Immunohistochemical study on hypoxia in spontaneous polycystic liver and kidney disease in rats.

Hypoxia-inducible factor (HIF) mediates homeostatic responses to hypoxia and activates transcription of hypoxia-inducible genes including vascular endothelial growth factor (VEGF). The aim of this study was to examine the expressions of VEGF, HIF-1alpha and HIF-3alpha in spontaneously occurring hepatorenal polycystic lesions in two Sprague-Dawley (Crj:CD) rats. Hepatic multiple cysts were derived from the interlobular and large bile ducts, while renal cysts were from the collecting ducts and distal tubuli. These findings were confirmed by a lectin peanut agglutinin (PNA) histochemistry. In the polycystic liver, VEGF immunoreaction was strongly evident in the cytoplasm of hepatocytes, whereas expression of HIF-3alpha, but not HIF-1alpha, was found in a few nuclei of hepatocytes. In the polycystic kidney, VEGF immunoreaction was increased in the cytoplasm of collecting ducts and distal tubuli, whereas nuclear expression of HIF-1alpha and HIF-3alpha was evident in the proximal tubuli and thin loop of Henle, respectively. The results suggest that hypoxia-related molecules may be induced by cystic alterations in a heterogeneous appearance.

Transforming growth factor beta production by spontaneous malignant mesothelioma cell lines derived from Fisher 344 rats.

We investigated whether transforming growth factor beta (TGF-beta) is involved in the growth of malignant mesothelioma (MM) cells in culture. TGF-beta production was examined in two mesothelioma cell lines (MeET-4 and -6) that were established from rat spontaneous MM in our laboratory. TGF-beta bioactivity in conditioned medium of these cell lines was analyzed using a CCL64 mink lung epithelial cell growth inhibition assay and found to be 30-70 times higher than that of normal rat mesothelial cells (MCs). The MM cell lines also showed considerably higher levels of TGF-beta mRNA expression when compared with MCs. The bioactivity and mRNA expression level were greater in MeET-4 than MeET-6. When MeET-4 was treated with antisense TGF-beta1 oligonucleotide (ODN), a significant decrease in both anchorage-dependent and -independent growth was observed. Treatment with exogenous TGF-beta resulted in no effects on the growth pattern of the MM cell lines, while proliferation of the MCs was slightly induced. It is considered that TGF-beta appears to be produced by rat spontaneous MM cells through an autocrine mechanism and could modulate the malignant growth of the tumor cells.

Pulmonary microcystic hamartoma in an adult dog.

Microcystic hamartoma was detected as a tumorlike mass in the left caudal lung lobe of a 12-year-old mixed-breed dog. Histologically, the mass was characterized by microcysts of various sizes that mimicked alveoli and were surrounded by thin fibrous septal tissue. However, unlike the adjacent lung parenchyma, bronchial or bronchiolar trees were absent, and the septal vascular channels were extremely underdeveloped. Immunohistochemically, the cells lining the microcysts were consistently positive for cytokeratin but not for vimentin, whereas the septal fibroblast-like cells were negative for cytokeratin and positive for vimentin. Electron microscopy confirmed that the microcysts were lined with a layer of type I and type II mature pneumocytes. This is the first description of the detailed morphologic features of microcystic hamartoma.

Age-related hematological changes in normal F344 rats: during the neonatal period.

In order to clarify age-related changes in hematological values of normal rats after birth, blood samples from neonatal F344 rats of both sexes were examined periodically during the period from 0 to 40 days postpartum. The erythrocyte count (RBC) increased with time after birth as a function of age. In contrast, the reticulocyte count (Retics) continuously decreased with time after birth. Hemoglobin (Hb), hematocrit (Ht), and the mean corpuscular hemoglobin concentration (MCHC) tended to decrease after birth until weaning (about 21 days postpartum), but they began to increase after weaning. Mean corpuscular volume (MCV) and mean corpuscular hemoglobin (MCH) also gradually decreased after birth until weaning, but they were unchanged thereafter. The platelet count (PLT) gradually increased after birth and reached a plateau at weaning. Microscopic examination of blood smears revealed that erythrocytes at birth had characteristic morphological features such as anisocytosis, polychromasia, basophilic stippling, Howell-Jolly body, and erythroblastosis. These characteristic features, however, disappeared by 30 days after birth. The total leukocyte count (WBC) gradually increased with time after birth, due to an increase in the number of lymphocytes. The lymphocyte count started to rapidly increase within several days after birth and the increase continued thereafter. Other differential leukocyte counts also showed various characteristic patterns of changes during the neonatal period. There were no apparent differences between males and females in these changes in hematological values.

