Y. Clermont

DEFINITION OF THE STAGES OF THE CYCLE OF THE SEMINIFEROUS EPITHELIUM IN THE RAT

Any area of any seminiferous tubule of the rat contains a few spermatogonia along the basement membrane, one or several layers of spermatocytes farther in, and groups of spermatids next to the lumen of the tubule. The present work is a systematic study of the various modes of association of these cells in what is known as the seminiferous epithelium. In the rat, development of any one generation of spermatogonia, spermatocytes, or spermatids is closely integrated with that of other generations present in the same area of the tubule. Thus, the cel!s are not arranged a t random, but are organized into well-defined cellular associations. I t was realized in the 1890’s that the various cell associations must succeed one another in time in any given area of the seminiferous tubule, and that the sequence must repeat itself indefinitely. The complete series of successive cellular associations has been named the “ spermatogenic cycle” by von Ebnerl and Regaud.? This designation is somewhat misleading and, more recently, has been used to describe the whole of spermat~genesis.~ To avoid confusion, the more exact designation “cycle of the seminiferous epithelium” or, simply, “cycle” will be used instead. The cycle of the seminiferous epithelium may be defined as that series of changes occurring in a given area of the seminiferous epithelium between two successive appearances of the same cellular association. The number of cellular associations identified by various authors within a cycle ot the seminiferous epithelium shows large (TABLE 1.) The discrepancies may be explained by the fact that many stages of development of the cells cannot be characterized with precision in preparations stained with hematoxylin. In the present work, the cellular associations were defined by the stage of development of the spermatids, which themselves were characterized by their staining reaction to the “periodic acid-Schiff”, also called “periodic acid-fuchsin sulfurous acid” technique (hereafter referred to as PAFSA). The stages through which the developing spermatids progress can be easily identified by this technique, as shown in a previous paper,l0 in which the bibliography of the subject may be found. Spermiogenesis may thus be divided into 19 stages, numbered from 1 to 19 (FIGURES 1-19). During the first 8 stages of the development of young spermatids, the seminiferous epithelium also contains a generation of older spermatids, which are released when the younger generation completes Stage 8. During the next few stages of development, the remaining spermatids are the only ones present, but when they reach Stage 15, a new generation again arises

Spermatogenesis in man: an estimate of its duration.

Testicular biopsies were obtained at various intervals after intratesticular injections of tritiated thymidine. Radioautographs revealed that the most mature labeled germ cells were the preleptotene spermatocytes at 1 hour, the midpachytene spermatocytes at 16 days, and the immature spermatids at 32 days after injection. These results indicated that one cycle of the seminiferous epithelium lasts 16 days, and the whole of spermatogenesis is estimated to consume approximately 64 days.

EXISTENCE OF A SPERMATOGONIAL CHALONE IN THE RAT TESTIS

Adult rats with normal or X‐irradiated testes were used in an experiment to test the possible existence of a chalone in the testis. On the 11th day following irradiation, i.e. as the type A spermatogonia proliferated actively to restore the partially destroyed spermatogonial population, the animals with irradiated testes were subdivided into three groups. Rats of the first group were injected intraperitoneally with a saline extract of normal adult rat testes. Animals of the second group were injected with an equal amount of physiological saline while the rats of the third group received equivalent injections of a saline liver extract. Two additional groups of rats with non‐irradiated testes, injected with the testicular extract or saline solution, served as controls. Following the last injection all animals were injected with 3H‐thymidine and sacrificed. From each animal one testis was used to determine the specific radioactivity of its DNA, the other testis was processed for radioautography. The testicular extract produced a significant decrease in uptake of radioactivity by the irradiated testes. There was no difference in the radioactivity uptake by the testes of non‐irradiated rats. Correspondingly the labeling index of type A spermatogonia was significantly lower in animals of the first group than in the other two groups of animals with irradiated testes. However, there was no difference in the labeling indices of Intermediate and type B spermatogonia or of preleptotene spermatocytes in the animals receiving the extracts or the saline solution. In animals with non‐irradiated testes there was no difference in the labeling indices of type A or other types of spermatogonia or of spermatocytes. These data were taken to indicate that a saline extract of normal adult testes contains a substance that can inhibit specifically the proliferation of type A spermatogonia during the repair phase of the spermatogonial population following irradiation. This substance was tentatively considered as a spermatogonial chalone.

