Effects of frozen and liquid hypothermic storage and extender type on calcium homeostasis in relation to viability and ATP content in striped bass (Morone saxatilis) sperm

Abstract The effect of hypothermic storage on striped bass sperm calcium homeostasis was determined by Fluo-3 flow cytometry. Calcium homeostasis was defined as the ability of cells to maintain a low concentration of intracellular free calcium as measured by Fluo-3 fluorescence. Sperm were stored frozen in striped bass extender (SBE) and Tris–NaCl medium (T350) modified with 50 mM glycine and 7.5% dimethylsulfoxide and in nonfrozen form diluted 1:3 (vol/vol) in SBE and T350 for 1, 24, and 48 hours at 4 °C in an oxygen atmosphere. Fluo-3 fluorescence was detected in less than 5% of fresh viable sperm cells indicating maintenance of calcium homeostasis. In contrast to sperm in fresh semen, frozen-thawed and nonfrozen sperm cells lost to a considerable extent the ability to maintain low intracellular free calcium even in the absence of exogenous calcium; positive Fluo-3 fluorescence was found in 26% and 39% of thawed sperm frozen in SBE- and T350-based freezing diluents, respectively, and increased (P 0.05) in SBE after 48 hours of nonfrozen storage but decreased (P