium phenobarbital (NaPB) led to a conspicuous increase in all situations, up to approximately 38-fold in female rat and mouse liver, with the exception of hamster kidney where PROD activity was only slightly affected. Unexpectedly, both sodium barbital (NaB) and phorone (PHR) moderately induce CYP2B1 isoforms in rat, the extent being highest in female kidney (PHR, 14-fold increase) and male lung (NaB, 4.5-fold). The degree of induction was maximal in the liver with some exceptions occurring in male mice where NaB induced up to 46- and 115-fold increases in lung and kidney and PHR up to 115-fold in kidney. Minimal, although significant induction of PROD activity following treatment with trans-1,2-dichloroethylene (1,2-DCE) occurred in all situations with the exception of hamster kidney and lung. Therefore, caution should be exercised when using PROD activity as specific enzymatic assay to probe CYP2B1-like induction.

Effects of barbital on neuro-oncogenesis in a transplacental carcinogenicity model using F344 rats.

Effects of barbital (BB) on neuro-oncogenesis were examined in a rat transplacental carcinogenesis model. Pregnant F344 rats were divided into 7 groups. Dams in group I received subcutaneous injections of 10 mg/rat 1-butyl-1-nitrosourea (BNU) on the days 15, 18 and 21 of pregnancy and dams in groups II-IV, 1mg/rat BNU on the same time schedule. In addition to the treatment with BNU, dams in group IV were given 0.125% BB solution as their drinking water from the day 12 of pregnancy to parturition. Offspring in groups III and IV received 0.125% BB solution as drinking water from 4 weeks of age until the termination of the study. Animals in groups V and VI were given 0.25% and 0.125% BB solutions, respectively, in the peri- and postnatal period without BNU treatment. Dams in group VII received 250 mg/kg BB subcutaneously on the days 15, 18 and 21 of pregnancy. Offspring in all groups were observed until 105-116 weeks of age. High yields of neurogenic tumors, such as gliomas and neurinomas, were observed in group I. In groups II, III and IV, single cases of a chordoma, a granular cell tumor, and a neurinoma and a malignant reticulosis, which are known to occur spontaneously, were noted, although no gliomas were found. No neurogenic tumors were observed in groups V-VII. With regard to lesions other than those in neurogenic organs, a significant increase in liver tumors was observed in group III compared to group II. In contrast, lung tumors were not found in group III, while they were observed in groups II and IV. These results suggest that BB has no neuro-carcinogenic activity in the rat transplacental carcinogenesis model.

Comparison of popliteal lymph node responses in various strains of rats.

Outbred (namely Wistar and Sprague-Dawley) and inbred (Wistar-Furth, Lewis, Fisher 344 and Brown-Norway) strains of rats were screened for their responses to reference compounds in the popliteal lymph node (PLN) assay. Streptozotocin and diphenylhydantoin gave positive responses as evidenced by increased weight and cellularity indices in all strains used whereas procainamide, isoniazid and barbital consistently gave negative responses. Although these findings overall are in agreement with previous investigations involving these compounds, the lack of marked interstrain differences in PLN responses argues against a strong immunogenetically controlled mechanism as could be assumed in presumably auto-immune reactions. The question is raised whether drug-induced side-effects predicted by the PLN assay are basically non-autoimmune as suggested by clinical and immunological findings in man.

Comparative hyaline droplet nephropathy in male F344/NCr rats induced by sodium barbital and diethylacetylurea, a breakdown product of sodium barbital.

Hyaline droplet nephropathy in male rats due to alpha 2u-globulin accumulation in proximal tubules is caused by chemicals from several chemical classes. We have previously shown that the well-known sedative/hypnotic barbiturate, sodium barbital, and its breakdown product, diethylacetylurea, are renal toxins and renal tumor promoters. To determine comparative induction of hyaline droplets in renal tubules by sodium barbital and diethylacetylurea, male F344/NCr rats, 6 weeks of age, were given diets containing 0, 170, 341, 500, or 1000 ppm of diethylacetylurea or containing 500, 1000, or 4000 ppm of sodium barbital for periods of 2 or 10 weeks. Rats were terminated at 2 or 10 weeks and the histology of the kidney was evaluated using light microscopy with hematoxylin and eosin staining and staining by the Heidenhain method. Quantitative analysis showed dose responses for the degree of droplet accumulation in the P2 and P3 segments of the proximal tubules. Diethylacetylurea was more potent. Immunohistochemistry and ultrastructural evaluation revealed the nature of the droplets. Western blotting confirmed the presence of alpha 2u-globulin. Renal tubular necrosis, regeneration, and increased levels of cell proliferation using proliferating cell nuclear antigen immunohistochemistry were also found. Female rats similarly exposed to each chemical did not show tubule droplet accumulations nor renal lesions. We confirm for the first time that these two chemicals can be added to the enlarging list of nephrotoxic chemicals inducing alpha 2u-globulin nephropathy and possessing tumor promoting and renal carcinogenic properties.

