methylazoxymethanol and Body-Weight

Several findings have indicated that schizophrenia may be connected with the impaired epigenetic regulation of gene transcription. The present study investigated the epigenetic modifications connected with histone H3 methylation at lysine (K)4 and K9 in the medial prefrontal cortex (mPFC) in a neurodevelopmental model of schizophrenia based on prenatal administration of methylazoxymethanol (MAM) at embryonic day 17, which impairs the sensorimotor gating process in adult but not adolescent animals. The effect of MAM was determined at different postnatal ages, pre-puberty (P15, P30 and P45) and post-puberty (P60 and P70), using western blot analyses. MAM treatment altered the levels of H3K9me2 before puberty. H3K9me2 was decreased at P15 and P45 but was increased at P30. In contrast, H3K4me3 was noticeably decreased in adult rats. Immunofluorescence experiments revealed that H3K9me2 protein levels were increased in neuronal cells at P30 and that H3K4me3 levels were decreased in astrocytes at P60 after MAM administration. Decreases in the methyltransferase ASH2L protein levels at P45, P60 and P70 were also observed, while the protein levels of the methyltransferase G9a did not change. In addition, levels of the demethylases LSD1 and JARID1c were analysed after MAM administration. LSD1 protein levels were increased at P15 but decreased at P30. JARID1c protein levels were increased in the MAM-treated animals at P60. Decreased Gad1 mRNA levels were found in adult MAM-treated animals, similar to alternation observed in schizophrenia. The present study indicates that prenatal MAM administration evokes changes in the methylation patterns of histone H3 during postnatal life.

Topics: Animals; Astrocytes; Body Weight; Brain; Enzyme Inhibitors; Glutamate Decarboxylase; Histone Demethylases; Histone-Lysine N-Methyltransferase; Histones; Male; Methylation; Methylazoxymethanol Acetate; Methyltransferases; Neurons; Oxidoreductases, N-Demethylating; Prefrontal Cortex; Rats; Rats, Wistar; RNA, Messenger; Sensory Gating; Teratogens; Valproic Acid

Rats exposed during prenatal life to methylazoxymethanol (MAM) display in postnatal age structural and behavioral deficits resembling those observed in schizophrenic patients. These deficits are associated with significant changes in brain nerve growth factor (NGF) and brain derived neurotrophic factor (BDNF), particularly in the hippocampus and entorhinal cortex. In the present study, we used the MAM model to investigate in young rats the effect of antipsychotics, Clozapine and Haloperidol, on brain and blood NGF and BDNF presence. Young animals were used because administration of antipsychotics during adolescence is a common feature of intervention. The results showed that administration of Clozapine and Haloperidol causes significant changes in the concentration of NGF and BDNF in the brain and bloodstream of MAM-treated rats. These findings indicate that these drugs may affect the synthesis and release of neurotrophins in the central nervous system and in the blood circulation. In addition, the MAM model can be a useful tool to investigate the biochemical and molecular mechanisms regarding the effects of antipsychotics.

Topics: Alkylating Agents; Animals; Antipsychotic Agents; Body Weight; Brain Chemistry; Clozapine; Entorhinal Cortex; Female; Granulocytes; Haloperidol; Hippocampus; Methylazoxymethanol Acetate; Nerve Growth Factors; Pregnancy; Prenatal Exposure Delayed Effects; Rats; Rats, Sprague-Dawley

Recent clinical and laboratory data suggest that there is a link between neuronal migration disorders (NMD) and increased seizure threshold. To characterize an animal model with features similar to human NMD and to assess seizure susceptibility, NMD were induced in the rat at the time of neuroblastic division (PG15) and three other gestational ages (PG 13, PG14, PG16) by transplacental exposure to methylaxozymethanol (MAM, 25 mg/kg). Offspring pups were monitored for spontaneous and electrographic seizures. At postnatal day 14, randomly selected rat pups were sacrificed for histological examination. In other MAM-exposed pups and controls, status epilepticus was induced by intraperitoneal administration of kainic acid. On histology, NMD were found in all PG 15 MAM-exposed rats, in comparison to 63% of PG 13, 70% of PG 14, 80% of PG16. Histological features included cortical laminar disorganization, ectopic neurons in the subcortical white matter and in cortical layer I, persistent granular layer, marginal glioneuronal heterotopia, and discrete areas of neuronal ectopia in the CA1 subfield of the hippocampus. Based on the severity of the neuronal migration abnormalities, rats were divided into three categories: severe, moderate, and mild. Severe and moderate NMD were only found in the PG 15 MAM-exposed rats. EEG recording in rats with NMD did not disclose spontaneous seizures; however, rats with severe NMD had higher slow wave activity compared to controls (P < .05). MAM-exposed rats with severe NMD were more susceptible to kainic-induced seizures compared to controls (P < .05). In rats with severe NMD, kainic acid-induced status epilepticus produced hippocampal damage in the CA3/4 region. These results demonstrate that MAM-induced NMD have histological and electrographic characteristics similar to human NMD. The severity of neuronal abnormality depends on the time of transplacental exposure as the most severe NMD were found after exposure to MAM at the time of neuroblastic division. The degree of NMD positively correlates with seizure susceptibility, since only rats with severe NMD have decreased seizure threshold. The occurrence of status epilepticus-induced hippocampal damage in pups with severe NMD suggests that the severely compromised hippocampus is less resistant to seizure-induced injury than the normal developing brain.