Oral teratogenicity studies of methyl bromide in rats and rabbits.

Teratogenicity studies of methyl bromide, a widely used fumigant, were conducted in rats and rabbits. Methyl bromide was dissolved in corn oil and administered orally to groups of 24 copulated female Crj:CD (SD) rats at dose levels of 0 (corn oil), 3, 10 or 30 mg/kg/day on days 6-15 of gestation and to groups of 18 artificially inseminated female Kbl:JW rabbits at 0, 1, 3 or 10 mg/kg/day on days 6-18 of gestation. Maternal rats and rabbits were euthanized on respective days 20 and 27 of gestation. Foetuses were examined for survival, growth and teratological alterations. Maternal toxicity was evident in the high-dose groups for both species. In these groups, maternal body weight gains and food consumption were significantly decreased during the dosing and post-dosing periods. Necropsy of maternal rats also revealed erosive lesions in the stomach and the surrounding organs. However, no treatment-related adverse effects were found in foetuses of the treated groups for both rat and rabbit studies. These results led to the conclusion that methyl bromide was not foetotoxic or teratogenic to rat and rabbit foetuses up to dose levels of 30 and 10 mg/kg/day, respectively, at which maternal toxicity was evident for both species.

Immunohistochemical localization of microcystin-LR in the liver of mice: a study on the pathogenesis of microcystin-LR-induced hepatotoxicity.

The relationship between the intralobular sites of hepatotoxic injury and the distribution of microcystin-LR (MCLR), an inhibitor of protein phosphatases 1 (PP1) and 2A (PP2A), was examined using an immunohistochemical method with a monoclonal antibody specific to MCLR on the livers of mice receiving a single i.p. injection of the MCLR. Immunoblotting and high-performance liquid chromatography analyses of liver extracts were also performed to determine the binding form of MCLR to PP1 and PP2A (MCLR-PP1/PP2A adducts) and free MCLR. Immunohistochemistry revealed a discernible intensity of staining in the centrilobular regions where hemorrhage and apoptosis occurred. In these regions, immunopositivity was evident in the cytoplasm and nuclei of the hepatocytes; some apoptotic cells were also immunopositive. In contrast, coagulative necrosis, which was mainly evident in the midlobular regions, was completely negative. Analysis of liver extracts demonstrated MCLR-PP1/PP2A adducts, but free MCLR was below detection limit. These results suggest that the immunohistochemical localization of MCLR in centrilobular hepatocytes is closely associated with the onset of hemorrhage and apoptosis and is related to adduct formation. The occurrence of coagulative necrosis however might also be related to other factors such as ischemia/hypoxia.

Morphological variations in transplanted tumors developed by inoculation of spontaneous mesothelioma cell lines derived from F344 rats.

Morphological and immunohistochemical features of the abdominal mesotheliomas that were developed by inoculation of 3 cell lines (MeET-4, -5 and -6) established from spontaneous abdominal mesotheliomas in male F344 rats. Although the original tumors of three cell lines showed signs of epithelioid growth with a predominantly simple papillary pattern, transplanted tumors revealed a variety of morphologic features including epithelioid with glandular structures, sarcomatous, and a mixture of these components. All tumor cells of transplanted tumors were positive for alpha-smooth muscle actin (ASMA) but almost negative for desmin as were epithelioid cells of the original tumors, and the cell lines were positive for desmin but not for ASMA. These results suggested that mesothelioma in the F344 rat had the potential for wide spectrum differentiation under in vitro conditions. The microenvironmental factors obtained in vivo can modify their potential ability and their morphological aspects. These factors may be related to tumor cell reexpression of ASMA of tumor cells that were masked under in vitro culture conditions.

Establishment and characterization of spontaneous mesothelioma cell lines derived from F344 rats.

