David M. Sever

Ultrastructure of the reproductive system of the black swamp snake (Seminatrix pygaea): Part I. Evidence for oviducal sperm storage

Oviducal sperm storage in the viviparous (lecithotrophic) colubrid snake Seminatrix pygaea was studied by light and electron microscopy. Out of 17 adult snakes examined from May–October, sperm were found in the oviducts of only two specimens. In a preovulatory female sacrificed 14 May, sperm were found in the oviducal lumen and sperm storage tubules (SSTs) of the posterior infundibulum. In a nonvitellogenic female sacrificed 9 June, sperm were found in the lumen and glands of the posterior uterus and anterior vagina, indicating a recent mating. The glands in the posterior infundibulum and vagina were simple or compound tubular, whereas glands in the uterus always were simple tubular. The epithelium of the sperm storage glands was not modified from that lining the rest of the oviduct. The cuboidal or columnar epithelium consisted of alternating ciliated and secretory areas. The secretory product released into the lumen by a merocrine process contained mucoprotein. Lipid droplets also were numerous in the epithelium. Portions of sperm sometimes were embedded in the apical cytoplasm or in secretory material. A carrier matrix containing a mucoid substance, desquamated epithelium, lipids, membranous structures, and possibly phagocytes was found around sperm in the posterior uterus. J. Morphol. 241:1–18, 1999.

Comparative biology of sperm storage in female salamanders.

Females in seven of the ten families of salamanders possess cloacal glands called spermathecae that store sperm. The annual cycle of sperm storage has been studied by light and electron microscopy in eight species representing five families. In these taxa, we recognized 14 characters associated with the spermathecae and traced their evolution on a phylogeny of salamanders based upon other characters. The plasticity and phyletic significance of the spermathecal characters varied greatly. Plethodontids have complex spermathecae while other families possess simple spermathecae; thus, this character has phyletic value as well as being highly conserved within the Salamandroidea. Other characters, such as carbohydrate histochemistry, are highly plastic and show no obvious phyletic trends. The significance of some of these variable characters, such as duration of sperm storage, is apparent only after including in the analysis other aspects of the reproductive cycle, such as length of the mating season. Additional comparative studies, employing the protocol used in this paper, will help further clarify the relationships between phyletic and functional variability in sperm storage mechanisms in salamanders.

Morphology of the mental hedonic gland clusters of plethodontid salamanders amphibia urodela plethodontidae

Mental hedonic gland clusters of 18 species of eastern United States plethodontids were studied by gross dissection and light microscopy. In the Plethodontinae examined, the glands are largely or completely posterior to the mandibular symphysis. In the Plethodontini, the glands are relatively short, vertically oriented in a circular cluster, and bordered dorsally by the dense inner layer of the dermis, the stratum compactum. Within the Hemidactylini genus Eurycea, the glands are more elongate and excretory ducts only occur ventral to and immediately posterior of the mandibular symphysis. The glands then pass horizontally between the intermandibularis posterior muscle ventrally and the geniohyoideus muscle dorsally. The stratum compactum is reduced to a thin sheath over the glands. Reduced or specialized glands resembling those of the Plethodontini occur in the hemidactyliines Hemidactylium, Typhlotriton, and E. multiplicata. From literature reports, the Bolitoglossini contain seven genera with glands like the Plethodontini and one (Oedipina) with glands like Eurycea. In the Desmognathinae, except for Desmognathus wrighti, the excretory ducts are in a fleshy protrusion anterior to the mandibular symphysis, and the short glands extend posteriorly, terminating ventral to and just posterior of the mandibular symphysis. In D. wrighti the gland cluster bifurcates anteriorly with elongate glands then extending posteriorly between the dermis and throat musculature along the medial borders of the dentaries. The excretory ducts in D. wrighti pass dorsally around the anterior ends of the dentaries.