Topics: Animals; Behavior, Animal; Body Weight; Brain; Brain Mapping; Cell Movement; Disease Models, Animal; Electroencephalography; Epilepsy; Female; Gestational Age; Hippocampus; Litter Size; Maternal-Fetal Exchange; Methylazoxymethanol Acetate; Neurons; Organ Size; Pregnancy; Rats; Rats, Sprague-Dawley; Seizures

A single injection of 20 mg/kg methylazoxymethanol acetate (MAM) on gestational day 14 in Brown Norway rats produced micrencephalic offspring (whole brain approximately equal to 65% of control). Despite the micrencephaly, MAM-induced alterations in behavior assessed here were relatively mild. The MAM-treated rats exhibited increased activity under darkened conditions in a complex maze and marginally increased activity after a challenge of methamphetamine. Open field activity, running wheel activity, and emergence behavior using a light/dark apparatus were not significantly affected. Compared with a similar study of Sprague-Dawley micrencephalics [Ferguson S.A., Racey F.D., Paule M.G. and Holson R.R. (1993) Behavioral effects of methyloxymethanol-induced microencephaly. Behav. Neurosci. 107, 1-101], frontal cortex and striatum weights were more reduced in Brown Norway micrencephalics. The MAM-induced behavioral alterations in the Brown Norway strain may have appeared attenuated compared to alterations shown by MAM-treated Sprague-Dawley rats due to differences in baseline between these two strains. Compared to control Sprague-Dawley rats in the previous study, control Brown Norway rats were more active in the open field and running wheels, but less active in the complex maze, exhibiting little to no learning. Emergence tests indicated increased dark preference in Brown Norway rats. Baseline behavior (increased activity and light shyness) of control Brown Norway rats was similar to that of MAM-treated Sprague-Dawley rats; a potential confound in the detection of behavioral effects of a compound. These findings emphasize the effects that strain selection may have on the outcome and interpretation of toxicological/teratological studies.

Topics: Animals; Behavior, Animal; Body Weight; Brain; Female; Male; Maze Learning; Methamphetamine; Methylazoxymethanol Acetate; Microcephaly; Organ Size; Rats; Rats, Inbred BN; Rats, Sprague-Dawley; Species Specificity; Teratogens

It has long been recognized that ethanol (EtOH) interferes with the hypothalamo-pituitary-gonad axis in adults of many species, and recent studies have provided evidence for similar effects after prenatal EtOH exposure. Since EtOH is capable of injuring dividing cells, we investigated the possibility that a single acute in utero EtOH exposure during the period of LHRH neuron formation might change the number of immunoreactive LHRH cells in the hypothalamus. Final LHRH cell division in Long-Evans rats was determined by [3H]thymidine autoradiography to take place over a short period between gestation days 12 and 13. Subsequently, pregnant rats were treated acutely with either EtOH or methylazoxymethanol (MAM), a known neuroteratogen, and the numbers of immunoreactive LHRH cells were counted. On gestation day 22, LHRH-positive cell numbers were significantly fewer than control numbers in both EtOH- and MAM-exposed offspring. On postnatal day 60, cell numbers in EtOH-exposed offspring did not differ from control numbers, whereas cells in MAM-exposed offspring remained significantly reduced. In controls, there were 40% fewer LHRH-positive cells on postnatal day 4 than in late gestation or at maturity. We conclude that 1) acute exposure to a high dose of EtOH at a critical time in early gestation can alter the expression of LHRH in late gestation; 2) exposure to MAM in the same period alters LHRH expression before birth and in the adult; and 3) in the early postnatal period, LHRH expression decreases profoundly.