Three mesothelioma cell lines (MeET-4, MeET-5, and MeET-6) established from ascitic fluid of F344 rats with spontaneous abdominal mesothelioma have been maintained through at least 60 passages on the DMEM with 10% FBS. Two of original tumours consisted of epithelioid cells growing in a papillary pattern, while one (original tumour of MeET-5) had sarcomatous areas composed of spindle-shaped tumour cells. The cell line originating from MeET-5 showed a constantly beiphasic growth pattern during the repetitive subcloning, while the other two lines retained a monophasic growth pattern. Although the growth pattern was different, the tumour cells in all three lines were positive for vimentin and keratin and ultrastructurally showed an abundant distribution of glycogen granules in the cytoplasm and numerous long microvilli on all surface. The modal chromosome number of cell lines varied from 41 to 71, and abnormal chromosomes were frequently seen. All cell lines established formed colonies on semi-solid medium and could be successfully transplanted, growing tumour masses in syngeneic rats and thus indicating their malignant nature. Cell lines grew even on a medium with a low concentration of FBS. The evidence suggests that they may produce growth factors that enable them to survive unfavourable medium conditions.

Intraabdominal lymphangiosarcoma in a Fischer-344 rat.

A Sarcoma arising in the abdominal cavity in an aged Fischer-344 rat was studied by immunohistochemistry and electron microscopy. The white-yellow soft mass was located on the lumbosacral vertebrae, compressing adjacent parenchymal organs. The tumor was made up of spindle shaped cells situated in a background of myxoid substance and a small amount of reticulin and collagen fibers. The tumor cells grew in a loose storiform pattern and often adhered to each other by their cell processes to form ovoid or slitlike spaces. Immunohistochemically, the tumor cells were strongly positive for vimentin but negative for keratin, macrophage ED1 antigen, alpha-smooth muscle actin, Factor VIII-related antigen, and S100 protein. Electron microscopy demonstrated the endothelial differentiation of the tumor cells, such as occasional luminal spaces, a small number of micropinocytotic vesicles, and interdigitating junctions with desmosomes between cell processes of adjacent cells. Furthermore, its endothelial origin was suggested by the presence of electron-dense rods resembling Weibel-Palade bodies. Instead of a definitive basement lamina surrounding the tumor cells, there were extracellular thin "anchoring filaments" that were attached to the cell surface at areas of increased electron density. These findings indicate that the tumor is of lymphatic vessel type rather than blood vessel type.

A safety study on rat's eye after 13-week oral administration with fenitrothion.

The potential of ocular toxicity of fenitrothion (O,O-dimethyl O-4-nitrom-tolyl phosphorothioate) was assessed in Sprague-Dawley (Crj:CD) rats of both sexes receiving a diet containing the test compound at concentrations of 0, 2.5, 5, 10, or 30 ppm for 13 weeks. The animals were observed daily for clinical signs and their body weights and food consumption were measured weekly during the study. At termination of treatment, surviving animals were subjected to ophthalmoscopy, electroretinography, and biochemical analyses of plasma, erythrocyte, and brain cholinesterase (ChE). Histopathological examinations of ocular tissues were performed on all animals by light microscopy and on two animals/sex/dose by electron microscopy. There were no treatment-related changes in clinical signs, body weights, and food consumption. A significant inhibition of ChE activity was observed in males (plasma and erythrocyte ChE) and females (plasma, erythrocyte, and brain ChE) at 30 ppm and in females (plasma ChE) at 10 ppm. Ophthalmological and histopathological examinations revealed neither functional nor morphological alterations in the visual system at any dose level. Under the conditions of the present study, there was no evidence of ocular toxicity of fenitrothion for male and female rats at dose levels up to 30 ppm (1.70 mg/kg/day for males and 1.96 mg/kg/day for females) where distinct inhibition of ChE activity was observed.

Morphological and ultrastructural study of the histogenesis of meningeal granular cell tumors in rats.

To clarify the cell of origin of granular cell tumors in the rat brain, light and/or electron microscopic examinations were performed on 40 cases of spontaneous meningeal tumors in Wistar (Jcl: Wistar) rats. The meningeal tumors were histologically subclassified into 3 types: meningothelial meningiomas (MMs), 3 cases; granular cell tumors (GCTs), 28 cases; and mixed forms (MIXs) of GCTs and MMs, 9 cases. Of these tumors, 2 MMs, 2 GCTs, and 3 MIXs were examined by transmission electron microscopy. Tumor cells of MMs were characterized by cytoplasmic intermediate filaments and prominent interdigitating cell processes often connected with cellular junctions without surrounding basal laminae. GCTs were composed of 2 cell types: granular cells with many dense bodies and filamentous cells with fine intermediate filaments. MIXs consisted of granular cells, filamentous cells, and intermediate cells. The tumor cells in GCTs and MIXs were apposed to each other and connected with cellular junctions. MIXs displayed a spectrum of cellular differentiation in that 2 MIXs had a close morphological resemblance to GCTs and the other one to MMs. These findings indicate there might be a histogenetic sequence among MMs, GCTs, and MIXs. The present study supports the suggestion that GCTs and MIXs may be variants of MMs from the meningeal arachnoid cell.