Possible involvement of NMDA receptor-mediated transmission in barbiturate physical dependence.

1. The competitive antagonists at the N-methyl-D-aspartate (NMDA) receptor, CGP39551 and CGP37849, protected against the barbiturate withdrawal syndrome in mice, as measured by ratings of convulsive behaviour on handling. 2. The effective doses of these compounds were lower than those required to prevent seizures due to NMDA in naive animals; these were in turn lower than those needed to prevent the convulsive effects of the alpha-aminobutyric acid (GABA) antagonist, bicuculline. 3. The NMDA-receptor antagonists did not alter the increase in the incidence of convulsions due to the GABAA antagonist, bicuculline, that is seen during barbiturate withdrawal, although the latencies to these convulsions during barbital withdrawal were significantly increased after CGP39551. 4. Barbiturate withdrawal did not affect the convulsive actions of NMDA, whether measured by the incidence of convulsions or by intravenous infusion. 5. The Bmax for [3H]-dizocilpine ([3H]-MK801) binding was significantly increased by chronic barbital treatment in cerebrocortical but not in hippocampal tissues, while the Kd remained unaltered in either case. 6. At 1 h and 24 h after administration of a single dose of barbitone, the Bmax for [3H]-dizocilpine binding was unaltered in cerebrocortical tissue. Acute addition of barbitone in vitro did not alter [3H]-dizocilpine binding or the displacement of binding of thienylcyclohexylpyridine.

Cell proliferation and renal carcinogenesis.

Enhanced cell proliferation occurs at several stages of renal tumorigenesis. Initiation by genotoxic nephrocarcinogens such as dimethylnitrosamine (DMN) is likely a result of DNA damage coupled with an initial burst of DNA synthesis associated with the cytotoxic effects of the compound. The level of initiation by DMN can be further enhanced by unilateral nephrectomy or hydronephrosis, which induces a brief burst of cell proliferation followed by tumorigenesis in the contralateral kidney. The role of sustained cell proliferation in renal tumor development is less well understood. The most compelling evidence comes from studies with nongenotoxic renal carcinogens such as unleaded gasoline and d-limonene, which induce alpha 2u-globulin (alpha G) nephropathy and renal epithelial tumors exclusively in male rats. Sustained increases in cell proliferation in these studies depend on the presence of a chemical-alpha G complex in phagolysosomes of P2 proximal tubule cells, which results in cytotoxicity and compensatory hyperplasia only in male F344 rats, but not female F344 rats or alpha G deficient male NBR rats. Furthermore, initiation-promotion experiments demonstrated a strong correlation between the dose-response of cell proliferation and the incidence of preneoplastic and neoplastic lesions. Clearly, similar correlative studies with a number of other renal carcinogens and non-carcinogens are warranted before general conclusions can be made. Cell proliferation is excessively elevated in tubules affected by chronic progressive nephropathy, but the significance of the lesion to renal carcinogenesis is unclear. Elucidating mechanisms of renal cell proliferation are necessary for our understanding of cause and effect relationships. An exciting recent finding is altered expression of transforming growth factor-alpha in hereditary rat renal cell carcinoma.(ABSTRACT TRUNCATED AT 250 WORDS)

Pathology of preneoplastic and neoplastic renal tubular lesions induced in F-344 rats by sodium barbital, a nongenotoxic renal carcinogen and nephrotoxin.

Sodium barbital (NaBB), a long-duration sedative/hypnotic barbiturate, is a nongenotoxic nephrotoxin and induces chronic persistent increases in rates of cell proliferation in renal cortical tubules of male F-344/NCr rats. In 5 of our 2-stage carcinogenesis experiments with NaBB at doses of 500, 1,000, or 4,000 ppm for periods of up to 106 wk of age, renal tubular cell tumors were found in incidences of up to 25% in rats receiving only NaBB while fewer than 1% of controls had renal epithelial tumors. We reviewed renal tubular proliferative lesions found in these studies and classified the lesions based on morphology, histogenesis, and immunohistochemical findings. Renal dysplastic tubules (DTs; atypical hyperplasia), putative preneoplastic lesions rarely seen in controls, were found in the renal cortex of more than 50% of the NaBB-exposed rats. DTs were classified into grades 1-3, based on lesion size and growth patterns. All renal adenomas were usually of the basophilic phenotype, and 70% of basophilic adenomas displayed solid patterns, while tumors with papillary, cystic, or tubular patterns were seen less commonly. By serial or step sectioning of the DTs and tumors, evidence was found indicating that the high grades (grade 2 or 3) of DTs, some of which arose in the P1 or P2 segment of the proximal tubules, were sometimes connected to the adenomas. Vimentin expression was demonstrated immunohistochemically in NaBB-induced renal tubular adenomas but not in normal tubules. Tumors were usually not immunoreactive for glutathione S-transferase, placental form, but heterogeneous immunoreactivity was also seen in some tumors. Lysozyme was absent in preneoplastic and neoplastic lesions induced by NaBB, while some intact normal proximal convoluted tubules were immunoreactive. The common tumor phenotype induced by NaBB, the basophilic solid adenoma, was similar to the most common type of spontaneous renal tumor found in untreated aging F-344 rats. NaBB may promote naturally occurring renal preneoplastic or neoplastic tubular lesions of this unique phenotype, but it is also possible that it may induce these lesions de novo.