Topics: Animals; Body Weight; Brain; Cell Count; Cell Division; Ethanol; Female; Gonadotropin-Releasing Hormone; Methylazoxymethanol Acetate; Neurons; Organ Size; Pilot Projects; Pituitary Gland; Pregnancy; Rats; Teratogens

The effect on colon and liver carcinogenicity in rats of a single X-irradiation exposure given either before or after methylazoxymethanol (MAM) acetate was studied in ACI/N rats of both sexes. A single dose of X-irradiation (3 Gy) was administered either 3 months before or after three weekly s.c. injections of MAM acetate (25 mg/kg body weight). At 365 days after the start, the incidence and multiplicity of MAM acetate-induced intestinal tumors were enhanced by X-irradiation either prior to or after the MAM acetate treatment. In addition, X-irradiation before MAM acetate increased the incidence of hepatocellular foci in either sex. In females, X-irradiation either before or after MAM acetate exposure decreased intestinal tumorigenesis. These findings suggest an apparent synergism of these agents in intestinal carcinogenesis of male rats.

Topics: Animals; Body Weight; Cocarcinogenesis; Colonic Neoplasms; Female; Liver; Liver Neoplasms, Experimental; Male; Methylazoxymethanol Acetate; Neoplasms, Radiation-Induced; Organ Size; Rats; Rats, Inbred ACI

Curcumin, a major yellow pigment of turmeric obtained from powdered rhizomes of the plant Curcuma longa Linn., is commonly used as a coloring agent in foods, drugs and cosmetics. Ascorbyl palmitate is a lipid soluble derivative of ascorbic acid. Both curcumin and ascorbyl palmitate have antioxidant activity and are potent inhibitors of 12-O-tetradecanoyl-phorbol-13-acetate-induced tumor promotion in mouse skin. The effects of dietary curcumin and ascorbyl palmitate on azoxymethanol (AOM)-induced hyperproliferation of colonic epithelial cells and the incidence of focal areas of dysplasia (FADs) were evaluated in female CF-1 mice fed an AIN 76A diet. Subcutaneous injections of AOM (10 mg/kg body wt. once weekly for 6 weeks) caused hyperplasia and the formation of FADs in the colon. Administration of 2% curcumin in the diet inhibited AOM-induced formation of FADs while administration of 2% ascorbyl palmitate in the diet did not demonstrate inhibition. This result suggests that dietary curcumin may inhibit AOM-induced colonic neoplasia in mice.

Topics: Animals; Antioxidants; Ascorbic Acid; Body Weight; Colon; Curcumin; Diet; Epithelium; Female; Methylazoxymethanol Acetate; Mice; Tetradecanoylphorbol Acetate

Micrencephalic neonatal pups were obtained from pregnant Crj:CD (SD) rats once treated with 5 mg/kg of N-methyl-N-nitrosourea (MNU) or 40 mg/kg of methylazoxymethanol (MAM) on day 12, 13, 14 or 15 of gestation (vaginal plug = day 0). They were reared by their own mothers and were subjected to various neurobehavioral tests during the suckling period, days 0 to 22 after birth. The brain weights in the MNU- and MAM-treated pups on postnatal day 22 were significantly less than those in the control pups. These micrencephalic pups were retarded in neurobehavioral ontogeny. By several tests, each of them showed an impaired performance such as paired limb movement, clumsy locomotion or hyperreflexive reaction. These behavioral disorders appeared different according to the day of treatment, without any substantial difference between the test compounds, MNU and MAM. The findings suggest that the different neurobehavioral characteristics in the micrencephalic pups may reflect their different brain disorders induced by the test compounds given on the different period of the treatment.

Topics: Animals; Behavior, Animal; Body Weight; Brain; Female; Locomotion; Male; Methylazoxymethanol Acetate; Methylnitrosourea; Microcephaly; Organ Size; Pregnancy; Prenatal Exposure Delayed Effects; Rats; Rats, Inbred Strains; Rodent Diseases; Swimming

