Chromosome aberrations in spermatogonia and sperm abnormalities in Curacron-treated mice

Curacron is an organophosphorus pesticide widely used in cotton fields. In order to assay its mutagenic potential in mammalian germ cells chromosomal aberrations in spermatogonial cells and sperm abnormalities were examined in mice after Curacron treatment. For studying chromosomal aberrations mice were treated both acutely (single treatment) and subacutely (for 5 consecutive days) with 3 dose levels of Curacron, 12, 36 and 72 mg/kg. Curacron was found to produce a significant increase in structural chromosomal aberrations after acute and subacute treatments. This increase was dose-dependent. A dose-dependent inhibition in mitotic activity in spermatogonia was also found. For studying sperm abnormalities mice were treated for 5 consecutive days with 20, 40 and 60 mg/kg. Morphological sperm abnormalities increased significantly after treatment with Curacron. The increase was dose-dependent. An inhibition of 40.2% in sperm count and of 74.5% in sperm motility occurred after treatment with 60 mg/kg Curacron. These results show that Curacron has a damaging effect on spermatogonial cells as well as on sperm morphology.     

Genetically Modified DR5-Specific TRAIL Variant DR5-B Revealed Dual Antitumor and Protumoral Effect in Colon Cancer Xenografts and an Improved Pharmacokinetic Profile

Despite the weak clinical efficacy of TRAIL death receptor agonists, a search is under way for new agents that more efficiently activate apoptotic signaling. We previously created a TRAIL DR5-selective variant DR5-B without affinity for the DR4, DcR1, DcR2, and OPG receptors and increased proapoptotic activity in tumor cells. Here we showed that DR5-B significantly inhibited tumor growth in HCT116 and Caco-2 but not in HT-29 xenografts. The antitumor activity of DR5-B was 2.5 times higher in HCT116 xenografts compared to TRAIL. DR5-B at a dose of 2 or 10 mg/kg/d for 10 days inhibited tumor growth in HCT116 xenografts by 26% or 50% respectively, and increased animal survival. Unexpectedly, DR5-B at a higher dose (25 mg/kg/d) inhibited tumor growth only during the first 8 days of drug exposure, while at the end of the monitoring, no effect or even slight stimulation of tumor growth was observed. The pharmacokinetic parameters of DR5-B were comparable to those of TRAIL, except that the half-life was 3.5 times higher. Thus, enhancing TRAIL selectivity to DR5 may increase both antitumor and proliferative activities depending on the concentration and administration regimens.     

Relative efficacy of short-term tests in detecting genotoxic effects of cadmium chloride in mice in vivo

The ability of intraperitoneally administered cadmium chloride (0.42-6.75 mg/kg) to induce genotoxic damage in somatic and germ cells of mice was evaluated using chromosomal aberrations, sister-chromatid exchanges (SCE), micronuclei and sperm-head abnormalities as end-points. A significant increase in the frequency of chromosomal aberrations and SCEs was observed in almost all treated series when compared to the negative control. Micronucleus formation in polychromatic erythrocytes was not affected significantly except at the highest concentration used (6.75 mg/kg). Significant differences were observed in the frequency of sperm with abnormal head morphology at all concentrations tested except the lowest one. The clastogenic effects of cadmium chloride in both somatic and germinal cells are found to depend directly on the concentrations used.     

Genotoxic effect of 2,4-dichlorophenoxy acetic acid and its metabolite 2,4-dichlorophenol in mouse.

The cytogenetic effect of 2,4-dichlorophenoxy acetic acid (2,4-D) and its metabolite 2,4-dichlorophenol (2,4-DCP) was studied in bone-marrow, germ cells and sperm head abnormalities in the treated mice. Swiss mice were treated orally by gavage with 2,4-D at 1.7, 3.3 and 33 mg kg(-1)BW (1/200, 1/100 and 1/10 of LD(50)). 2,4-DCP was intraperitoneally (i.p.) injected at 36, 72 and 180 mg kg(-1)BW (1/10, 1/5, 1/2 of LD(50)). A significant increase in the percentage of chromosome aberrations in bone-marrow and spermatocyte cells was observed after oral administration of 2,4-D at 3.3 mg kg(-1)BW for three and five consecutive days. This percentage increased and reached 10.8+/-0.87 (P<0.01) in bone-marrow and 9.8+/-0.45 (P<0.01) in spermatocyte cells after oral administration of 2,4-D at 33 mg kg(-1)BW for 24 h. This percentage was, however, lower than that induced in bone-marrow and spermatocyte cells by mitomycin C (positive control). 2,4-D induced a dose-dependent increase in the percentage of sperm head abnormalities. The genotoxic effect of 2,4-DCP is weaker than that of 2,4-D, as indicated by the lower percentage of the induced chromosome aberrations (in bone-marrow and spermatocyte cells) and sperm head abnormalities. Only the highest tested concentration of 2,4-DCP (180 mg kg(-1)BW, 1/2 LD(50)) induced a significant percentage of chromosome aberrations and sperm head abnormalities after i.p. injection. The obtained results indicate that 2,4-D is genotoxic in mice in vivo under the conditions tested. Hence, more care should be given to the application of 2,4-D on edible crops since repeated uses may underlie a health hazard.     

