JOURNAL zyxwv OF MORPHOLOGY 223:35-46 (1995) zy Morphology and Evolutionary Implications of the Annual Cycle of Secretion and Sperm Storage in Spermathecae of the Salamander zyxwv Ambystoma zyxw opacum (Amphibia: Ambystomatidae) DAVID M. SEVER, JOHN D. zyxwv KRENZ, KRISTIN M. JOHNSON, AND LISA zyxwvutsr C. MIA Department zyxwvut of Biology, Saint Mary’s College, Notre Dame, Indiana 46556 (D M.S., K.M J., L.C.R ); Savannah River Ecology Laboratory, Drawer E, Aiken, South Caroltna 29802 and Institute of Ecology, University of Georgia, Athens, Georgia 30602 zyxwvu (J,D.K.) ABSTRACT Females of the marbled salamander, Ambystoma opacum, store sperm in exocrine glands called spermathecae in the roof of the cloaca. Eggs are fertilized by sperm released from the spermathecae during oviposition. Some sperm remain in the spermathecae following oviposition, but these sperm degenerate within a month and none persists more than 6 mo after oviposition. Thus, sperm storage between successive breeding seasons does not occur. Apical secretory vacuoles are abundant during the fall mating season and contain a substance that is alcian blue+ at pH 2.5. Production of secretory vacuoles decreases markedly after oviposition, and the glands are inactive by the summer months. Ambystoma opacum is a terrestrial breeder, and some mating occurs prior to arrival at pond basins where oviposition occurs. Mating prior to arrival at the ovipository site may prolong the breeding season, leading to fitness implications for both males and females. Females have opportunities for more matings, and the possibilitiesfor sperm competition in the spermathe- cae are enhanced. zyxwvu o 1995 Wiley-Liss, Inc Ambystomatidae is a North American fam- ily of salamanders composed of 27 species (Shafl‘er et al., ’91). The marbled salamander, Ambystoma opacum, is unique among am- bystomatids in that mating and egg-laying occur on land. Adult females and males mi- grate from terrestrial habitats to the borders of ephemeral or permanent ponds in autumn (Noble and Brady, ’33). Eggs are oviposited in areas that will be flooded by rising water, and females typically remain with the eggs until the nests are inundated. Krenz and Scott (’94) reported that 31- 49% ofAmbystoma opacum females captured substantial distances from the nesting area and held in isolation subsequently oviposited fertile eggs. Thus, either males courted and inseminated females before females entered the “breeding site,” or females stored sperm from a previous breeding season. Like most other salamanders, females of Ambystoma opacum possess sperm storage glands called spermathecae in the roof of the cloaca. Eggs are fertilized in the cloaca by sperm released from the spermathecae dur- ing oviposition (Boisseauand Joly, ’75). Sever and Kloepfer (’93) found sperm in the sper- mathecae of females of A. opacum collected at a nesting area before oviposition and while tending eggs after oviposition,but sperm were absent and the spermathecae inactive in an individual sacrificed 62 days after removal from her nest. Herein, we describe the annual cycles of sperm storage and spermathecal activity in a population ofAmbystoma opacum from South Carolina. We used females that oviposited in natural field enclosures in the autumn of 1992 and remained there isolated from males, some for nearly a year. We used morphologi- cal criteria to test the null hypothesis that female A. opacum cannot store sperm be- tween successive breeding seasons. MATERIALS AND METHODS The Ambystoma opacum utilized in this study are indicated in Table 1. Specimens were collected, using a drift fence and pitfall trap technique (Gibbonsand Semlitsch, ’821, along the margin of Bay 23, a Carolina bay on D 1995 WILEY-LISS. INC.