Quantitative structure-activity studies of benzoylphenylurea larvicides: I. Effect of substituents at aniline moiety against Chilo suppressalis walker

The larvicidal activity of a series of N-2,6-difluoro- and N-2,6-dichlorobenzoyl-N′-(4-substituted phenyl)ureas against nondiapause larvae of the rice stem borer, Chilo suppressalis Walker, was measured by topical application and oral administration methods under conditions with and without piperonyl butoxide as an inhibitor of oxidative metabolism. The effects of substituents at the anilide moiety on the larvicidal activity were analyzed quantitatively using physicochemical substituent parameters and regression analysis. The results indicate that the oxidative metabolism in the larval body which is favored by electron-donating substituents is significant in determining the activity. The activity, when the metabolic factor is eliminated, is enhanced by electron-with-drawing and hydrophobic substituents and lowered by bulky groups. The inhibitory activity against new cuticle formation of the same series of compounds was also measured using cultured integument of the rice stem borer diapause larva. The comparison of the quantitative analyses between larvicidal and integument-level activities shows that inhibition of cuticular development is the most important factor governing larvicidal activity.     

Inhibitory effects of pesticides on growth and respiration of cultured cells

Effects of various drugs including pesticides on the growth and respiration of cultured cells were evaluated comparatively using cell lines derived from mosquito ovary and subcutaneous mouse tissues. The concentration producing 50% inhibition of cell growth, I₅₀ (M), was determined for each of 42 drugs. Inhibitors of respiration and nucleic acid and protein biosyntheses such as rotenone, piericidin A₁, actinomycin D, and puromycin had very high pI₅₀ values of approximately 8. Except for the compounds known to be uncouplers of oxidative phosphorylation, the drugs suppressed the respiration rate of the cultured cells to various degrees. The pI₅₀ value (and the pEC₁₅₀, 150% enhancement of the control, value for uncouplers) was determined for each compound. By examining the relation of pI₅₀ (and pEC₁₅₀) values between cell growth and respiration, the compounds could be classified into two groups according to their modes of inhibitory action against the cultured cells. One mode relates to the inhibition of energy synthesis and the other, perhaps, to interference with the biosynthesis of biomacromolecules.     

Nerve membrane as a target of pyrethroids

Nerve membrane is one of the most important target sites of pyrethroids. Changes in excitability of nerve fibres caused by allethrin can be satisfactorily accounted for in terms of ionic permeabilities of the membrane. Temperature has a profound effect on the allethrin-induced permeability changes.     

Immunolocalization of steroidogenic enzymes in the corpus luteum and placenta of the Japanese Shiba goat

In this study, we performed immunohistochemistry of cholesterol side-chain cleavage cytochrome P450 (P450scc), 3beta-hydroxysteroid dehydrogenase (3betaHSD), cytochrome 17alpha-hydroxylase P450 (P450c17), and cytochrome P450 aromatase (P450arom) in the corpus luteum and placenta of Shiba goats. The aim was to clarify the steroidogenic capability of the corpus luteum and placenta of Shiba goats. Ovaries containing corpora lutea were obtained from four adult Shiba goats during the luteal phase (day10; n=2) and pregnancy (90 and 120 days of gestation). Placenta was obtained from one Shiba goat on day 120 of gestation. The sections of the ovaries and placentae were immunostained using the avidin-biotin-peroxidase complex method (ABC) with polyclonal antibodies generated against steroidogenic enzymes of mammalian origin. All luteal cells expressed P450scc, 3betaHSD, P450c17 and P450arom. The distribution of P450scc, 3betaHSD, P450c17 and P450arom were not different during the luteal phase and pregnancy. P450arom showed a weak positive staining in late pregnancy (120 days). In addition, immunoreactions for P450c17 and P450arom were observed in syncytiotrophoblast of the placenta of one Shiba goat. These results indicate that, in Shiba goats, corpus luteum is not only an important source of progesterone but also has the ability to synthesize androgen and estrogen during the luteal phase and pregnancy. Also the placenta has the ability to synthesize androgen and estrogen in late pregnancy.     

Insulin-like growth factor-I stimulated DNA replication in mouse endometrial stromal cells

Much evidence has suggested that sex steroid hormone-induced growth of uterine cells is mediated by polypeptide growth factors synthesized in uterine tissues. The present study aimed to clarify the effect of insulin-like growth factor-I (IGF-I) on the proliferation of mouse endometrial stromal cells obtained from immature mice. IGF-I and IGF-I receptor (type I) mRNAs were detected in the endometrial stromal cells. IGF-I increased bromodeoxyuridine (BrdU) uptake in the endometrial stromal cells, indicating an increase in DNA replication. E2 increased IGF-I mRNA levels in the endometrial stromal cells. IGF-I receptor is a tyrosine kinase receptor, and treatment with genistein, a tyrosine kinase inhibitor, reduced IGF-I-induced BrdU-uptake in the endometrial stromal cells. IGF-I signaling pathways involve mitogen-activated protein (MAP) kinase and phosphatidylinositol-3 kinase (PI-3 kinase). Treatment with 10(-7) M of the MAP kinase inhibitor PD098059 and 10(-5) M of the PI-3 kinase inhibitor LY294002 decreased IGF-I-induced BrdU-uptake in the endometrial stromal cells. However, LY294002 (10(-5) M) also decreased the BrdU-uptake in the absence of IGF-I treatment. These results suggest that endometrial IGF-I is involved in the proliferation of endometrial stromal cells in a paracrine or autocrine manner, and that the MAP kinase pathway is involved in DNA replication of endometrial stromal cells.     