Effects of dexamethasone and chlorpromazine treatment on X-Y dissociation and multinucleated giant cell formation in hyperthermic mice

The effects of anti-stress drugs on X-Y dissociation and multinucleated giant cell formation in the testes of hyperthermic mice were determined for the possible use of such drugs in animal production. Mice were injected with dexamethasone 1 mg/kg or 2 mg/kg, chlorpromazine 1 mg/kg or 4 mg/kg, or 0.85% saline, and heat stressed for 4 d at 35 +/- 1 C and 65 +/- 1 % relative humidity. Mice were killed 5 d after stress, and the testes processed for observation of meiotic chromosomes and testicular histology. Heat stress caused a significant increase in X-Y bivalent dissociation in diakinesis-metaphase I spermatocytes and a significant increase in formation of multinucleated giant cells. Drug treatment increased X-Y dissociation but had no significant effect on giant cell formation.     

Teratogenicity of carbamazepine in rats

The teratogenicity of carbamazepine (CBZ) was investigated in Sprague-Dawley CD rats at doses of 0, 200, 400, and 600 mg/kg administered by gavage in corn oil on days 7-18 of gestation in a dosage volume of 2 ml/kg. The CBZ-600 dose was maternally toxic in that dams in this group weighed 30.6% less than controls by E20. This group had significantly increased resorptions, reduced live fetal weight (51.6% less than controls), and increased skeletal and visceral abnormalities. The CBZ-400 dose also significantly reduced maternal weight gain during gestation to 26.6% less than controls by E20. No significant increase in resorptions occurred in this group; live fetuses weighted 42.9% less than controls and showed an increase in visceral, but not skeletal, abnormalities. The CBZ-200 dose did not significantly affect maternal weight gain or increase resorptions or fetal abnormalities but did reduce fetal body weight (20.3% less than controls). Maternal serum total CBZ concentrations 1 hr after the final dose were 22.9, 27.9, and 34.4 micrograms/ml for the 200, 400, and 600 mg/kg groups, respectively. These levels were little changed 6 h post-treatment. CBZ was 65-70% serum protein bound across dose groups. Human therapeutic levels of CBZ are 4-12 micrograms/ml and the drug is typically 80% serum protein bound. This suggests that abnormalities in rats occur at concentrations well above the human therapeutic range. However, a no-effect level was not found for fetal body weight. Further experiments will be required to determine how much lower doses will need to be in order to find a no-effect level for fetal body weight. Nevertheless, the present data suggest that CBZ is not potent at inducing malformations in rats.     

Anti-tumor activity of new orally bioavailable 2-amino-5-(thiophen-2-yl)benzamide-series histone deacetylase inhibitors, possessing an aqueous soluble functional group as a surface recognition domain

New orally bioavailable 5-(thiophen-2-yl)-substituted 2-aminobenzamide-series histone deacetylase inhibitors were synthesized. These compounds possess a morpholine or piperadine-derived moiety as an aqueous soluble functional group. Among them, 8b, having a 4-ethyl-2,3-dioxopiperazine-1-carboxamide group as a surface recognition domain, showed promising inhibitory activities against HCT116 cell growth and HDAC1/2. Notably, unlike MS-275, this compound did not induce apoptosis in the cell cycle tests. We therefore conducted antitumor tests of 8b and MS-275 against HCT116 cell xenografts in nude mice. Compound 8b reduced the volume of tumor mass to T/C: 60% and 47% at 45 and 80mg/kg over 16days, respectively. These values were comparable to the rate (T/C: 51% at 45mg/kg) for MS-275. Furthermore, 8b, at neither 45 nor 80mg/kg, induced the weight loss which was observed in the mice given MS-275 at 45mg/kg.     

