Spatial Analysis of Slowly Oscillating Electric Activity in the Gut of Mice Using Low Impedance Arrayed Microelectrodes

Smooth and elaborate gut motility is based on cellular cooperation, including smooth muscle, enteric neurons and special interstitial cells acting as pacemaker cells. Therefore, spatial characterization of electric activity in tissues containing these electric excitable cells is required for a precise understanding of gut motility. Furthermore, tools to evaluate spatial electric activity in a small area would be useful for the investigation of model animals. We thus employed a microelectrode array (MEA) system to simultaneously measure a set of 8×8 field potentials in a square area of ∼1 mm². The size of each recording electrode was 50×50 µm², however the surface area was increased by fixing platinum black particles. The impedance of microelectrode was sufficiently low to apply a high-pass filter of 0.1 Hz. Mapping of spectral power, and auto-correlation and cross-correlation parameters characterized the spatial properties of spontaneous electric activity in the ileum of wild-type (WT) and W/W^v mice, the latter serving as a model of impaired network of pacemaking interstitial cells. Namely, electric activities measured varied in both size and cooperativity in W/W^v mice, despite the small area. In the ileum of WT mice, procedures suppressing the excitability of smooth muscle and neurons altered the propagation of spontaneous electric activity, but had little change in the period of oscillations. In conclusion, MEA with low impedance electrodes enables to measure slowly oscillating electric activity, and is useful to evaluate both histological and functional changes in the spatio-temporal property of gut electric activity.     

A 5-bp Insertion in Mip Causes Recessive Congenital Cataract in KFRS4/Kyo Rats

We discovered a new cataract mutation, kfrs4, in the Kyoto Fancy Rat Stock (KFRS) background. Within 1 month of birth, all kfrs4/kfrs4 homozygotes developed cataracts, with severe opacity in the nuclei of the lens. In contrast, no opacity was observed in the kfrs4/+ heterozygotes. We continued to observe these rats until they reached 1 year of age and found that cataractogenesis did not occur in kfrs4/+ rats. To define the histological defects in the lenses of kfrs4 rats, sections of the eyes of these rats were prepared. Although the lenses of kfrs4/kfrs4 homozygotes showed severely disorganised fibres and vacuolation, the lenses of kfrs4/+ heterozygotes appeared normal and similar to those of wild-type rats. We used positional cloning to identify the kfrs4 mutation. The mutation was mapped to an approximately 9.7-Mb region on chromosome 7, which contains the Mip gene. This gene is responsible for a dominant form of cataract in humans and mice. Sequence analysis of the mutant-derived Mip gene identified a 5-bp insertion. This insertion is predicted to inactivate the MIP protein, as it produces a frameshift that results in the synthesis of 6 novel amino acid residues and a truncated protein that lacks 136 amino acids in the C-terminal region, and no MIP immunoreactivity was observed in the lens fibre cells of kfrs4/kfrs4 homozygous rats using an antibody that recognises the C- and N-terminus of MIP. In addition, the kfrs4/+ heterozygotes showed reduced expression of Mip mRNA and MIP protein and the kfrs4/kfrs4 homozygotes showed no expression in the lens. These results indicate that the kfrs4 mutation conveys a loss-of-function, which leads to functional inactivation though the degradation of Mip mRNA by an mRNA decay mechanism. Therefore, the kfrs4 rat represents the first characterised rat model with a recessive mutation in the Mip gene.     

The APC/C activator Cdh1 regulates the G2/M transition during differentiation of placental trophoblast stem cells

Differentiation of placental trophoblast stem (TS) cells to trophoblast giant (TG) cells is accompanied by transition from a mitotic cell cycle to an endocycle. Here, we report that Cdh1, a regulator of the anaphase-promoting complex/cyclosome (APC/C), negatively regulates mitotic entry upon the mitotic/endocycle transition. TS cells derived from homozygous Cdh1 gene-trapped (Cdh1^GT/GT) murine embryos accumulated mitotic cyclins and precociously entered mitosis after induction of TS cell differentiation, indicating that Cdh1 is required for the switch from mitosis to the endocycle. Furthermore, the Cdh1^GT/GT TS cells and placenta showed aberrant expression of placental differentiation markers. These data highlight an important role of Cdh1 in the G2/M transition during placental differentiation.     