Nerve growth factor (NGF) in rat brain following long-term barbital treatment: relation to convulsions and cognitive function.

Nerve growth factor (NGF) protein has been implicated in alterations of cognitive function either following brain damage, selective lesions or aging. Groups of rats were given long-term (48 weeks) oral barbital treatment or tap water and following an extended period of abstinence (14 weeks) were tested for spatial learning ability in the Morris swim maze. Following the maze test, they were sacrificed and the NGF content of hippocampal and cortical brain regions were analyzed. Barbital treated rats were divided into convulsing and non-convulsing groups. It was found that there was a slight, significant increase (12%) in NGF content of the hippocampus in convulsing rats. Correlations between maze learning performance, brain weight and NGF in the cortex indicated a significant negative relationship between (a) performance and brain weight on day 1 of testing and (b) NGF content and performance on day 2. These data indicate some involvement of NGF in functions derived from a considerably different animal model to those applied previously.

Lack of renal tumour-initiating activity of a single dose of potassium bromate, a genotoxic renal carcinogen in male F344/NCr rats.

The renal tumour-initiating activity of potassium bromate (KBrO3), a known genotoxic rat renal carcinogen, was investigated in male F344/NCr rats. 6-wk-old rats were given KBrO3 intragastrically as a single dose of 300 mg/kg body weight, which was confirmed by our preliminary toxicity study as a maximum tolerated single dose for this strain of rat. Starting 2 wk after KBrO3 treatment, groups of 39 rats received either a basal diet or a diet containing 4000 ppm barbital sodium (BBNa) as a promoting regimen and were killed at 30, 52, or 104 wk. Control rats received either dietary BBNa (4000 ppm) or the basal diet alone from wk 2 to 52 or 104 wk. Nephropathy was observed in all rats treated with KBrO3 followed by BBNa at 30 wk and in rats receiving BBNa alone, but not in rats exposed to KBrO3 alone. Dysplastic renal tubular cell foci (DTF), putative preneoplastic renal tubular cell lesions were found associated with nephropathy in rats exposed to KBrO3 followed by BBNa from 47 wk. The incidences and multiplicities of DTF and renal tubular cell tumours observed from 31 to 104 wk revealed no initiating effect of KBrO3 treatment. These results indicate that the KBrO3 dose of 300 mg/kg did not initiate renal carcinogenesis.

Susceptibility to, tolerance to, and physical dependence on ethanol and barbital in two inbred strains of rats.

1. Ethanol-induced sleep time was significantly longer in F344 than LEW rats. However, there is no difference in barbital-induced sleep time between F344 and LEW. 2. Development of tolerance to ethanol-induced motor impairment was slightly faster in F344 than in LEW rats. While, LEW rats more easily developed tolerance to the impairment by barbital in comparison with F344 rats. 3. F344 and LEW rats were chronically treated with liquid diet containing ethanol or with barbital-admixed food. After the termination of ethanol and barbital treatments, various withdrawal signs occurred in F344 rats, including tremor and convulsions, whereas LEW rats showed no convulsions. Withdrawal scores of ethanol and barbital were significantly higher in F344 than in LEW rats. 4. These results suggest that strain differences in physical dependence on ethanol and barbital may be mainly influenced by the susceptibility to ethanol and the development of tolerance to barbital, respectively.

Learning deficits in aged rats pretreated chronically with barbital and tested late in abstinence: alleviation by tetrahydroaminoacridine.

Physostigmine and tetrahydroaminoacridine (THA) have been reported to improve cognitive function in patients with Alzheimer's disease. Two experiments were conducted to examine the effects of these anticholinesterase agents on learning in aged rats pretreated chronically with barbital. In the first experiment animals received barbital in their drinking water for 46 weeks. Controls were given only water. On days 100-104 of abstinence, when the animals were 20 months old, acquisition of the Morris maze task was initiated after treatment with physostigmine. It was found that physostigmine improved learning of the maze task in control but not barbital treated rats. In the second experiment animals received barbital solution or water as in experiment one. On days 100-103 of abstinence they were injected with THA before being tested in the Morris water maze. It was found that THA improved learning in both barbital treated and control rats. These results corroborate clinical findings of improved cognitive function following treatment with THA, and suggest that the therapeutic effects of THA may be mediated by mechanisms distinct from cholinesterase inhibition. Furthermore chronic barbital treatment could be used as a model to study cognitive disturbances in experimental animals.