Association of cyclophosphamide-induced male-mediated, foetal abnormalities with reduced paternal germ-cell apoptosis

To investigate the mechanism by which malformed offspring can result from the exposure of males to mutagens, we treated adult male rats with 0, 1.4, 3.4 or 5.1 mg/kg cyclophosphamide, 6 days per week for 9 weeks, a treatment regimen known to induce heritable abnormalities. Testis samples from some of the animals were then collected for fixation in Carnoy's fluid and subsequent analysis of germ-cell apoptosis and proliferation. The remainder were mated, resulting in a greater than 11-fold increase in the proportion of abnormal offspring produced in the 5.1 mg/kg group. The number of apoptotic cells per stage XII/XIII tubular cross-section decreased with increasing dose, significantly so at 5.1 mg/kg (P<0.05). No statistically significant effect was found on spermatocyte numbers at this dose, indicating that a reduction in the amount of cells available to undergo apoptosis cannot explain the decrease. The inappropriate survival of damaged germ-cells caused by a lowering of the incidence of apoptosis may, therefore, account for the rise in the proportion of foetal malformations.     

Induction of micronuclei by vinyl acetate in mouse bone marrow cells and cultured human lymphocytes

A dose-dependent increase in micronucleated polychromatic erythrocytes was observed in the bone marrow of male C57B1/6 mice 30 h after a single intraperitoneal injection of vinyl acetate (250, 500, 1000 or 2000 mg/kg b.wt.; (9-14 animals per group). The effect was statistically significant at 1000 mg/kg (1.33 +/- 0.29% vs. 0.6 +/- 0.10% in olive oil-treated controls) and at 2000 mg/kg (1.57 +/- 0.19%) of vinyl acetate. These doses were fatal to 6 (1000 mg/kg) and 8 (2000 mg/kg) out of 14 animals in both groups. The ratio of polychromatic to normochromatic cells decreased as a function of vinyl acetate dose. Cyclophosphamide (20 mg/kg), used as a positive control chemical, induced a clear increase in micronucleated polychromatic erythrocytes (2.07 +/- 0.20%). None of the treatments affected the number of micronuclei in normochromatic erythrocytes. In human whole-blood lymphocyte cultures, micronucleus induction by a 48-h treatment with vinyl acetate (0.125, 0.25, 0.5, 1 and 2 mM; 24 h after culture initiation) was studied in lymphocytes with preserved cytoplasm from smear slides prepared by a method involving the removal of erythrocytes at harvest by sodium cyanide treatment to improve preparation quality. The frequency of micronucleated lymphocytes reached a peak at 0.5 mM (3.2 +/- 1.0% vs. 0.9 +/- 0.1% in control cultures) and 1 mM (3.1 +/- 0.7%), with a decline at 2 mM probably because of a toxic effect resulting in mitotic inhibition.     

Cytotoxicity of zinc chloride in mice in vivo

Intraperitoneal administration of zinc chloride (ZnCl2) to Swiss albino mice in vivo induced a significant (p less than or equal to 0.05) increase in the frequencies of chromosomal aberrations of the bone-marrow cells at all concentrations used following acute (7.5, 10, 15 mg/kg body weight) and chronic (2.0, 3.0 mg/kg body wt) treatment. The degree of clastogenicity was directly proportional to the concentrations (p less than or equal to 0.05, trend test) and indirectly to the period of treatment (p less than or equal to 0.05, ANOVA test). It induced a dose-dependent, statistically significant increase (Mann-Whitney U statistics, Student's t-test) in sperm-head abnormalities. The data designate ZnCl2 as a potent clastogen and as a toxic chemical at the concentrations used.     