Identification of two novel amyloid A protein subsets coexisting in an individual patient of AA-amyloidosis

Amyloid A protein (AA), the major fibril protein in AA-amyloidosis, is an N-terminal cleavage product of the precursor protein, serum amyloid A (SAA). Using mass spectrometry and amino-acid sequencing, we identified and characterized two novel AA protein subsets co-deposited as amyloid fibrils in an patient having AA-amyloidosis associated with rheumatoid arthritis. One of the AA proteins corresponded to positions 2–76 (or 75) of SAA2α and the other corresponded to positions 2–76 (or 75) of known SAA1 subsets, except for position 52 or 57, where SAA1α has valine and alanine and SAA1β has alanine and valine in position 52 and 57, respectively, whereas the AA protein had alanine at the both positions. Our findings (1), demonstrate that not only one but two SAA subsets could be deposited together as an AA-amyloid in a single individual and (2), support the existence of a novel SAA1 allotype, i.e., SAA1^52,57Ala.     

Loss of androgen dependency with preservation of functional androgen receptors in androgen-dependent mouse tumor (Shionogi Carcinoma 115).

Shionogi Carcinoma 115 (SC 115) is an androgen-dependent mouse tumor. Chiba Subline 2 (CS 2) is an androgen-independent subline derived from SC 115. CS 2 contains androgen receptors (AR), but is refractory to androgen and does not exhibit androgen-related responses which are observed in SC 115. In the present study the structure and function of AR in SC 115 and CS 2 are examined using cloned cells. There were no gross rearrangements or deletions in the AR genes of these cell lines when compared by Southern blot analysis with the AR gene in the mouse seminal vesicle. SC 115 and CS 2 expressed AR mRNA of normal size. When the cDNA containing DNA- and androgen-binding domains of the AR genes of both cell lines were amplified by polymerase chain reaction, no mutations were found in these regions. SC 115 and CS 2 were transfected with a plasmid containing a long terminal repeat of mouse mammary tumor virus linked to the chloramphenicol acetyltransferase (CAT) gene. Androgen stimulation of these transfectants resulted in equal elevation of CAT activity. These results indicated that the androgen-independent CS 2 contained functionally normal AR which were identical to those in the androgen-dependent parent tumor.     

Fine structural and immunohistochemical studies of goat adenohypophysial cells

Summary The fine structure of each type of anterior pituitary cell in the male goat was studied through the application of a superimposition technique in which adjacent thick sections were used to identify individual cells beforehand by light-microscopic immunohistochemistry. A cone of the pars intermedia protrudes into the pars anterior, being surrounded by the narrow pituitary cleft; the immunohistochemical appearances of the cells forming the cone resemble those of the pars anterior. Several follicles appear in the pars anterior. Ultrastructurally GH cells resemble prolactin cells. The secretory granules of both types are spherical; the diameter of the former is about 340 nm, whereas that of the latter is about 440 nm. ACTH cells are polygonal in shape with secretory granules, about 180 nm in diameter, scattered throughout the cytoplasm. TSH cells, which are spherical in shape, contain the smallest secretory granules, 150 nm in diameter. The highly electron-dense LH cells contain numerous secretory granules about 210 nm in diameter. Their nuclei are irregular with incisures. Thus, the anterior pituitary cells of the goat are ultrastructurally characteristic and species-specific.     

A new snailfish of the genus Careproctus (Cottoidei: Liparidae) from the Pacific coast of southern Japan

Ichthyological Research - A new snailfish, Careproctus tomiyamai, is described on the basis of four specimens collected from Suruga Bay, Tosa Bay, and the Hyuga-nada Sea, southern Japan...     

Effect of gravity stress on fidelity of DNA double-strand break repair.

DNA double strand break (DSB) causes many cytotoxic effects such as cellular lethality, somatic mutation, and carcinogenesis. Fidelity of DSB repair is a important factor that determines the quality of genomic stability. It is known that the most of DSBs are properly repaired on the earth, however, little is known whether those are rejoined at the same fidelity even under the space environment. One of the DSB repair pathway, homologous recombination (HR), allows the cells to repair their DSBs with error free. Therefore, the efficiency of HR is a good index to assess the fidelity of DSB repair. In order to clarify the effect of gravity stress on HR pathway, we established a cell line that can detect a site-specific DNA repair via HR. The cells carrying a reporter construct for HR were incubated under hypergravity condition after induction of site specific DSB. Our preliminary results suggest that the gravity stress may affect the HR efficiency.