Benzylselenocyanate (BSC), a novel organoselenium compound, has been found to inhibit azoxymethane (AOM)-induced colon carcinogenesis in rats during initiation. To investigate its mechanism of action, we examined the effects of BSC feeding on the following parameters: (a) metabolism of [14C]AOM to 14CO2 in vivo; (b) metabolic activation of AOM to MAM and of MAM to formic acid and methanol by rat liver microsomes in vitro; and (c) AOM-induced DNA methylation in rat livers and colons. Five-week-old male F344 rats were fed modified (23% corn oil) AIN-76A diets containing 0 (control), 25, or 50 ppm of BSC or benzylthiocyanate (BTC), a sulfur analogue of BSC which does not inhibit the colon carcinogenicity of AOM. After 3 weeks, rats were either sacrificed for the isolation of liver microsomes or were given 15 mg/kg of [14C]AOM s.c. to determine the rate of carcinogen metabolism in vivo. No difference in [14C]AOM metabolism was found between rats fed the BTC diets and those fed the control diet. In contrast, the rate of [14C]AOM metabolism, as determined by exhaled radioactivity, was 2-3 times higher in rats fed the BSC diets. While liver microsomes from rats fed the BTC diets metabolized AOM and MAM at rates not significantly different from those obtained with control liver microsomes, the metabolic activation of AOM as well as of MAM was stimulated severalfold when assayed with liver microsomes from rats fed the BSC diets. An increase in total liver cytochrome P-450 was also observed in the BSC-fed rats. Following the administration of 15 mg/kg AOM, significantly less O6-methylguanine and 7-methylguanine was present in the colon DNA from rats consuming the BSC diets than in rats fed the BTC or control diets. The body weight gains of rats fed the 25- and 50-ppm BSC-containing diets for 3 weeks were less (27 and 43%, respectively) than those of rats fed either the control or BTC-containing diets. These results indicate that dietary BSC significantly induces the hydroxylation of AOM and the oxidation of MAM in rat liver. An increase in the rates of AOM and MAM metabolism in the liver due to enzyme induction by BSC will result in decreased delivery of MAM to the colon via the bloodstream. This will be reflected in decreased DNA alkylation, as observed, and is likely to be a major factor in the inhibition of AOM-induced colon carcinogenesis by BSC.

Topics: Animals; Azoxymethane; Body Weight; Colonic Neoplasms; Cyanates; Cytochrome P-450 Enzyme System; DNA; Drug Screening Assays, Antitumor; Guanine; Male; Methylation; Methylazoxymethanol Acetate; Microsomes, Liver; Organoselenium Compounds; Rats; Rats, Inbred F344; Selenium

Dietary quercetin (QU) and rutin (RU), phenolic flavonoids commonly found in many fruits and vegetables, were provided to CF1 female mice for 50 weeks to assess the ability of these compounds to inhibit azoxymethanol (AOM)-induced colonic neoplasia. In addition to a control group fed an AIN 76A diet, five other groups received that diet to which was added either 0.1, 0.5 or 2.0% QU and 1.0 or 4.0% RU. Acute studies revealed that, among saline controls, no alteration of any proliferative parameters of colonic epithelial cells was observed among those groups receiving any dose of QU or RU. However, among the AOM-treated mice, both 2% QU and 4% RU significantly reduced hyperproliferation and inhibited the shift of S-phase cells to the middle and upper portion of crypts. Moreover, mice fed these concentrations of QU and RU had significantly fewer AOM-induced focal areas of dysplasia (FADs) than those fed the control diet (0.2 +/- 0.4 and 0.4 +/- 0.5 versus 3.6 +/- 2.3 respectively). Tumors occurred more frequently in the distal half of the colon, regardless of treatment. Compared with controls, mice fed 2% QU had a significantly reduced tumor incidence (25.0% versus 5.9%, P = 0.03). Those fed 4% RU showed only a trend toward inhibition (25% versus 9.7%, P = 0.11). Nevertheless, both 2% QU and 4% RU suppressed tumor multiplicity, i.e. fewer tumors/animal arose in these groups than in the AOM-treated control mice (1.2 versus 2.3, P = 0.005; 1.1 versus 2.3, P = 0.003 respectively). Clearly, QU and RU exhibit significant activity in reducing AOM-induced hyperproliferation of colonic epithelial cells and FAD incidence. This behavior successfully forecast the ability of both flavonoids to suppress tumor multiplicity and ultimately tumor development.

Topics: Animals; Body Weight; Carcinogens; Cell Division; Colonic Neoplasms; Female; Methylazoxymethanol Acetate; Mice; Quercetin; Rutin

Prenatal administration of methylazoxymethanol acetate (MAM), which kills neuroblasts undergoing mitosis, was used to lesion striatal somatostatin neurons. Previous [3H]thymidine autoradiographic studies had indicated that striatal somatostatin neurons undergo their final mitotic division at Gestational Days (G) 15 and 16. Therefore, pregnant Sprague-Dawley rats received an intraperitoneal injection of MAM (25 mg/kg) on G15. Neurochemical and histological examination of the mature offspring indicated the loss of half the striatal aspiny interneurons in which somatostatin, neuropeptide Y, and NADPH diaphorase coexist, with relative sparing of the cholinergic interneurons and medium spiny projection cells. This prenatal MAM treatment was without apparent effect on the patch-matrix organization of the striatum.