Immuno-potentiating effects of the antler-shaped fruiting body of Ganoderma lucidum (Rokkaku-Reishi)

The immuno-potentiating effects of the antler-shaped fruiting body of Ganoderma lucidum (Rokkaku-Reishi, RR), which has been used as a traditional supplement for human health, were investigated in mice. BALB/c mice were administered orally with RR for 3 days at a dose of 50 mg/kg or 500 mg/kg, and interferon-gamma (IFN-gamma) production by splenocytes in response to lipopolysaccharide (LPS) was examined on day 4. The oral administration of 500 mg/kg of RR resulted in a significant increase (p<0.05) in IFN-gamma production. Stimulation of splenic adherent cells from these mice with LPS also resulted in a significant increase (p<0.05) in interleukin-12 (IL-12) production compared with that from the control mice, suggesting that splenic macrophages were activated by RR administration. Furthermore, 500 mg/kg of RR administered for 14 days resulted in a significant increase (p<0.05) in IFN-gamma production by splenocytes in response to both LPS and concanavalin A (Con A). These results suggest that not only splenic macrophages but also T cells were activated by the long-term treatment with RR in vivo. On the other hand, the production of interleukin-4 (IL-4), which is known as an allergic disease-related cytokine, was not affected by the long-term treatment with RR. Our results suggest that the oral administration of RR resulted in Th1-associated immuno-potentiating activities in vivo.     

Induction of sperm abnormalities in mice by ifosfamide and trofosfamide

The antitumor drugs ifosfamide (IF) and trofosfamide (TF) were evaluated for their capability to induce sperm abnormalities in (C3H X C57BL/6)F1 mice. A statistically significant increase in teratospermia was observed at the 35th day after 5 daily consecutive intraperitoneal injections of the drugs at doses of 25, 50, 100 mg/kg b.w. of TF and 100 mg/kg b.w. of IF. Thus, IF and TF are able to interfere with the differentiation process of spermatogenic cells.     

Developmental toxic effects of antifungal flusilazole administered by gavage to mice

BACKGROUND: The developmental toxicity of flusilazole was studied in CD-1 mice after oral administration. METHODS: Pregnant mice were given flusilazole at doses of 0 (corn oil), 10, 20, and 40 mg/kg/day, by gavage, on gestational days (GD) 6-15. RESULTS: Maternal toxicity, as evidenced by reduction in body weight gain and signs of toxicity, was observed at the middle- and high-dose groups. No significant incidence of resorptions or death was observed in any of dose groups. There was a pronounced reduction in fetal weight, which was significantly lower than control from 20 and 40 mg/kg/day. There was no significant increase in the incidence of fetuses with external or visceral malformations in any of dose groups, but there was a significant increase in the incidence of skeletal malformations was observed at 20 and 40 mg/kg/day. CONCLUSIONS: The results of this study reported marked maternal toxicity, growth retardation, and skeletal abnormalities in the mid- and high-dose groups. It seems likely that marked maternal toxicity contributed to the observed alterations in fetal growth retardation and skeletal development. The no-observed-effect level in the present study for maternal and developmental toxicity was 10 mg/kg/day.     

Novel combination of 2-methoxyestradiol and cyclophosphamide enhances the antineoplastic and pro-apoptotic effects on S-180 ascitic tumour cells

Sarcoma 180 (S-180) tumour cell line is a stable murine tumour cell line with 98–99% stumour takes capacity in Swiss albino mouse - Mus musculus. 2 Methoxyestradiol (2ME) - a promising anti-neoplastic and anti-angiogenic agent, showed toxicity to host body in higher concentration. Cyclophosphamide (CP), the anti-neoplastic agent has long been used as a chemotherapeutic drug for treatment of different cancers. Our studies have shown that the combination effect of 2ME and CP on S-180 tumour cell line is anti-proliferative and less toxic. The treatment with lower concentrations of 2ME and CP (6.5 mg 2ME/kg body weight + 75 mg CP/kg body weight) antagonistically increased the life span of tumour bearing mice and synergistically inhibited the viable cell population. 2ME or CP treatment individually induces G2/M arrest. The combination treatment of 2ME + CP (6.5 mg 2ME/kg body weight + 75 mg CP/kg body weight) produced a significant increase of cells in the G₀ which is the indication of cell arrest or apoptosis. Reduction of cell viability by 2ME + CP treatments is due to apoptotic cell death. This combination therapy produced a significant inhibitory effect of cell proliferation and augmentation of cell accumulation in the G₀ phase (i.e. apoptosis). Apoptosis is validated by Fluorescence staining of control and treated S-180 tumour cells with Acridine Orange and EtBr dye. Moreover, a steady increase in the frequency of complex chromosomal aberrations (i.e. tri-, qudri-radial translocations) in tumour cells was noted in that particular concentration of combination therapy treated series along with the increase in dead cell frequency and tumour regression pattern. It is assumed that, these chromosomal abnormalities or damages recorded in higher frequency prevent the affected metaphases to enter into the next cell cycle through apoptosis or necrosis. This study introduces a novel combination, where this particular concentration of 2ME + CP (i.e. 6.5 mg 2ME/kg body weight + 75 mg CP/kg body weight) not only enhanced the life span of tumour bearing mouse and decreased the tumour volume antagonistically but also inhibited the viable cell population synergistically, which could serve as a potential effective regimen for cancer treatment.     

