Yoji Hakamata

To clarify a function of brain-type ryanodine receptor (RyR3) and its regulation, we established a stable cell line expressing rabbit RyR3 by transfection of Chinese hamster ovary cells (CHO cells) with the cDNA and investigated characteristics of the RyR3. Scatchard analysis of [3H]- ryanodine binding to the membrane from CHO cells expressing RyR3 showed two distinct binding sites. The Kd values of high and low affinity binding sites were 1.92 and 25.9 nM, respectively. [3H]-ryanodine binding to the membrane from CHO cells expressing RyR3 was dependent on pCa. Extracellular Ca2+ (2- 10 mM) and high concentration (more than 30 mM) of caffeine activated the RyR3 in CHO cells and increased its intracellular Ca2+ concentration. The enhancement of [3H]-ryanodine binding to the membrane from CHO cells expressing RyR3 was observed by bromoeudistomin D (BED), a caffeine -like powerful Ca2+ releaser, at pCa 5.5. Stably expressed RyR3 in CHO is useful for characterization of its function.

We have cloned and sequenced the cDNA of the human brain ryanodine receptor (RyR3), which is composed of 4866 nmino acids and shares characteristic structural features with the rabbit RyR3. Northern blot analysis shows that the human RyR3 mRNA is abundantly expressed in hippocampus, caudate nucleus and amygdala as well as in skeletal muscle. The human RyR3 mRNA is also detected in several cell lines derived from human brain tumors. Functional expression of RyR3 and a chimeric RyR suggests that RyR3 forms a calcium-release channel with a very low Ca2+ sensitivity.

The male hypogonadism rat (hgn/hgn) shows a characteristic male sterility as a single autosomal recessive trait. Recently, the female homozygotes for hgn, assumed to be fertile, could be detected by a /jgji‐associated hypoplastic kidney (hpk/hpk). The present study was to investigate a possible influence of the hgn gene on female reproduction. The hgn/hgn females showed a significant growth retardation as compared with the phenotypically normal ones (+/? +/hgn or +/+). The litter size at birth and number of implantation traces were significantly less in the hgn/hgn than in the +/? females. The hgn/hgn females became anestrous and infertile much earlier than the +/? did. Histologically, there were a few corpora lutea, some atretic follicles at different stages of maturation and abundant abnormal interstitial cells with pyknotic or karyorexic nuclei in the ovaries of hgn/hgn females that have been infertile. The birth rate expressed by per cent litter size at birth against number of implantation traces was comparable between the hgn/hgn and the +/? female, suggesting that the small litter size of hgn/hgn female could not be due to the embryonic death in utero. Nevertheless, the number of the tubal ova at estrous was comparable in the hgn/hgn and +/? females. Therefore, it was suggested that the half of ova or embryos may be lost during the period from the fertilization to the implantation. Histological appearances of the neonatal ovary in the hgn/hgn seemed hypoplastic. The number of cells including oocytes and interstitial cells, enzymatically separated from neonatal ovary, was significantly less in the hgn/hgn than in the +/hgn. These results suggest that the gene product(s) coded by normal allele of hgn gene(s) involves normal gonadal development in both sexes the defect may lead testicular dysmorphology in the male and reduced fertility in the female. Copyright © 1992, Wiley Blackwell. All rights reserved

The presence of a basic fibroblast growth factor-like immunoreactive substance was demonstrated in the nuclei of germ cells at stages from spermatocyte to spermatid in adult rat testis by using immunohistochemistry with an antibody raised against a synthetic peptide corresponding to residues 1-10 of bovine basic fibroblast growth factor [1-146]. The fluorescence was very weak in the nuclei and cytoplasm of spermatogonia, Sertoli cells, and most of the interstitial compartments, except for capillary endothelial cells. This is the first study to demonstrate the presence of basic fibroblast growth factor-like immunoreactive material in the nuclei of haploid cells in vivo.

Abstract Bilateral hypoplastic kidneys have been found to associate with the male hypogonadism rat (hgn/hgn). The hypoplastic kidney was persistent to the male homozygote for hgn and it was also seen in some females with unknown genotype. The affected kidney was apparently smaller in size than phenotypically normal one. When females with hypoplastic kidney were mated to proven heterozygous males for hypogonadism (hgn/+), the ratio of the affected testis to phenotypically normal one was 32: 20 in the Fl generation. This segregation ratio did not deviate siginificantly from the 1: 1 ratio expected from the hypothesis that the genotype of females with hypoplastic kidney in parent generation for hypogonadism would consist of hgn/hgn alone. The ratio expected from the other hypotheses was denied statistically. The ratio of the affected kidney to phenotypically normal one in both sexs of the Fi was 53: 50, fitting to 1: 1 hypothesis for a single autosomal recessive trait. There was neither a case showing the phenotypically normal kidney with the affected testis, nor the hypoplastic kidney with the phenotypically normal testis. The results suggest that the gene responsible for the hypoplastic kidney and the gene for hypogonadism would be identical or both genes reside in the close vicinity in a chromosome. The results also suggest that the homozygotes for hgn can be selected by the presence of hypoplastic kidney. Copyright © 1991, Wiley Blackwell. All rights reserved