Yoshinobu Iwaki

Full-Field Electroretinography Obtained Using a Contact Lens Electrode with Built-In High-Intensity White-Light-Emitting Diodes Can Be Utilized in Toxicological Assessments in Rats

Full-field electroretinography (ERG) using contact lenses with built-in LED was performed on albino rats, and used to evaluate the visual toxicity of sodium iodate (NaIO3). Experiment 1 was carried out to determine the optimal conditions in rats relating to stimulus intensity, background illumination, and light adaptation period. As a result, we found that a full-field ERG was recorded under the following conditions: stimulus intensity: –3.5 log cd s/m2 in rod response; background intensity and light adaptation period: 10 cd/m2 and 10 min in cone and flicker responses. Experiment 2 was carried out to confirm the usefulness of full-field ERG using rats with retinal toxicities induced by NaIO3. Male rats were given NaIO3 intravenously at a dose of 50 mg/kg. ERG was recorded before administration and after 3, 8, 24 h, and 7 days of administration, and histopathological analysis was conducted after 8 h of administration. The rod response disappeared completely at 3 h, based on a reduced maximal response and oscillatory potentials. On the other hand, cone and flicker responses were still present at 8 h. All responses disappeared on the 7th day. These findings indicate that the retinal toxicity induced by NaIO3 was expressed first in rods, followed by cones. There were no microscopical changes after 8 h of administration, although the rod responses had completely disappeared by this time. These results suggest that full-field ERG in rats using an LED contact lens is useful for the separate evaluation of toxic effects on rods and cones.

Evaluation of Bone Toxicity in Various Bones of Aged Rats

The aim of the present study was to provide a method for evaluating bone toxicity induced by drugs in various bones in aged rats. Male Crl:CD (SD) rats at 46 weeks of age were administered 15 mg/m2 body surface area of doxorubicin, which effects the growth plate in weanling rats, weekly for 9 weeks by intravenous injection, and the femur, sternum, humerus and tibia were examined histopathologically. In the doxorubicin-treated group, thinning of the growth plate was remarkably observed in the proximal tibia and humerus; however, these changes were not observed in other regions. In addition, the osteoclast number per bone perimeter in the proximal tibia was significantly higher than others in control aged rat. Thus, recognizing the various histological reactions related to the time of epiphyseal closure is important for evaluating bone toxicity in aged rats.

Evaluation of Myelotoxicity in Dietary Restricted Rats

The purpose of this study was to clarify the effect of decreased food consumption on evaluation of myelotoxicity in routine general toxicity studies. Male rats were divided into the following 7 groups: 12, 15, and 18 mg/kg 5-fluorouracil (5-FU) treatment groups (FU12, FU15 and FU18); dietary restriction groups (R12, R15 and R18 receiving the same amount of food as the rats in the FU12, FU15 and FU18 groups, respectively); and a nontreated control group (NT). We compared the changes in body weight, hematology and the results of cytological analyses of bone marrow and histopathology among the groups after administration and recovery periods of 14 and 7 days, respectively. At the end of the administration period, the FU15 and FU18 groups showed decreases in many hematologic and bone marrow parameters that were all similar to those in the corresponding dietary restriction groups (R15 and R18). A granulocyte abnormality (polyploidy: frequency of 1% or less) was also observed in all 5-FU treated groups. At the end of the recovery period, increases in the reticulocyte and platelet counts and extramedullary hematopoiesis of the spleen were observed in the 5-FU treated groups. These results indicate that the results of general toxicity studies in rats should be evaluated in consideration of dietary restriction effects when food consumption is decreased at about 30-40% or more. Careful morphological observation of hemocytes would be helpful in distinguishing the effect of a drug from that of dietary restriction in relation to hematological and bone marrow parameters. Performance of a recovery test to determine the reactive response of hematopoiesis is also recommended.

Effect of estrogen on electroretinographic responses in streptozotocin-induced diabetic female rats

The aim of this study is to investigate the effects of estrogen on functional changes in the retinas of streptozotocin (STZ)-induced diabetic rats by using an electroretinography. Female rats were randomly divided into four treatment groups: (1) Control (sham operation and vehicle administration); (2) STZ (sham operation and STZ administration); (3) OVX (ovariectomy and vehicle administration); and (4) OVX + STZ (ovariectomy and STZ administration). Full-field electroretinograms (ERGs) were recorded before OVX and STZ administration and 4 and 12 weeks after STZ administration. At 4 weeks after STZ administration, although there were no differences in the STZ and OVX groups compared with the Control group, the amplitude of the cone-response was significantly lower in the OVX + STZ group than in the Control group (P = 0.013). At 12 weeks after STZ administration, this response showed a similar tendency in the STZ and the OVX + STZ groups. At 12 weeks after STZ administration, the implicit times of OP3 and OP4 and of the cone-response were significantly delayed in the STZ and OVX + STZ groups (OP3: P = 0.030 and 0.050, OP4: P = 0.0060 and 0.0053, cone-response: P = 0.014 and 0.039), compared with in the Control group. Thus, the retinal functions in STZ-induced diabetic female rats were aggravated by OVX. OVX-induced estrogen deficiency resulted in earlier changes in the amplitudes of cone-response, especially in the diabetes, although this is a transient effect and it is difficult to explain. Recognizing the early neurosensory change would enable a better understanding of the effect of estrogen in the retina.