Influence of liver tumor promoters and structurally related chemicals on survival of normal adult rat hepatocytes in primary culture.

The influence of four liver tumor promoters and two structurally related chemicals (non-promoters) on survival of normal adult rat hepatocytes was examined in primary culture. Of the four promoters, butylated hydroxytoluene (BHT), dichlorodiphenyltrichloroethane (DDT), barbital sodium and phenobarbital sodium, only phenobarbital efficiently prolonged the hepatocyte survival and maintained the morphological features of the cells. Both BHT and DDT were toxic to hepatocytes within the dose ranges tested. Barbital was also ineffective for maintenance of primary cultured hepatocytes but not toxic to the cells. Of the two non-promoters, barbituric acid and amobarbital, barbituric acid also showed no maintenance effect on hepatocytes. However, amobarbital resembled phenobarbital in its effect on the maintenance of hepatocytes in primary culture. DNA synthesis of primary cultured hepatocytes was severely suppressed by phenobarbital in a dose-dependent manner. The results clearly show that the ability to support long-term survival of primary cultured hepatocytes is a common property of some barbiturates but not of liver tumor promoters, and that the maintenance of hepatocytes by phenobarbital is not due to a counterbalance of stimulated proliferation and death of the cells.

Effect of GABAergic drugs on motor impairment from ethanol, barbital and lorazepam in rat lines selected for differential sensitivity to ethanol.

The effect of GABAergic drugs on the motor-impairing effects of ethanol, barbital, and lorazepam were studied in the ethanol-sensitive ANT (Alcohol Nontolerant) and ethanol-insensitive AT (Alcohol Tolerant) rat lines, selected for differential ethanol-induced motor impairment on the tilting plane. The basic population from which these rat lines were derived, the mixed (M) line, was also included in the study. The ANT rats were more sensitive to the intoxicating effects of ethanol, barbital, and lorazepam than the AT and M rats at the dose ranges tested. Picrotoxin antagonized motor impairment from all three drugs. Flumazenil (Ro 15-1788) antagonized only the effects of lorazepam, and isoniazid did not modify motor impairment induced by any of the three drugs. These results confirm that the selection of AT and ANT lines has not been specific to ethanol, and that it has increased sensitivity to ethanol, barbital, and lorazepam in the ANT rats rather than decreasing it in the AT rats relative to the M rats. The finding that picrotoxin counteracted motor impairment from ethanol, barbital, and lorazepam support the view that the GABAA receptor complex is important in mediating the intoxicating effects of these drugs. These results also suggest that the genetically-determined difference in sensitivity to ethanol between the rat lines involves GABAergic mechanisms, but it remains to be determined whether any part of the GABAA receptor itself has been affected by the selection program.

Barbital sodium, a tumor promoter for kidney tubules, urinary bladder, and liver of the F344 rat, induces persistent increases in levels of DNA synthesis in renal tubules but not in urinary bladder epithelium or hepatocytes.

The nephrotoxicity of barbital sodium (NaBB), a renal and urinary bladder tumor promoter, was investigated in male F344/NCr rats. In an 8-week toxicity study, NaBB was administered to 6-week-old male rats for 8 weeks at dietary levels of 16,000, 8000, 4000, 2000, 1000, or 0 ppm. Rats tolerated NaBB at levels of 4000 ppm and below with no weight depression or mortality. Liver-to-body weight ratios, however, were significantly increased at 4000 ppm, and toxic renal lesions were observed histologically. Light microscopic studies of male rats fed 1000 ppm NaBB for 2-52 weeks or 4000 or 8000 ppm for 8 weeks revealed elevated levels of DNA synthesis in renal tubular cells as measured with tritiated thymidine autoradiography or 5-bromo-2'-deoxyuridine immunohistochemistry that were associated with degenerative and regenerative nephrotoxic lesions. Increased labeling indices of urothelium and hepatocytes were not seen in rats exposed to 1000 ppm NaBB which is effective as a bladder and liver tumor promoter at these doses. These studies provide evidence for the chronic nephrotoxicity and renal tubular hyperplasia induced by NaBB in F344 rats, which are associated with the tumor-promoting activity of NaBB at the doses studied. Hyperplasia in the urinary bladder or liver was not found, however, for this bladder and liver tumor promoter. The conflicting findings in liver, bladder, and kidney are discussed.