Chapter 11 Three-Dimensional Structure of the Golgi Apparatus

Publisher Summary This chapter discusses the Golgi apparatus of Sertoli cells and spermatids. In both cases, the backbone of the Golgi apparatus is made up of saccular regions interconnected by intersaccular regions composed of anastomotic tubules. On the cis face, the cis -osmiophilic element displays a similar appearance in both types of cells. In contrast, the transtubular network observed in the Sertoli cells on the transface of the Golgi apparatus is replaced in the spermatid by elements of the medullary zone, the three-dimensional organization of which remains to be elucidated. Finally, relations between endoplasmic reticulum (ER) and Golgi elements, which in the Sertoli cells are restricted to the transface of the Golgi backbone, are more numerous in the spermatid, where they are observed not only on both faces of this organelle but also in the intersaccular and saccular regions. The Golgi apparatus forms a continuous structure made up of alternating saccular and intersaccular connecting regions. In these regions, the saccules and bridging tubules form the main component or backbone of the Golgi apparatus.

Glucose-6-phosphatase activity of endoplasmic reticulum and Golgi apparatus in spermatocytes and spermatids of the rat: an electron microscopic cytochemical study

Summary— The glucose‐6‐phosphatase (G6Pase) activity of cytoplasmic components of spermatocytes and spermatids of the rat was examined by electron microscope cytochemistry using cerium chloride as a capture agent. G6Pase activity, a recognized ER‐resident enzyme, was present in all ER cisternae of spermatocytes. In spermatids, while some ER cisternae were G6Pase‐reactive, others were negative or only slightly reactive, indicating an unequal distribution of the enzymatic activity throughout the network of ER cisternae in these cells. In spermatocytes, the cis‐ and trans‐elements of the stacks of Golgi saccules were slightly but significantly reactive for G6Pase. In the Golgi apparatus of spermatids, the cis‐element, 4 or 5 underlying saccules, as well as one or two thick trans Golgi elements were G6Pase reactive. The G6Pase activity of the various Golgi elements, like that of the ER cisternae was not affected by the pH of the medium and was completely inhibited by Na‐vanadate, a known G6Pase inhibitor. Sertoli and Leydig cells, submitted to the same cytochemical conditions, showed complete G6Pase reactivity of their ER; however in Sertoli cells, all Golgi components were consistently negative while in Leydig cells the cis‐ and trans‐elements of the Golgi stacks were slightly reactive, as in spermatocytes. Thus, the G6Pase reactivity of Golgi elements, appeared variable from one cell type to another. The compact juxtanuclear Golgi apparatuses of spermatocytes and spermatids were both associated with numerous G6Pase reactive ER cisternae; some were present at their surface, others crossed their cortices between Golgi stacks and formed elaborate networks in their cores. Other ER cisternae, in turn were closely apposed to or intertwined with trans‐elements of the Golgi stacks, a relationship raising the possibility of molecular exchanges at this pole of the stacks in germinal cells.

Biogenesis of Specialized Cytoskeletal Elements of Rat Spermatozoa

Our results on the formation of the ODF and perforatorium are diagrammatically summarized in Figures 30 and 31. The developmental expression of proteins making up these two cytoskeletal elements differs in timing, duration and intracellular localization. The ODF proteins are synthesized exclusively during the latter part of spermiogenesis, well after transcriptional activity in the haploid germ cell nucleus has ended. This implies that these major integral proteins of the tail are translationally regulated and that mechanisms must exist for the storage and eventual release of the mRNAs encoding these proteins. The perforatorial proteins, on the other hand, begin to be synthesized during the meiotic prophase reaching a peak of production in early spermiogenesis just before the initiation of the condensation of the spermatids nucleus, at which time RNA synthesis stops. Another major difference between ODF and perforatorial protein production is that there seems to be a coordinated activity between the synthesis and the assembly of the ODF proteins, whereas there appears to be an almost 25 day delay between the initial meiotic synthesis and final condensation of perforatorial proteins in the subacrosomal space at the end of spermiogenesis. As for the intracellular localization of ODF and perforatorial proteins both have unprecedented distributions. The ODF proteins appear to be concentrated in a particular type of granular body which is especially abundant in the elongated spermatid at the time of peak ODF assembly. The perforatorial proteins, on the other hand, appear to be concentrated in the nuclei of pachytene spermatocytes and round spermatids until their displacement into the cytoplasm during nuclear condensation. Both forms of localization suggest a storage role for these proteins uniquely adapted by the spermatid to regulate the assemblies of the respective cytoskeletal elements.