Comparison of the induction of hepatic peroxisome proliferation by the herbicide oxadiazon in vivo in rats, mice, and dogs and in vitro in rat and human hepatocytes.

Oxadiazon [5-ter-butyl-3-(2,4-dichloro-5-isopropoxyphenyl)- 1,3,4-oxadiazol-2(3H)-one] was administered orally at 20-500 mg/ kg body wt/day to male Sprague-Dawley CD rats for 14 days, at 20-200 mg/kg body wt/day to male CD1 [CR1/CD-1(1GR)BR] mice for 28 days, and at 500 mg/kg body wt/day to male beagle dogs for 28 days. Although liver enlargement was observed in the three species, morphological studies indicated that peroxisome proliferation only occurred in rats and mice. Parallel biochemical investigations showed that there was a dose-dependent increase in the peroxisomal cyanide-insensitive palmitoyl CoA oxidase and acetyl carnitine transferase activities in treated rats and mice. Acetyl carnitine activity appeared to correlate well with the number and volume of peroxisomes as determined histologically. The increases in enzyme activities at 200 mg/kg body wt/day oxadiazon were comparable in rats and mice indicating that both rodent species were equally sensitive to oxadiazon-induced peroxisome proliferation. When added in vitro to cultured rat hepatocytes at concentrations ranging from 2.5 to 10 x 10(-5) M, oxadiazon induced a dose-dependent increase in the activities of palmitoyl CoA oxidase and acetyl carnitine transferase. The ratio obtained by comparing oxadiazon and clofibric acid on acetyl carnitine transferase activity at 5 x 10(-5) M in the present in vitro study on rat hepatocytes are equivalent to those that can be calculated from the results on this enzyme activity obtained in vivo in the rat with 500 mg/ kg body wt/day oxadiazon (this study) and clofibric acid (literature values), indicating that the rat hepatocyte cultures gave satisfactory results regarding peroxisome proliferation. Neither oxadiazon nor clofibric acid modified the activities of palmitoyl CoA oxidase and acetyl carnitine transferase in cultured human hepatocytes. The results presented here demonstrate clearly that oxadiazon induces peroxisome proliferation in rodents in vivo and in vitro, as determined both biochemically and morphologically, whereas dogs in vivo and human hepatocytes in vitro were refractory to peroxisome proliferation. This observation extends to the herbicide oxadiazon, which is structurally unrelated to other known peroxisome proliferators, the generally observed marked species difference in sensitivity to peroxisome proliferation, and has important implications in the human safety evaluation of this herbicide.

Effects of age on endometrial carcinogenesis induced by concurrent oral administration of ethylenethiourea and sodium nitrite in mice.

Aging effects on the susceptibility to chemical endometrial carcinogenesis were investigated in ICR female mice. The animals were divided into 3 groups of different ages: 1 month (young), 6 months (middle), and 12 months (old) at initiation of treatment. They received weekly oral administration of mixture of ETU (100 mg/kg body weight) and sodium nitrite (70 mg/kg body weight) for 6 months followed by a withdrawal period of 3 months. All animals were subjected to histopathology. The incidence of endometrial adenocarcinomas was highest in the middle age group (8/20), secondary in the old age group (4/20), and lowest in the young group (1/20). The incidence of atypical glandular hyperplasia, a precursor lesion of the tumor, was also higher in the middle age group. The endometrial adenocarcinomas showed morphological similarities among all age groups and the nuclei of tumor cells lost almost all staining reactivity to estrogen receptors. The labeling indices with bromodeoxyuridine (BrdU) were notably higher in the old age group than in the young and middle age groups. A further investigation on the aging process of female genital organs in control mice revealed that their senility seemed to be preceded by the formation of ovarian cysts which first appeared at 6 months of age with a concomitant elevation of plasma 17 beta-estradiol level. These results indicate that the susceptibility of the mouse endometrium to the carcinogenic effects of N-nitroso ETU could be closely linked with the stage of aging process of the genital organs and it appears to be most susceptible when initiated at around 6 months of age. However, the mitotic activity of neoplastic endometrial glandular cells seems to be higher in older mice than younger ones.

