and Maturation of Spermatozoa
The first studies on the process of spermatogenesis and the spermatogonia of the domestic bull were reported in the latter part of the nineteenth century [147], but it was not until 1931 that the chromosome number of spermatogonia and spermatocytes was determined and that the diploid number was 60 [148]. Subsequent studies in the era after the Second World War described the germ cells and their cytoplasmic structures in more detail [147]. It is now well established that spermatogenesis is a complex cellular process whereby spermatozoa, the male haploid germ cell or gamete, are formed from the diploid spermatogonia stem cells through a series of cellular transformations. These complex transformations occur in the seminiferous tubules of the mammalian testes and may proceed over an extended period of time, which is species dependent. Spermatogenesis has been described morphologically in distinct and recognizable cellular “stages” or “phases” that progress through highly organized and precisely timed cycles [35]. During fetal development primordial germ cells migrate to the embryonic testes where they undergo mitotic division to form gonocytes. Just prior to puberty, gonocytes differentiate into the primary pool of A0 spermatogonia, the stem cells from which all subsequent classes of spermatogonia arise. Spermatogenesis proceeds through three distinct stages within the seminiferous tubules of the testis. The first stage is spermatocytogenesis, a proliferative phase where spermatogonia undergo a series of mitotic divisions to form primary spermatocytes. The second stage is meiosis where the primary spermatocytes undergo reduction division of the chromosomal number, from primary spermatocytes (4n) to secondary spermatocytes which are diploid (2n) to the final division that produces round haploid (1n) spermatids. The final stage is differentiation of the haploid spermatids in a process referred to as spermiogenesis, where the rounded spermatids undergo a series of metamorphic changes to form the elongated and flagellated spermatozoa [35, 149].
Spermatocytogenesis
The initial proliferation occurs at the basal membrane compartment of the seminiferous tubules where the germ cells reside. Spermatogonia emerge from the basal germ cell layer, where spermatogonia progress through the differentiation cycle and undergo a series of mitotic divisions, the number of which may be species dependent, forming A1 to A4, I and B spermatogonia. Spermatogonia move forward from the basal lamina of the seminiferous tubules as a cohort connected to each other by intercellular bridges, thereby allowing interconnection of cytoplasm between cells. These cytoplasmic bridges may facilitate intercellular communication and thus support the synchronized development of the cohort of spermatogonia. The bull and ram undergo four mitotic divisions during spermatocytogenesis, yielding 16 primary spermatocytes from each active spermatogonium [18]. During this process, a pool of stem germ cells (dormant spermatogonia A1 and A2) is maintained that provides for new generations of spermatogonia (stem cell renewal). This allows the continual production of spermatozoa in the adult male, but the precise mechanism of this regeneration process or renewal of germinal stem cells is not completely understood [150]. During this process, there may be a level of cell loss resulting from cellular degeneration. These cells are resorbed by Sertoli cell cellular phagocytosis.
Meiosis
Mitotic division provides a continuous supply of spermatogonia, while meiotic division is a process whereby B spermatogonia undergo reduction division to form haploid spermatocytes [35]. This meiotic process not only halves the number of chromosomes in spermatocytes, but also ensures genetic diversity by DNA replication and crossing‐over, random events that produce genetically unique individual spermatozoa. This replication and crossing‐over of DNA material occurs during the first meiotic division, while the second meiotic division results in the formation of the haploid spermatocytes. Spermatocytes may be found at all stages of development in seminiferous epithelium due to the prolonged nature of the meiotic period of spermatogenesis, which ranges from 18 to 21 days in the bull (Figure 2.4a). At the completion of this meiotic process in bulls, each B spermatogonium will result in the production of four haploid spermatids, with a total of 64 spermatids emerging from a single active (A3) spermatogonium [152–154].
Figure 2.4 Schematic representations of the germinal epithelium of the bull testis during stage IV and stage VIII of the seminiferous epithelium cycle. Stage IV of the cycle (a) is characterized by the very active generation of primary spermatocytes into secondary spermatocytes during meiosis I and progress on with the completion of the meiotic division of meiosis II and the formation of spermatids. Stage VIII of the cycle (b) is characterized by the emerging free spermatozoa that have undergone differentiation from the round spermatids.
Source: From [151], © 2003, Germinal Dimensions Incorporated.
Spermiogenesis
The final phase is the differentiation or transformation of the round spermatids into elongated, flagellated, and highly condensed mature spermatozoa that are released into the seminiferous tubule lumen [35]. This process is remarkably similar across domestic species and involves a complex series of events whereby the spermatid undergoes metamorphosis into a highly organized motile cellular structure [33]. Differentiation consists of four main phases that occur in the adluminal compartment between adjacent Sertoli cells: the Golgi phase, the cap phase, the acrosomal phase, and the maturation phase. During the Golgi phase, small Golgi vesicles within the spermatid cytoplasm fuse to form a larger complex structure, the acrosomic vesicle or acrosome. The acrosome is a membrane‐bound vesicle or lysosome that contains several enzymes including acid hydrolase, acrosin, esterases, hyaluronidase, and zona lysine. As differentiation progresses, the acrosome begins to migrate to form a cap (capping) over the nucleus of the cell. During capping, the acrosomic vesicle flattens and covers approximately one‐third of the nucleus [35]. At the same time the nucleus and cytoplasm undergo elongation, and the nucleus begins to occupy the head region of the spermatid. While the head region of the spermatid is undergoing elongation, the midpiece and tail are being formed, thus initiating the maturation phase. Mitochondria migrate in the cytoplasm to form a spiral assembly around the flagellum posterior to the nucleus, which defines the midpiece of the spermatid. The flagellum originates from the distal centriole that gives rise to the axoneme, which is composed of nine pairs of microtubules arranged radially around two central filaments [21]. The axoneme is connected to the base of the nucleus and extends through the midpiece and continues on to form the principal piece or “tail” of the spermatid. As the cytoplasm from the spermatid is shed during the formation of the tail, a cytoplasmic droplet is formed on the neck of the spermatozoon. Once the metamorphosis is complete, the flagellated and elongated spermatid is now referred to as a spermatozoon (Figure 2.4b). These metamorphic changes are all important developmental steps to ensure that the spermatozoa have the ability to be not only motile but also capable of fertilization and delivery of nuclear material when they come in contact with the female gamete (oocyte).
Spermiation
The final stage of development results in the release of the spermatozoa into the lumen of the seminiferous tubules by a process referred to as spermiation [33]. As the final stages of spermiogenesis are completed, the spermatozoa emerge from the adluminal compartment of the seminiferous tubules tail toward the apex of the Sertoli cells and the tubule lumen. During the process of migration from the rete testis through the epididymis and ejaculate, the spermatozoa shed their cytoplasmic droplets, but not all emerge as normal spermatozoa in the ejaculate (Figure 2.5). Before spermatozoa are capable of fertilization they must undergo additional maturation in the epididymis. Capacitation, the final stage of maturation where spermatozoa gain the ability to penetrate the zona pellucida (the outer membrane of the oocyte), occurs in the female