Ultrastructure of the reproductive system of the black swamp snake (Seminatrix pygaea). II. Annual oviducal cycle

This article is the first ultrastructural study on the annual oviducal cycle in a snake. The ultrastructure of the oviduct was studied in 21 females of the viviparous natricine snake Seminatrix pygaea. Specimens were collected and sacrificed in March, May, June, July, and October from one locale in South Carolina during 1998–1999. The sample included individuals: 1) in an inactive reproductive condition, 2) mated but prior to ovulation, and 3) from early and late periods of gravidity. The oviduct possesses four distinct regions from cranial to caudal: the anterior infundibulum, the posterior infundibulum containing sperm storage tubules (SSTs), the uterus, and the vagina. The epithelium is simple throughout the oviduct and invaginations of the lining form tubular glands in all regions except the anterior infundibulum and the posterior vagina. The tubular glands are not alveolar, as reported in some other snakes, and simply represent a continuation of the oviducal lining with no additional specializations. The anterior infundibulum and vagina show the least amount of variation in relation to season or reproductive condition. In these regions, the epithelium is irregular, varying from squamous to columnar, and cells with elongate cilia alternate with secretory cells. The secretory product of the infundibulum consists largely of lipids, whereas a glycoprotein predominates in the vagina; however, both products are found in these regions and elsewhere in the oviduct. In the SST area and the anterior vagina, tubular glands are compound as well as simple. The epithelium of the SST is most active after mating, and glycoprotein vacuoles and lipid droplets are equally abundant. When present, sperm form tangled masses in the oviducal lumen and glands of the SST area. The glands of the uterus are always simple. During sperm migration, a carrier matrix composed of sloughed epithelial cells, a glycoprotein colloid, lipids, and membranous structures surround sperm in the posterior uterus. During gravidity, tubular glands, cilia, and secretory products diminish with increasing development of the fetus, and numerous capillaries abut the basal lamina of the attenuated epithelial lining of the uterus. J. Morphol. 245:146–160, 2000

Sperm storage and degradation in the Spermathecae of the salamander Eurycea cirrigera

Spermathecae are exocrine glands in the roof of the female cloaca that store sperm. Cytological and histochemical data indicate that the one type of secretion into the lumen is a glycoprotein. After a period of stasis in the summer, production of the secretion is initiated in the fall, coincident with an increase in ovarian follicular size. By the time of maximal follicular development and most intense mating activity in March and April, the spermathecal epithelium is filled with secretory granules. The secretory material is released into the lumen, enveloping the sperm. Many sperm remain in the spermathecae after oviposition, and most of these sperm are degraded in the spermathecal epithelium or pass through interruptions in the spermathecal walls caused by desquamation. Sperm in contact with the stromal environment are phagocytized by leukocytes. Some sperm, however, may survive in the lumen until at least the following fall. These sperm retain normal cytology, but whether or not they remain fertile and intact until a subsequent ovipository cycle is unknown.

Spermatophore Formation in Desmognathus ochrophaeus (Amphibia: Plethodontidae)

The relationship between cloacal gland secretion and spermatophore structure is described for Desmognathus ochrophaeus. The spermatophore cap consists of spermatozoa and of granular and globular secretions. Upon initial formation of the cap, these materials are mixed. After being held in the female cloaca, the cap is organized so that the granular and globular material are separate. The portion of the cap deepest in the female cloaca consists of a mass of granular material that fills the inner circumference of the cloacal orifice. The globular material is in layers superficial to the granular material and covers the cloacal orifice. Spermatozoa are scattered throughout the granular material but are most numerous near the globular layers. The granular material of the cap is largely secreted by Kingsburys glands and anterior ventral glands, although some material is added by dorsal and caudal pelvic glands. This material stains positively for acid mucopolysaccharides. The globular material is secreted by pelvic glands and is in two layers. The inner layer, which stains positively for lipids and proteins, is secreted by dorsolateral pelvic glands. The outer layer, which stains positively for proteins, is secreted by dorsal and caudal pelvic glands. The base of the spermatophore stains positively for acid mucopolysaccharides and is secreted by posterior ventral glands. Depletion of glandular products after spermatophore deposition is documented histologically. Spermatophore production and sperm transfer are discussed in the context of courtship behavior in natural situations.

Reproduction of the salamander Siren intermedia le conte with especial reference to oviducal anatomy and mode of fertilization