An epithelioid cell type of amelanotic melanoma of the pinna in a Fischer-344 rat: a case report.

An epithelioid cell type of spontaneous amelanotic melanoma in the pinna of an aged albino Fischer-344 (F-344) rat is described. When the rat was necropsied at 109 wk of age, the pinnal tumor was recognized as a white spherical mass of 4 mm in diameter. Histologically, the tumor was observed in the dermis and composed of large round or polyhedral epithelioid eosinophilic cells that were arranged in various-sized cell nests. Immunohistochemistry demonstrated that the tumor cells were positive for S-100 protein and vimentin but negative for keratin and a rat macrophage antigen. Ultrastructurally, a few single membrane-bound premelanosomes that were accompanied by many primary lysosome-like bodies containing very fine granules were present in the cytoplasm of the tumor cells. This study has disclosed that, in addition to spindle cell variants of amelanotic melanomas, an epithelioid cell type of amelanotic melanoma may occur spontaneously in the skin of F-344 rats.

Endometrial carcinogenesis induced by concurrent oral administration of ethylenethiourea and sodium nitrite in mice.

Endometrial carcinogenesis induced by concurrent oral administration of ethylenethiourea (ETU) and sodium nitrite (NaNO2) was investigated in ICR (Crj:CD-1) female mice. A mixed solution of ETU (100 mg/kg) and NaNO2 (70 mg/kg) was given to animals orally once a week for up to 6 months and all surviving animals were killed at 12 months of study. During the study, estrous cycle was monitored by vaginal smear and five or 10 selected animals were subjected to interim killing at 3 month interval to observe time-related carcinogenic responses of the uterus. Treatment with ETU and NaNO2 resulted in development of endometrial adenocarcinomas in the uterine horn and the incidence reached 42% in the surviving animals at 12 months. Prior to the development of the tumor, atypical hyperplasia of endometrial glands was frequently observed and regarded as the precancerous lesion. Immunohistochemistry for bromodeoxyuridine (BrdU) incorporation revealed higher labeling indices in both hyperplastic and neoplastic endometrial glandular cells, and the index in the adenocarcinoma was more than 20% on average at any stage of the estrous cycle. Overexpression of p53 protein, which is frequently demonstrated in virulent phenotypes of human corpus cancers, was seen in three out of eight (38%) adenocarcinomas, but not in the atypical hyperplasia or normal endometrial glands. There were no treatment-related changes in the estrous cycle on vaginal smears at any interval of the study. The analyses for plasma ovarian hormones at 12 months disclosed a marked depression of progesterone in the treated animals, while the 17 beta-estradiol (E2) level was comparable to the controls. These results suggest that endometrial carcinogenesis by ETU and NaNO2 could be initiated with atypical hyperplasia of the endometrial gland and a decrease in plasma progesterone level may play an important role in the development of endometrial carcinogenesis. In addition, inactivation of the p53 gene may play a significant role in the malignant transformation of endometrial epithelial cells in mice.

Different inhalation lethality between micron-sized and submicron-sized aerosols of organophosphorus insecticide, chlorfenvinphos, in rats.

This study was designed to evaluate that a difference in the route of absorption or the mode of lethality is responsible for the higher inhalation lethality of micron-sized organophosphorus insecticide, chlorfenvinphos (CVP), aerosols than the submicron-sized aerosols. Male Fischer 344 rats were exposed to the micron-sized (> 1 micron) or the submicron-sized aerosols (< 1 micron) for 4 h using the nose-only exposure system. LC50 of the micron-sized and the submicron-sized aerosols was 0.13 mg/l and 0.51 mg/l, respectively. Placing a drain cannula in the esophagus markedly increased LC50 of the micron-sized aerosols to 0.49 mg/l, but not that of the submicron-sized aerosols. There was no qualitative difference in lethal profile in cardiorespiration between 2 types of aerosols. The higher lethality of the micron-sized aerosols could be ascribed to swallowed CVP.

Carcinogenicity studies of oxolinic acid in rats and mice.