Male Pseudohermaphroditism in a raccoon dog (Nyctereutes procynoides)

A wild raccoon dog (Nyctereutes procynoides) which died due to a traffic accident on 18 October 2001, and was determined to be 4.5 years old, was examined. Visual appearance of the external genitalia in this animal showed to be female with a large penis-like clitoris protruding from the vulvar juncture. Visual examination of the internal genitalia revealed that the animal possessed both testes and uterus. Histological appearance of the removed gonads showed only Sertoli cells but no spermatogenesis. Using polymerase chain reaction with skin biopsy directed against the sex-determining region on the Y chromosome (SRY) gene, the genomic SRY gene was expressed as a single band and sequenced. Based on these findings, this raccoon dog was diagnosed as male pseudohermaphrodite.     

Carcinogenicity and chronic toxicity in mice and rats exposed by inhalation to para-dichlorobenzene for two years

Carcinogenicity and chronic toxicity of para-dichlorobenzene (p-DCB) were examined by exposing 50 BDF1 mice and 50 F344 rats of both sexes by inhalation to p-DCB vapor at a target concentration of 0 (control), 20, 75 or 300 ppm for 6 hr/day, 5 days/week and 2 years. Incidences of hepatocellular carcinomas, hepatoblastomas and hepatic histiocytic sarcomas in the 300 ppm-exposed male mice, and hepatocellular adenomas and carcinomas and hepatoblastomas in the 300 ppm-exposed female mice were increased. An increase in the incidences of most of those liver tumors was dose-related. No increase in tumor incidence was found in any p-DCB-exposed rat of either sex. Centrilobular hypertrophy of hepatocytes and papillary mineralization and pelvic urothelial hyperplasia of the kidney were noted in the 300 ppm-exposed male rats. Treatment- and age-related increases in incidences of the eosinophilic globules of the respiratory and olfactory epithelia in female rats and incidences of the respiratory metaplasia of the nasal gland epithelium in mice and rats and the olfactory epithelium in mice were noted. The nasal lesion was the most sensitive endpoint of chronic inhalation toxicity. Induction of the mouse hepatocarcinogenicity and lack of the rat nephrocarcinogenicity found in the present study were compared with the mouse liver tumors and the rat renal tumors reported by the NTP gavage study, and discussed in light of the estimated p-DCB uptake into the body through the inhalation and the oral administration.     

Participation of Rhipicephalus sanguineus (Acari: Ixodidae) in the epidemiology of canine visceral leishmaniasis

The vectorial competence of the tick Rhipicephalus sanguineus is discussed in relation to the epidemiology of canine visceral leishmaniasis, taking into account its strict association with dogs and the low indices of natural infection presented by its known vector, the phlebotomine sand fly Lutzomyia longipalpis. In order to evaluate natural infection by Leishmania chagasi and the infectivity of these parasites in the tick, 39 specimens (6 females, 11 males and 22 nymphs) of R. sanguineus were removed from 21 dogs showing diverse symptoms of zoonotic visceral leishmaniasis (ZVL). Six ticks (15.4%) gave positive results for the genus Leishmania using the PCR technique. To determine the infectivity of the parasites, 36 hamsters were inoculated orally and peritoneally with macerates of ticks removed from nine dogs symptomatic for visceral leishmaniasis. After 6 months the hamsters were sacrificed and necropsied. Serum was removed for IFAT, as well as spleen and liver fragments to make imprint smears and for PCR. Eight (88.9%) of these dogs presented ticks that were infective for 14 hamsters (41.2%), 12 (85.7%) of them infected peritoneally and two (14.3%) orally. PCR revealed 27 smears (40.9%) to be positive, 20 (62.5%) of them infected peritoneally and seven (20.6%) orally. IFAT showed 14 positive animals (41.2%). Based on these findings, we suggest that the vectorial capacity of R. sanguineus for L. chagasi should be evaluated further, opening new perspectives in the epidemiology of ZVL.     

Fipronil modulation of GABAA receptor single-channel currents

Fipronil is the first phenylpyrazole insecticide introduced for pest control. Although fipronil is known to inhibit GABA receptors, the detailed mechanism of action remains to be seen. In order to elucidate the mechanism of fipronil interaction with the mammalian GABAA system, single-channel patch clamp experiments were performed using rat dorsal root ganglion neurons. The amplitude of main conductance state (27pS) current was not significantly altered by co-application of 10 microM fipronil and 10 microM GABA. The histograms of open time distribution were fitted to a sum of three exponential functions. After application of 10 microM fipronil, the proportion of the fastest component increased slightly and that of the slowest component decreased slightly. Thus, the mean open time was decreased from 11.4 ms to 7.8 ms by fipronil. The histograms of closed time distribution were fitted to a sum of four exponential functions. Fipronil 10 microM prolonged the slowest time constant resulting in a prolongation of the mean closed time from 29.7 ms to 52.8 ms. Thus, the frequency of channel openings was reduced. Thus, the fipronil suppression of GABA-induced whole-cell currents is caused in part by decreases in the channel open time and the frequency of channel openings.     

Immunohistochemical localization of steroidogenic enzymes in corpus luteum of wild sika deer during early mating season

We analyzed the localization of steroidogenic enzymes (P450 scc, 3 beta HSD, P450 arom and P450 c17) in the corpora lutea of two Hokkaido sika deer (Cervus nippon yesoensis) during the early mating season. Two corpora lutea were found in each female and the timing of formation of the corpora lutea seemed different. P450 scc, and 3 beta HSD, positive luteal cells were found in both corpora lutea. The existence of two functional corpora lutea from the early mating season through pregnancy suggests that progesterone secreted by two or more corpora lutea is necessary for maintenance of pregnancy in sika deer.