Reproduction was studied in a South Carolina population of the paedomorphic salamander Siren intermedia with emphasis on anatomy of the female oviduct. The oviduct forms 67–79% of the snout‐vent length in this elongate species and can be divided into three portions. The atrium, 7–13% of oviducal length, is the narrow anteriormost portion, with the ostial opening immediately caudad of the transverse septum. The ampulla, 63–75% of oviducal length, is the highly convoluted, middle portion in which gelatinous coverings are added to the eggs during their passage. Hypertrophy of the oviducal glands in the ampulla causes the ampulla to increase in diameter during the ovipository season. The secretion of the eosinophilic oviducal glands is intensely positive following staining with the periodic acid‐Schiff procedure and does not react with alcian blue at pH 2.5. This staining reaction, coupled with the presence of abundant rough endoplasmic reticulum and Golgi complexes, indicates that the secretion contains a glycoprotein. The ovisac, 16–25% of oviducal length, is the most posterior portion of the oviduct and holds up to 10–11 eggs prior to oviposition. Oviducal glands similar to those in the ampulla are absent in the ovisac. Oviposition in female sirens occurs during February‐April in this population, and male spermiation is concurrent. Entire oviducts were sectioned from three females collected during the ovipository season and from two collected prior to the breeding season, and sperm were not found in the oviducts of these specimens. Thus no evidence was found for internal fertilization or sperm storage in the oviducts of sirens.

Histological analysis of spermatogenesis and the germ cell development strategy within the testis of the male Western Cottonmouth Snake, Agkistrodon piscivorus leucostoma

Cottonmouth (Agkistrodon piscivorus leucostoma) testes were examined histologically to determine the germ cell development strategy employed during spermatogenesis. Testicular tissues from Cottonmouths were collected monthly from swamps around Hammond, Louisiana. Pieces of testis were fixed in Trumps fixative, dehydrated in ethanol, embedded in Spurrs plastic, sectioned with an ultramicrotome, and stained with toluidine blue and basic fuchsin. Spermatogenesis within Cottonmouths occurs in two independent events within a single calendar year. The testes are active during the months of March-June and August-October with spermiation most heavily observed during April-May and October. To our knowledge, this is the first study that describes bimodal spermatogenesis occurring in the same year within the subfamily Crotalinae. During spermatogenesis, no consistent spatial relationships are observed between germ cell generations. Typically, either certain cell types were missing (spermatocytes) or the layering of 3-5 spermatids and/or spermatocytes within the same cross-section of seminiferous tubule prevented consistent spatial stages from occurring. This temporal pattern of sperm development is different from the spatial development found within birds and mammals, being more reminiscent of that seen in amphibians, and has now been documented within every major clade of reptile (Chelonia, Serpentes, Sauria, Crocodylia). This primitive-like sperm development, within a testis structurally similar to mammals and birds, may represent an intermediate testicular model within the basally positioned (phylogenetically) reptiles that may be evolutionarily significant.

Regionalization of eccrine and spermiophagic activity in spermathecae of the salamander Eurycea cirrigera (Amphibia: Plethodontidae)

The spermathecae of female Eurycea cirrigera are compound alveolar glands; narrow neck tubules connect the distal bulbs to a common tube that opens onto the roof of the cloaca. The common tube and neck tubules produce apical secretory vacuoles that contain a periodic acid‐Schiff (PAS)+ substance for merocrine export into the lumen. This substance is produced throughout the year, although secretory vacuoles are less numerous during the period of reproductive inactivity in the summer. When sperm are present, the product from the secretory vacuoles bathes sperm in the lumen. Sperm are in orderly arrays and never are embedded in the cytoplasm of the common tube or neck tubules. The distal bulbs do not produce PAS+ secretory vacuoles, and are actively spermiophagic as long as sperm are present. Sperm become embedded in the epithelium of the distal bulbs where lysosomes degrade sperm.

Sperm Storage in the Spermatheca of the Red-Back Salamander, Plethodon cinereus (Amphibia: Plethodontidae)

In northern Indiana, the mating season of Plethodon cinereus occurs after hibernation from March until June, when oviposition begins. During the mating season, a female stores sperm in its spermatheca, a compound tubular gland in the roof of the cloaca. The apical cytoplasm of the spermathecal epithelium is filled with large secretory vacuoles whose product is released while sperm are stored. Females induced to oviposit in June and July by injections of human chorionic gonadotropin (hCG) still retain much sperm 1 month after oviposition, but secretory vacuoles are absent in all specimens sacrificed in July and August. Instead, some sperm are embedded in the spermathecal epithelium with resultant spermiophagy involving lysosomes. A female sacrificed in September 2 months after oviposition possesses scant sperm, but spermiophagy alone does not seem extensive enough to account for the decrease in sperm numbers. Females sacrificed in October prior to hibernation lack sperm in their spermathecae; some secretory vacuoles are present, but they are not as numerous or as enlarged as in specimens collected in March and May. Inter‐ and intrafamilial differences in the cytology of sperm storage may not be phyletically informative at the family level but related to species‐specific reproductive adaptations. J. Morphol. 234:131–146, 1997.