The ultrastructure of testicular interstitium in young and aged adult rats was analysed using morphometric methods, and the plasma testosterone concentration was measured. With increasing age there was an augmentation in the volume of collagen fibrils in the intercellular matrix and in blood vessels. During the aging process (approximately two years) the average volume of the Leydig cell decreased from 1364 microns 3 to 637 microns 3, but the number of Leydig cells in paired testes increased from 53 x 10(6) to 113 x 10(6). The absolute volume of smooth surfaced endoplasmic reticulum (SER) per Leydig cell amounted in aged rats to 78% of that in young adult rats. The total amount of SER in paired testes increased by 62% with aging. The present analysis suggests that the ability of SER to maintain peripheral testosterone concentration decreases with age. In young adult rats the absolute volume of peroxisomes per Leydig cell correlated significantly with the concentration of testosterone in blood and also with the absolute volume of SER per Leydig cell. These results combined with ultrastructural observations of close apposition of peroxisomes and SER suggest that peroxisomes have a role in testosterone secretion by Leydig cells.

ABSTRACT. The interaction between simian virus 40(SV40)-induced endocytotic vacuoles and the nuclear membrane was investigated using cationized ferritin (CF) and concanavalin A (Con A) as cell membrane markers. These markers bound to the cell surfaces of CV-1 cells together with SV40 at 4°C. Following incubation of these modified cells at 37°C in serum-free medium, the cell membranes showed many invaginations. After incubation for 60 min at 37°C in the same medium, many various-sized vacuoles were present that contained membrane-bound CF, Con A and SV40. After 2 h of incubation at 37°C, Con A was present in some areas of the perinuclear cisterna along the nuclear membrane. The control experiment, however, showed no localization of Con A-binding on the nuclear membrane. These results provide evidence that SV40-induced endocytotic vacuoles migrate toward the nucleus and fuse with its membrane.

Key words: SV40/cell membrane markers/endocytotic vacuoles/nuclear envelope/diaphragm/interaction ABSTRACT. The transfer of endocytosed simian virus 40 (SV40) to the nuclear position was investigated ultrastructurally using cationized ferritin (CF), ferritin labelled concanavalin A (Fer-Con A) and Con A as cell membrane markers. In the cells incubated with these markers and SV40 at 4°C, and then chased for 2 h at 37°C in serum-free medium, ferritin particles representing CF and/or Fer-Con A binding sites were found in vacuoles with SV40. The membraneof some vacuoles seemedto be in contact with the outer nuclear membrane.Several ferritin particles were located in the perinuclear cisterna and within the nucleoplasm, but not within the nuclear pores. In addition, there were vacuoles with ferritin particles and SV40near the nuclear membrane, which looked like a single diaphragm with heterochromatins inside it. The outer nuclear and vacuole membraneswere often obscure in the areas where the vacuole was very close to the diaphragm. In the case of cells incubated with CF, SV40 and Con A at 4°C, chased for 2 h at 37°C, and then reacted with horseradish peroxidase (HRP), HRP activity showing Con A-binding sites was also observed along the nuclear side of the inner nuclear membraneas well as in the perinuclear cisterna along the outer membrane. These results confirm that SV40-induced endocytotic vacuoles fuse with the outer nuclear membrane,and further indicate that someendocytotic vacuoles may well interact directly with the diaphragm, suggesting another path for migration of SV40 into CV-1 cell nuclei besides the path going through the process of fusion of the vacuole membrane with the outer nuclear membrane.

JapanKey words: testosterone-bovine serum albumin conjugate/target cell nuclei/colloidal gold/silver enhancement/in vivo/electron microscope ABSTRACT. Weexaminedwhether testosterone-bovine serum albumin conjugate (testosterone-BSA) showed similar distribution to radiolabeled testosterone in vivo, by injecting 2-nm colloidal gold labeled-testosterone-BSA (testosterone-BSA-gold) from rat tail vein. The testosterone-BSA-gold with the silver enhancement becamevisible as silver deposits under electron microscope in nuclei of Leydig cells, Sertoli cells, spermatogonia, spermatocytes and spermatids in the testis, those of the epithelial cells in the seminal vesicle and of the cardiac muscle cells in the heart of rat killed 2 h after the injection. Few deposits were present on the non-target cell nuclei in thymus and spleen. In the liver cells, the deposits were observed in the cytoplasm, but few in the nucleus. At high-power magnification without silver enhancement, the gold particles were found in the target cell nuclei in the testis. In control rat injected with BSAlabeled with 2-nm colloidal gold, the percentages of nuclei showing the deposits were fewer than those in the rat injected testosterone-BSA-gold in the target cells. The deposits were also few in the nuclei of non-target cells in control rat. These results suggest that testosterone-BSAgold is useful for morphological study of testosterone target cells, and imply that BSAconjugated with testosterone can enter the target cell nuclei of the rat.

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