In vivo growth inhibitory effect of Withania somnifera (Ashwagandha) on a transplantable mouse tumor, Sarcoma 180.

Withania somnifera is a medicinal plant used in the treatment of a variety of ailments in the Ayurvedic system. Alcoholic extract of the root of the plant was injected(ip) at daily doses of 200 to 1000 mg/kg body wt for 15 days starting from 24 hr after intradermal inoculation of 5 x 10(5) cells of S-180 in BALB/c mice. Solid tumor growth was monitored for 100 days. Doses of 400 mg/kg and above produced complete regression of tumor after an initial growth, the percentage of complete response (CR) increasing with increasing drug dose. A 55% CR was obtained at 1000 mg/kg drug administration, but this dose also produced some mortality among the animals. A significant increase in the volume doubling time and growth delay was seen when the drug dose was increased from 500 to 750 mg/kg body wt, but further increase in drug dose to 1000 mg/kg did not produce any significant increase in these responses. Cumulative doses of 7.5 to 10 g at daily doses of 500 or 750 mg/kg seems to produce a good response in this tumor.     

Adverse effects of cyclophosphamide on progeny outcome can be mediated through post-testicular mechanisms in the rat.

Previous studies from our laboratory have suggested that, in addition to an effect on spermatozoa in the testis, cyclophosphamide may have an adverse effect on spermatozoa after they leave the testis, during epididymal transit. To elaborate on this post-testicular effect on germ cells and to determine at which site(s) in the epididymis germ cells are most sensitive to cyclophosphamide treatment, three experiments were undertaken. First, the time course of the effect of treatment of male rats with cyclophosphamide on the outcome of their progeny was determined. Male rats were treated daily by gavage with saline or one of two doses of cyclophosphamide (6.8 mg/kg or 10.0 mg/kg) for 1, 4, or 7 days. At the end of each treatment period, males were mated to assess the effect on pregnancy outcome. No effect was observed on pre-implantation loss at any time among any of the groups, but there was a time-dependent and dose-related increase in post-implantation loss. Post-implantation loss was significantly increased after 4 days of treatment and reached nearly 40% after 7 days of drug exposure (10.0 mg/kg). Second, the effect of treatment with single high doses of cyclophosphamide was studied. Male rats were treated with a single dose of cyclophosphamide (10, 30, or 70 mg/kg) and bred 1 day and 4 days post-treatment. No significant change in pre-implantation loss was observed at either time point; no change in post-implantation loss was found after 1 day post-treatment. However, a significant increase in post-implantation loss was observed in the two high-dose groups 4 days post-treatment.(ABSTRACT TRUNCATED AT 250 WORDS)     

Cyclophosphamide teratogenesis: evidence for compensatory responses to induced cellular toxicity

Cyclophosphamide (CP) administered ip to pregnant mice on day 10 of gestation (day of plug = day 0) is teratogenic (exencephaly, cleft palate, and limb malformations) at 20 mg/kg and embryolethal at higher doses. In the present study, CP was administered at 1, 5, 10, or 20 mg/kg on day 10 of gestation. Embryos were removed at 8 and 28 hr postdosing, and two embryos from each litter were immediately stained with Nile blue sulfate (NBS) to identify areas of cell death. The remaining embryos were frozen and forelimb buds subsequently removed for flow cytometric (FCM) analysis of the cellular DNA synthetic cycle. Additional litters were examined near term (day 17) for morphological abnormalities; these data were correlated with embryonic toxicity as detected by NBS staining and FCM analysis. Only the highest dose produced malformations. In marked contrast, a dose-related increase in the percentage of limb bud cells in the S (DNA synthetic) phase of the cell cycle was detectable at all doses. Inhibition of DNA synthesis was detected at all doses 8 hr post exposure and persisted through 28 hr for doses greater than or equal to 10 mg/kg. NBS staining indicated increased cell death in the alar plate of the neural tube 28 hr after exposure to 10 mg/kg CP and generally increased cell death in areas of rapid cell proliferation throughout the embryo at 20 mg/kg. The absence of an overt teratogenic response at dose levels that produced significant perturbation of the cell cycle indicates that a measure of embryonic damage can be compensated for or repaired. The implications of these findings for the existence of thresholds in developmental toxicity are discussed.     