EFFECT OF A SPERMATOGONIAL CHALONE ON THE GROWING RAT TESTIS

Immature rats were used in an experiment to test the possible influence of a spermatogonial chalone on the expanding spermatogonial population in their developing testes. An extract from adult rat testes was injected intraperitoneally into 33‐day‐old rats and control animals were injected with an equal amount of saline. Two groups of normal adult rats similarly injected with the testicular extract and saline solutions served as additional controls. Following these injections, all animals were administered a dose of 3H‐thymidine 10 hr before sacrifice. An analysis of the labeling indices of the various types of spermatogonia revealed that in young rats injected with testicular extract the percentage of labeled type A spermatogonia was significantly lower than in control animals. In contrast, the labeling indices of Intermediate and type B spermatogonia were similar in the two groups of young rats. In the two groups of adult animals, there was no difference in the labeling indices of type A or of other types of spermatogonia. These data indicated that the saline extract of adult testes contained a substance, a spermatogonial chalone, inhibiting specifically the proliferation of some type A spermatogonia. The results also support the concept that a spermatogonial chalone may intervene, through its action on the spermatogonial stem cell population, to arrest the growth of the seminiferous tubules as the animal reaches maturity.

TRANSFORMATIONS OF MEMBRANE-BOUND ORGANELLES IN SEC14 MUTANTS OF THE YEASTS SACCHAROMYCES CEREVISIAE AND YARROWIA LIPOLYTICA

In early descriptions of ultrastructural alterations of secretory (sec) mutants of the yeast Saccharomyces cerevisiae, two mutants, sec7 and sec14, were shown to produce cell structures, the so‐called Berkeley bodies thought at first to correspond to Golgi structures. In sec7 mutants grown at restrictive temperature, secretion granules soon disappeared, whereas networks of Golgi tubules increased in size and transformed into stacks of seven to eight flattened elements. At these time intervals, structures resembling Berkeley bodies appeared to be extensions of the endoplasmic reticulum (Rambourg et al., 1993). It is the purpose of the present study to examine by electron microscopy S. cerevisiae sec14 mutants and to compare the modifications along their secretory pathway with those occurring in a homologous mutant of Yarrowia lipolytica.

Modifications of the golgi apparatus in Saccharomyces cerevisiae lacking microtubules

Disassembly of cytoplasmic microtubules by nocodazole in cultured mammalian cells leads to the disruption of the continuous ribbonlike Golgi apparatus and dispersal of the Golgi elements from their normal juxtanuclear location, close to the microtubule‐organizing center (MTOC), toward the cell periphery. Clearing of the drug induces reassembly of the microtubules from the MTOC and reorganization of the Golgi elements into a continuous ribbonlike juxtanuclear structure. In the yeast Saccharomyces cerevisiae, the Golgi apparatus does not form a continuous structure as in mammalian cells but instead constitutes independent units dispersed throughout the cytoplasm. It is the purpose of this article to investigate the role of microtubules in the structure and distribution of the Golgi elements in S. cerevisiae by studying the ultrastructure of cell organelles either in mutant cells deficient in β‐tubulin or in wild‐type cells treated with the microtubule‐depolymerizing drug nocodazole.

Dynamics of Human Spermatogenesis

The production of spermatozoa by the seminiferous epithelium of mammals has been and still is the object of numerous cytological and histological investigations (1,2,3). Spermatogenesis in man has not been completely ignored by microscopists, even if it is difficult to obtain properly fixed sections of testis from normal adult individuals for investigative purposes. From the morphological point of view the phenomena occurring within the seminiferous tubules may be classified as being either cytological or histological. Amongst the cytological phenomena there are: the mode of proliferation, renewal and differentiation of spermatogonia, the meiotic divisions of spermatocytes, the metamorphosis of spermatids into spermatozoa, and the release of spermatozoa from the seminiferous epithelium. At the histological level, considering the seminiferous epithelium as a whole, there is a grouping of germ cells into distinct cellular associations. The chronological evolution of these cellular associations is known as the “cycle of the seminiferous epithelium”, while the orderly distribution of the cell associations along the seminiferous tubule is referred to as the “wave of the seminiferous epithelium”. In the following brief discussion three topics concerning the dynamics of human spermatogenesis will be examined: firstly the steps of spermatogenesis, secondly the cycle of the seminiferous epithelium and its duration, and thirdly, the mode of renewal of spermatogonia.