Toru R. Saito

Fine structure of three types of olfactory organs in Xenopus laevis

There is no report on the fine structure of three types of olfactory organs in Xenopus laevis. Their functional assignments in olfaction are not yet established. The fine structure of three types of olfactory organs, olfactory epithelium (OE), vomeronasal organ (VNO), and middle chamber epithelium (MCE), was examined in Xenopus laevis by light and electron microscopy. The olfactory cells of the OE and the sensory cells of the VNO were equipped with cilia and microvilli, respectively, similar to terrestrial animals that possess both the OE and the VNO. On the other hand, the sensory cells of the MCE were classified into two types, the sensory cells with cilia and the sensory cells with microvilli, like those of the OE in fish. These findings suggest that the OE and the VNO in Xenopus laevis detect different kinds of odoriferous molecules in air, whereas the MCE is involved in the perception of odorants in water. Anat. Rec. 252:301–310, 1998.

Carcinogenicity of methylmercury chloride in ICR mice: preliminary note on renal carcinogenesis.

ICR mice (60/group) were fed diets containing methylmercury chloride (MMC) in 0, 15 or 30 ppm for 78 weeks. The majority of mice in the 30-ppm group died due to the neurotoxicity by week 26. The first renal mass was grossly seen in a male of the 15-ppm group at week 58. Histopathology on kidney tissue from all animals surviving after 53 weeks revealed renal tumors in 13 of 16 males of the 15-ppm group in contrast with 1 of 37 males of the control. No renal tumors were seen in the female treated groups and/or control groups.

Obesity induced changes to plasma adiponectin concentration and cholesterol lipoprotein composition profile in cats.

Feline obesity generally results in aberrations to plasma metabolite levels, such as lipid concentrations and lipoprotein composition. This study sought to investigate the resultant effect of obesity on cholesterol lipoprotein composition and circulating adiponectin concentrations in cats. Plasma glucose, lipids (triglyceride, cholesterol and free fatty acid), insulin and adiponectin concentrations, and cholesterol lipoprotein composition were measured and compared between body condition score (BCS) determined normal healthy control and obese cats. Although the obese group demonstrated higher levels of plasma cholesterol, glucose, and triglycerides, as compared to healthy controls, the difference was insignificant thus indicating that the BCS determined obese cats may have been overweight and not morbidly obese. Plasma insulin levels were significantly higher (25-30%) versus healthy control animals thereby possibly hinting at the ensuing emergence of obesity induced insulin resistance. However, the BCS determined obese cat demonstrated a significant reduction (p<0.05) in plasma adiponectin concentration and a significant increase (p<0.05) in LDL-cholesterol % as compared to age matched healthy control animals. This would indicate that changes in plasma adiponectin concentration and cholesterol lipoprotein composition may be good early indicators of obesity in cats.

Effects of estrogen on food intake, serum leptin levels and leptin mRNA expression in adipose tissue of female rats

The integration of metabolism and reproduction involves complex interactions of hypothalamic neuropeptides with metabolic hormones, fuels, and sex steroids. Of these, estrogen influences food intake, body weight, and the accumulation and distribution of adipose tissue. In this study, the effects of estrogen on food intake, serum leptin levels, and leptin mRNA expression were evaluated in ovariectomized rats. Seven-week-old female Wistar-Imamichi rats were ovariectomized and divided into three treatment groups: group 1 (the control group) received sesame oil, group 2 was given 17β-estradiol benzoate, and group 3 received 17β-estradiol benzoate plus progesterone. The body weight and food consumption of each rat were determined daily. Serum leptin levels and leptin mRNA expression were measured by ELISA and quantitative RT-PCR, respectively. Food consumption in the control group was significantly higher (P<0.05) than that in groups 2 and 3, although body weight did not significantly differ among the three groups. The serum leptin concentration and leptin mRNA expression were significantly higher (P<0.05) in groups 2 and 3 than in group 1, but no significant difference existed between groups 2 and 3. In conclusion, estrogen influenced food intake via the modulation of leptin signaling pathway in ovariectomized rats.

Characterization of multiple first exons in murine prolactin receptor gene and the effect of prolactin on their expression in the choroid plexus

Prolactin (Prl) receptor (Prlr) gene is expressed in various brain regions, with the highest level present in the choroid plexus, a site for receptor-mediated PRL transport from the blood to cerebrospinal fluid. We investigated the regulatory mechanism of Prlr gene expression by PRL in the murine choroid plexus. We first examined the organization of the alternative first exons in murine Prlr gene. In addition to the three known first exons, mE1(1), mE1(2), and mE1(3), two first exons, mE1(4) and mE1(5), were newly identified by cDNA cloning. Each first exon variant of Prlr mRNA exhibited tissue-specific or generic expression. In the choroid plexus of mice, the expression levels of mE1(3)-, mE1(4)-, and mE1(5)-Prlr mRNAs were increased in the lactating mice compared with those in the diestrus mice. Furthermore, the expression level of mE1(4)-Prlr mRNA was decreased in the PRL-deficient (Prl(-/-)) mice compared with the PRL-normal (Prl(+/+) and Prl(+/-)) mice. In the ovariectomized Prl(-/-) mice, the expression level of mE1(4)-Prlr mRNA was significantly increased by PRL administration but not by 17β-estradiol administration. The expression levels of the two last exon variants of Prlr mRNAs, encoding the long and short cytoplasmic regions of PRLR, were also increased in the lactating mice and decreased in the Prl(-/-) mice. These findings suggest that PRL stimulates the Prlr gene expression through the transcriptional activation of mE1(4) first exon, leading to increases in the long- and short-form variants of Prlr mRNA in the murine choroid plexus.

Immunohistochemical analysis of cAMP response element-binding protein in mouse testis during postnatal development and spermatogenesis.