Topics: Animals; Body Weight; Catecholamines; Corpus Striatum; Female; Fetus; Methylazoxymethanol Acetate; Mitosis; Neurons; Rats; Rats, Inbred Strains; Somatostatin

Various doses (0, 1, 5, 10, 15, 20, or 25 mg/kg) of methylazoxymethanol acetate (MAM), a potent alkylating agent, were injected singly into pregnant rats intraperitoneally on day 15 of gestation. Relationships between brain weights and neurochemical changes in the cerebral hemispheres (CHs; cerebral cortex and subjacent white matter, hippocampus, amygdala) and remainder of the brain (BGDM; basal ganglia, diencephalon, and mesencephalon) were examined at 60 days of age in offspring; varying degrees of microencephaly were observed. Dose-dependent reductions in the weights of CH and BGDM were observed. Reductions in total DNA content positively correlated with decreases in brain weights also observed. Dose-dependent elevations of noradrenaline (NA) and dopamine (DA) were observed in CH at MAM levels 10 mg/kg and above; dose-dependent elevations of 5-hydroxytryptamine (5-HT) were observed at 15 mg/kg and above; and in BGDM at 20 mg/kg and above dose-dependent elevations for NA and 5-HT were observed; dose-dependent elevations at 15 mg/kg and above were observed for DA. Monoamine concentrations were negatively correlated with brain weights or total DNA contents. NA and DA concentrations increased to the extent of approximately 1.3 times of control at a time when an 18% loss of CH weight was noted in animals treated with 10 mg/kg MAM. It is suggested that the above variables might be appropriately sensitive neurochemical markers for detecting minor developmental anomalies in the brain.

Topics: Alkylating Agents; Animals; Azo Compounds; Body Weight; Brain; DNA; Dopamine; Dose-Response Relationship, Drug; Female; Methylazoxymethanol Acetate; Norepinephrine; Organ Size; Pregnancy; Rats; Serotonin

The influence of diet restriction on induction of intestinal tumors in Sprague-Dawley rats by two unique carcinogenic agents was investigated: methylazoxymethanol acetate [(MAM) CAS: 592-62-1], which requires metabolic activation, and N-methylnitrosourea [(MNU) CAS: 684-93-5], which is a direct-acting carcinogen. Most of the tumors induced by MAM developed in the small intestine and less frequently in the colon, but MNU produced tumors predominantly in the colon. Among rats fed the restricted diet (12 g/day), the production of tumors by MAM was significantly reduced compared to that of counterpart rats on the ad libitum diet. However, dietary restriction did not modify the production of tumors in rats by MNU. The same relationship of diet restriction to tumor induction was demonstrable when MAM and MNU were administered to the same test rats: Numbers of MAM-related tumors, especially in the small intestine, were reduced and numbers of MNU-related tumors in the colon were unchanged. Dietary restriction modified the tumorigenic response of rats to MAM but not to MNU.

Topics: Animals; Body Weight; Colonic Neoplasms; Diet; Intestinal Neoplasms; Intestine, Large; Intestine, Small; Male; Methylazoxymethanol Acetate; Methylnitrosourea; Rats; Rats, Inbred Strains

Methylazoxymethanol (MAM), an alkylating agent which kills dividing cells, produces microcephaly when administered to rats at 15 days gestation. Rats treated prenatally with MAM were tested on a variety of behavioral tests. The MAM-treated animals performed better than controls in the acquisition of a food-reinforced operant response, but poorer than controls on a passive avoidance procedure. When required to reverse the passive avoidance procedure by actively avoiding the portion of a chamber that was associated with shock, MAM-treated rats performed better than controls. The MAM-treated rats were microcephalic and were also hyperactive compared to controls. It was postulated that the behavioral changes observed in the conditioning tasks may be attributable to hyperactivity. A possible neurochemical basis for this hyperactivity is discussed.

Topics: Alkylating Agents; Animals; Avoidance Learning; Azo Compounds; Body Weight; Conditioning, Operant; Female; Methylazoxymethanol Acetate; Motor Activity; Pregnancy; Prenatal Exposure Delayed Effects; Psychomotor Performance; Rats; Rats, Inbred Strains

Topics: Abnormalities, Drug-Induced; Analysis of Variance; Animals; Azo Compounds; Behavior, Animal; Body Weight; Brain; Carcinogens; Female; Fetus; Learning; Male; Methylazoxymethanol Acetate; Motor Activity; Organ Size; Pregnancy; Rats; Rats, Inbred Strains; Reproduction