Importance of Starting Age for Myelotoxicity Study in Dietary Restricted Rats

The aim of this study was to prove our hypothesis that adult rats with lowering of body weight gain, rats at 12 weeks of age as an example, are suitable for evaluation of myelotoxicity. Age-related differences between young rats (6-week-old study) and adult rats (12-week-old study) were analyzed in hematological examination values. The data of the young rats were reprinted from our previous report (Miyata et al., 2009) since our hypothesis was verified by comparison with that previous report. Several experimental groups were defined for the 12-week-old study as well as for the 6-week-old study; these included 5-fluorouracil (5-FU) treated groups receiving 12, 15 and 18 mg/kg/day (FU12, FU15 and FU18), pair-feeding groups (R12, R15 and R18 receiving the same amount of food as in the FU12, FU15 and FU18 groups, respectively) and a nontreated control group. Numerous hematologic and bone marrow parameters in the 5-FU treated groups were comparable to those in the corresponding pair-feeding groups in both age studies. Generally, the influences of undernutrition were more apparent in the young rats than in the adult rats. Histopathological examinations showed a decrease in hematopoiesis in the bone marrow in the 5-FU treated and pair-feeding groups. No apparent differences were observed in the decreased hematopoiesis between the 5-FU treated and pair-feeding groups in the 6-week-old study, but a difference between these groups was noted in the 12-week-old study; decreased hematopoiesis was more frequently noted in the 5-FU treated groups. These facts suggest that adult rats are more suitable than young rats for evaluation of 5-FU-induced myelotoxicity.

Visually evoked cortical potentials obtained using checker patterns can detect ethambutol-induced visual toxicity in albino rats

We determined whether visually evoked cortical potentials obtained using checker patterns (P-VECPs) and albino rats would reveal visual damage induced by ethambutol (EB). Findings were compared in cases of detection of visual damage between by P-VECPs and by flash visually evoked cortical potentials (F-VECPs). Twelve adult albino male Crj:CD(SD)IGS rats were grouped into four, three per group: control, 250PS, 500PS, and 500SC groups. In the 250PS and 500PS groups, rats were administered EB orally for the first 2 weeks and then subcutaneously for the second 2 weeks to 250 and 500 mg/kg, respectively. In the 500SC group, rats were given 500 mg/kg of EB subcutaneously for 4 weeks. Rats in the control group were given the vehicle orally for the first 2 weeks and then subcutaneouly for the second 2 weeks. P-VECPs and F-VECPs were carried out prior to initiation of drug administration and at the 1st, 2nd, 3rd, and 4th weeks of the administration. Prolongation of P1 latency in the P-VECPs was evident in both the 500PS and the 500SC groups at the 4th week, while no marked changes were observed in the F-VECPs. Thus, P-VECPs in albino rats can detect visual damage induced by EB even when F-VECPs cannot do so. These studies suggest that P-VECPs are useful for evaluating the visual toxicity of drugs.

Evaluation of Short-term Myelotoxicity Study in Dietary Reduced Rats

This study attempted to prove our hypothesis that a short-term toxicity study, using a 4-day dosing regimen as an example, is suitable for evaluating myelotoxicity in rats. We compared the hematological, bone marrow cytological and histopathological results of 5-fluorouracil (5-FU) treated and pair-feeding groups after a 4-day administration period. Several experimental groups were defined for this 4-day study as well as for our previously reported 14-day study (Miyata et al., 2009); these included 5-FU treated groups receiving 12, 15 and 18 mg/kg/day (FU12, FU15 and FU18), pair-feeding groups (R12, R15 and R18 receiving the same amount of food as the FU12, FU15 and FU18 groups, respectively) and a nontreated control group. Although severe reductions in body weight gain and food consumption were reported in the 14-day study, only slight reductions were observed in the 4-day study. In the 4-day study, a decrease in blood reticulocytes and a decreasing trend of marrow erythroid cells were only observed in the FU18 group, and no effects were observed in the pair-feeding groups. The erythroblastic changes observed in this 4-day study were thought to reflect the direct influence of 5-FU administration. Since concerns regarding the influence of secondary changes related to undernutrition were minimized in the 4-day study, it was thought to clarify the direct influence of 5-FU administration on erythroblastic cells. Thus, a 4-day study protocol might be helpful for distinguishing secondary changes related to undernutrition.