Based on light microscopic findings representing the full sequence of spermatogenesis and spermiogenesis from the capsule to the luminal surface (Fig. 1), ultrastructural features were carefully examined. In individual primary spermatogonia, comparatively large oval nuclei with prominent nucleoli in euchromatin were tightly packed and laid on the folded basement membrane beneath the testicular capsule. Several mitochondria and fine electron-dense ribosomes were present in scanty cytoplasm (Fig. 2A, B). In much more advanced spermatogonia, patches of heterochromatin were noted in the nucleus, and the cytoplasm was slightly more voluminous containing cytoplasmic organelles, especially mitochondria and ribosomes (Fig. 2C, D). Eight primary spermatocytes, in accordance with established understanding that these came from 3 mitotic divisions of an individual spermatogonium, formed a group with a rosette-like appearance, gradually developing a central common mass called a cytophore (Fig. 3A, B). In early spermatocytes, pale round nuclei were larger than those of spermatogonia, and each had an obvious nucleolus (Fig. 3C). The synaptonemal complexes were also demonstrated in the nuclei of early prophase-I meiotic division of the primary spermatocytes (Fig. 3D, E). At late prophase-I meiotic division, an irregularly shaped nucleus without a nucleolus was noted, in which the chromatin was condensed into chromosomes (Fig. 3F, G).

Meiosis-I is known to produce 16 secondary spermatocytes. During this stage, the volume of the cytophore gradually increased. The nuclei of secondary spermatocytes were much more electron-dense, 4-fold smaller than those of primary spermatocytes and with an increased nucleo-cytoplasmic ratio (Fig. 4A). The nuclei were dislocated away from the central cytophore which showed completely fused cell membranes with the neighbors. The proximal plasma membranes between neighboring spermatocytes became highly convoluted (Fig. 4B). It was followed by meiosis-II which produces 32 spermatids from 16 spermatocytes. The earliest spermatids were of elongated oval shape, and the cell membranes at their apical margins were associated with electron-dense undercoats (Fig. 5A, B). Their nuclei were oval in shape and had chromatin of fine thread-like reticular arrangement intermingled with nucleoli. At the stage of spermiogenesis, in which the spermatids become differentiated into the spermatozoa, the spermatid nuclei were increasingly elongated and curved in shape. The nuclear chromatin changed into bundles of electron dense lamellar scroll-like configurations which were arranged in parallel with the longitudinal axis of the nuclei, resulting in a honeycomb-like appearance in cross section (Fig. 5C-E). A cone-shaped projection of the cytoplasm appeared on the surface of the cytophore in conjunction with each nucleus. It contained several kinds of cytoplasmic organelles such as mitochondria, Golgi apparatus, rough endoplasmic reticulum and free ribosomes (Fig. 6A). As the spermatid-cone grew outward, its base sank slightly into the cytoplasm of the cytophore, forming a shallow ring-like groove delineated by arching membranes of increasing electron density. The thickening membranes close to the cytophore, called zone of differentiation, showed a density increase associated with the arching membranes, which was found to be a single row of microtubules immediately underneath the cell membrane (Fig. 6A). The median cytoplasmic process (MCP) arising away in an opposite direction from the cytophore was also shown (Fig. 6A). The flagellar complex also arose from the zone of differentiation, and was composed of a middle piece of the intercentriolar body sandwiched by 2 flagellar axonemes which anchored the basal bodies and the striated rootlets, respectively (Fig. 6B-D).

The flagellar axonemes consisted of 9 doublet tubules concentrically arranged around a single central core complex (Fig. 6C, D). At the early stage of flagellar development, they were arranged almost perpendicular to the MCP axis (Fig. 6B-D). As the development proceeded, the emergent flagella were rotated and arranged closer and in parallel to the longitudinal axis of the MCP of the developing spermatozoon (Fig. 7A-C) and they finally fused with the MCP. The arching membrane portions also gradually moved centrally to make a narrow constricted portion (Fig. 7A-D) and then finally pinched off the head portion of the spermatid from the central mass of the cytophore, resulting in formation of the spermatozoon (Fig. 7E). In synchrony with this change, mitochondria grew in the long axis of the spermatid and spermatozoon. Subsequently large vacuoles appeared in the cytophore, which represents a stage of degeneration (Fig. 7F). Spermatozoa, when pinched off, did not contain the central body, centriole and striated rootlets anymore. However, axonemes still remained and they were already fused with the MCP (Fig. 8). Two different types of spermatozoa were distinguished; the first type with translucent cytoplasm and the second type with densely packed glycogen granules (Fig. 8). Rows of microtubules were arranged underneath the cell membrane of the head and middle portions of spermatozoa of both types. Additionally, the elongated mitochondria also appeared centrally in the core of both types. Elongated nuclei of high electron density were also apparent in both types.

The SEM observations were focused on the spermatids and spermatozoa. Flagella of fine thread-like appearance were shown to extend from the cone-shaped proximal region of the spermatids (Fig. 9A). The pinched-off head and thicker elongated tails of the spermatozoa were also shown (Fig. 9C). Spermatids and mature spermatozoa were shown to be characterized by frequent occurrence of flagella and tails bundles respectively (Fig. 9).