Effect of cortisol on the cellular proliferation and maturation in the cerebrum and the cerebellum of the rat: Importance of the age of the animals at the beginning of treatment]

Young rats were given either a single subcutaneous injection (1 mg at 0, 1, 4 or 8 days), or four consecutive daily injections (0.2 mg/day between 0 and 3 days; 0.4 mg/day between 4 and 7 days; 0.6 mg/day between 8 and 11 days) of cortisol acetate in order to test the influence of age on the action of corticosteroids on the biochemical maturation of the cerebrum and cerebellum in terms of their DNA, RNA, and protein contents. The results showed that: 1 The diminution of the DNA content at 35 days was greater in the cerebellum (- 16 to - 32%) than in the cerebrum (- 9 to 20%); the DNA content of the cerebrum was more affected by treatment at birth, whereas that of the cerebellum was more affected by the delayed treatments. Results were different when expressed in terms of reduction of the normal increase: the gain of DNA decreased more in the cerebrum (-70%) than in the cerebellum (-40%); but the most delayed treatment induced a greater effect in both organs. These abnormalities were not always accompanied by a significant decrease of the body weight. 2 Generally, the treatments led to an increase of the mean cell territory, expressed either in terms of decrease of the DNA concentration, or in terms of increase of the organ weight/DNA ratio. Moreover, the increase of the RNA/DNA and the protein/DNA ratios constituted an indication of an accelerated cellular maturation.     

Cyclosporine lowers seizure threshold in an experimental model of electroshock-induced seizures in Munich-Wistar rats

We have developed a model of cyclosporin A (CsA) central nervous system toxicity in the Munich-Wistar rat in which CsA, 20 mg/kg/day i.p., produces significant EEG abnormalities and mortality. In the present study we used cohorts of Munich-Wistar rats to assess effects of CsA on the threshold for tonic-clonic electroshock-induced seizures. Rat cohorts were begun on cremephore, CsA-10 mg/kg/day, or CsA-20 mg/kg/day. On day 7 and day 14 of the dosing protocol, cohorts of animals were exposed to maximal electroshock (MES) using a minimal staircase method within each cohort. Multiple logistic regression models were used to determine differences between groups on the relative odds of producing a MES-induced seizure while controlling for other variables. Seizure threshold was significantly affected by shock amperage and body weight, but not by SUN, creatinine, bilirubin, sodium, potassium, weight loss or day the shock was delivered. The odds ratios of seizure induction in the CsA-treated groups versus placebo group were 1.91 for CsA-10 mg/kg/day and 3.63 for CsA 20-mg/kg/d, both statistically significant. These results suggest that cyclosporine lowers seizure threshold and probably increases susceptibility to seizures, the etiology of which may be multifactorial clinically.     

Effects of chemically- and environmentally-induced hypothermia on micronucleus induction in rats

We investigated micronucleus induction in rats treated with chlorpromazine and reserpine, drugs that induce hypothermia. We administered chlorpromazine (31.3--250mg/kg) or reserpine (500--2000 mg/kg) intraperitoneally and measured temperature rectally. Chlorpromazine at 62.5-250mg/kg and reserpine at all doses significantly decreased rectal temperature, although the hypothermic response was weaker than previously reported in mice. Only chlorpromazine at 250mg/kg decreased rectal temperature transiently to <33 degrees C for 20h and induced a statistically significant increase in micronucleated polychromatic erythrocyte frequency. When rats treated with reserpine at 500mg/kg were exposed to an environmental temperature of 16 degrees C for 6, 12, or 24h to keep their body temperature under 33 degrees C, only the 24h treatment group significantly induced micronuclei. In addition, relatively large micronuclei (diameter of micronucleus> or = 1/4 diameter of cytoplasm) accounted for 33.0% of the induced micronuclei, suggesting that hypothermia affected the mitotic apparatus. The hypothermic response to chlorpromazine and reserpine was weaker in rats than in mice, and it was correspondingly more difficult to induce micronuclei in rats with those drugs.