A chronic feeding study was conducted to determine the carcinogenic potential of oxolinic acid, an antimicrobial agent, in rats and mice. Oxolinic acid was administered in the diet to Wistar rats (0, 30, 100, 300 or 1000 ppm; 50 rats/dose/sex) for 104 wk and to ICR mice (0, 50, 150 or 500 ppm; 50 mice/dose/sex) for 78 wk. Clinical signs, body weight, food consumption, autopsy findings and histopathological data were noted. Mortality was unaffected by oxolinic acid administration in neither species. In rats, body weight gain was suppressed in both sexes at 1000 ppm. Histopathological examinations conducted after autopsy at 104 wk revealed a slight increase in benign Leydig cell tumours of the testis at 1000 ppm, which did not appear until late in the lifetime of rats. No other treatment-related neoplastic lesions were observed in rats. Non-neoplastic lesions in males at 1000 ppm included Leydig cell hyperplasia and tubular atrophy of the testes. In mice, decreased body weight gain was observed in both sexes at 500 ppm, but no non-neoplastic or neoplastic lesions attributable to the treatment with oxolinic acid occurred in either sex. In conclusion, oxolinic acid induced benign Leydig cell tumours of the testis in rats at the highest dose level tested (1000 ppm). The no-effect level for tumour induction was confirmed to be 300 ppm (10.9 mg/kg/day) in rats. None was induced in mice.

A two-generation reproduction study in rats with methyl bromide-fumigated diets.

Crj:CD(SD) rats were fed basal (control) diet or methyl bromide-fumigated diet containing 80, 200 or 500 ppm total bromine for two successive generations (18 wk for each generation). Compared with controls, food consumption was significantly lower in F1 parental females of the 500-ppm group during the second half of the dosing period, and F2 female pups of the 500-ppm group showed lower body weights throughout the lactation period. No other treatment-related changes were found at any dietary level of total bromine in either parental animals (F0, F1) or their offspring (F1, F2) for any of the parameters studied (i.e. clinical signs, oestrous cycle, sperm count and morphology, mating, fertility, gestation, parturition, litter size, pup viability, organ weights, and gross or histopathological examination). It was concluded that residues of 200 and 500 ppm total bromine in diets fumigated with methyl bromide are the no-observed-effect level and the minimum toxic level, respectively, for both parental rats and their offspring, and that bromine residues do not affect reproductive performance at dietary levels of up to 500 ppm.

Tumor induction by concurrent oral administration of ethylenethiourea and sodium nitrite in mice.

Carcinogenic potential of ethylenethiourea (ETU) in combination with sodium nitrite was investigated in ICR mice of both sexes. Groups of 30 males and 30 females each were given 10 weekly oral administrations of ETU and sodium nitrite with the following combinations of dosing (ETU vs sodium nitrite, mg/kg/wk): 0 vs 0, 100 vs 0, 0 vs 70, 25 vs 17.5, 50 vs 35, and 100 vs 70. Thereafter, the animals were allowed to live without treatment up to 18 mo after the first administration. Concurrent administration of ETU and sodium nitrite caused earlier development of tumors and/or dose-dependent increases in the incidences of tumors in the lymphatic tissue, lung, forestomach, Harderian gland, and uterus, whereas treatment with either ETU or sodium nitrite failed to show carcinogenic activity. In addition, carcinomas in the forestomach and uterine horn were limited to mice receiving concurrent administrations of ETU and sodium nitrite. These results indicate that ETU is most probably converted in vivo into N-nitroso ETU and that the N-nitroso ETU has a greater carcinogenic potential in mice than ETU alone.

Quantitative analysis of neuronal damage induced by tri-ortho-cresyl phosphate in Wistar rats.

A quantitative analysis of neuronal damage was performed on the fasciculus gracilis (FG) of the cervical spinal cord in male Wistar rats that received orally a single dose of tri-ortho-cresyl phosphate (TOCP) at 1500 mg/kg. FG tissues were sampled at 1, 2, and 3 weeks after treatment and examined histopathologically. Wallerian degeneration of myelinated nerve fibers was observed in FG at 2 weeks. Morphological changes were most evident at 3 weeks after treatment and the number of fibers was reduced. Ultrastructurally, axonal swelling due to the accumulation of cytoplasmic contents was observed near the node of Ranvier in the affected animals, indicating paranodal degeneration. Axonal atrophy and swelling in organophosphorus-induced delayed neuropathy (OPIDN) were evaluated quantitatively using a computer-assisted image analyzer. Morphometric examinations on semi-thin sections and frozen sections stained with Nauta's method were demonstrated to be useful for objective evaluation of OPIDN in the rat.