Basal activity and cellular localization of cAMP response element-binding protein (CREB) was examined in mouse testis during postnatal development and spermatogenesis. Testes of ICR mice sampled on postnatal day (PND) 3, 7, 14, 21, 28, 35, 42, and 49 were analyzed using Western blotting. Basal CREB activity was significantly higher in early phase (PND 3–7) developing testes than in intermediate- and late-phase developing (PND 14–42) and adult testes (PND 49). Furthermore, immunohistochemical analysis demonstrated the change of CREB phosphorylation in various testicular cell types during postnatal development. In particular, CREB phosphorylation in seminiferous tubules of the adult testis varied according to the spermatogenic cycle, while phosphorylation was evident in spermatogonia during all stages. Phosphorylation was moderate in pachytene spermatocytes of stages I–III and intense in round and elongate spermatids of spermiogenesis in stages XII–IX. These results suggest that CREB plays an important role in cell proliferation and differentiation in the early phase of postnatal development and spermatogenesis of mouse testis.

Use of long term dermal sensitization followed by intratracheal challenge method to identify low-dose chemical-induced respiratory allergic responses in mice

The inhalation of many types of chemicals, including pesticides, perfumes, and other low-molecular weight chemicals, is a leading cause of allergic respiratory diseases. We attempted to develop a new test protocol to detect environmental chemical-related respiratory hypersensitivity at low and weakly immunogenic doses. We used long-term dermal sensitization followed by a low-dose intratracheal challenge to evaluate sensitization by the well-known respiratory sensitizers trimellitic anhydride (TMA) and toluene diisocyanate (TDI) and the contact sensitizer 2,4-dinitrochlorobenzene (DNCB). After topically sensitizing BALB/c mice (9 times in 3 weeks) and challenging them intratracheally with TMA, TDI, or DNCB, we assayed differential cell counts and chemokine levels in bronchoalveolar lavage fluid (BALF); lymphocyte counts, surface antigen expression of B cells, and local cytokine production in lung-associated lymph nodes (LNs); and antigen-specific IgE levels in serum and BALF. TMA induced marked increases in antigen-specific IgE levels in both serum and BALF, proliferation of eosinophils and chemokines (MCP-1, eotaxin, and MIP-1beta) in BALF, and proliferation of Th2 cytokines (interleukin (IL)-4, IL-10, and IL-13) in restimulated LN cells. TDI induced marked increases in levels of cytokines (IL-4, IL-10, IL-13, and IFN-gamma) produced by restimulated LN cells. In contrast, DNCB treatment yielded, at most, small, nonsignificant increases in all parameters. Our protocol thus detected respiratory allergic responses to low-molecular weight chemicals and may be useful for detecting environmental chemical-related respiratory allergy.

Detection of low-level environmental chemical allergy by a long-term sensitization method

Multiple chemical sensitivity (MCS) is characterized by various signs, including neurological disorders and allergy. Exposure may occur through a major event, such as a chemical spill, or from long-term contact with chemicals at low levels. We are interested in the allergenicity of MCS and the detection of low-level chemical-related hypersensitivity. We used long-term sensitization followed by low-dose challenge to evaluate sensitization by well-known Th2 type sensitizers (trimellitic anhydride (TMA) and toluene diisocyanate (TDI)) and a Th1 type sensitizer (2,4-dinitrochlorobenzene (DNCB)). After topically sensitizing BALB/c mice (9 times in 3 weeks) and challenging them with TMA, TDI or DNCB, we assayed their auricular lymph nodes (LNs) for number of lymphocytes, surface antigen expression of B cells, and local cytokine production, and measured antigen-specific serum IgE levels. TMA and TDI induced marked increases in levels of antigen-specific serum IgE and of Th2 cytokines (IL-4, IL-5, IL-10, and IL-13) produced by ex vivo restimulated lymph node cells. DNCB induced a marked increase in Th1 cytokine (IL-2, IFN-gamma, and TNF-alpha) levels, but antigen-specific serum IgE levels were not elevated. All chemicals induced significant increases in number of lymphocytes and surface antigen expression of B cells. Our mouse model enabled the identification and characterization of chemical-related allergic reactions at low levels. This long-term sensitization method would be useful for detecting environmental chemical-related hypersensitivity.

A simple method for prevention of coprophagy in the mouse

A series of studies of coprophagy in mice has been conducted from the viewpoints of behaviour and nutrition. Since it was necessary to prevent coprophagy to investigate its significance, an attempt was made to devise a re-strainer. A polyethylene bottle was cut and fixed to the mouse so as to prevent its head from reaching its anus when faeces were excreted. The animals were also housed in cages with floors of larger mesh size in order to let the faeces fall through.

Quantitative and qualitative analysis of rat pup ultrasonic vocalization sounds induced by a hypothermic stimulus

Ultrasonic vocalizations (USVs) are essential communicative sounds used between rodent pups and their mother. Rat pups emit USVs in stressful situations, such as when they are cold or separated from the nest. We verified the ontogenetic changes in USVs emitted by infant rats isolated from their mother during the pre-weaning period. The number of calls, and the median frequency and first peak of frequency of the calls were measured at 1, 3, 5, 7, 10, 12, and 14 days postnatal age in Wistar-Imamichi rats. Pups were placed in a cold glass beaker and USVs were recorded for 5 min. The number of calls increased to a peak on day 5 and then gradually decreased. The median frequency of calls decreased slowly during the first 12 days, and then increased slightly on day 14. Similarly, the first peak frequency of calls was the highest on day 1, and then decreased gradually by day 12. A small increase was observed on day 14. These changes in frequency were correlated with the physical development of the pups, whose body weights increased significantly with age except during postnatal days 7-10.