title: Effects of cryopreservation methods on post-thaw motility of spermatozoa from the Japanese pearl oyster, Pinctada fucata martensii.
abstract: In order to develop cryopreservation techniques for Japanese pearl oyster spermatozoa, the effects of various cryopreservation conditions on post-thaw motility were examined. Spermatozoa cryopreserved with 10% methanol (MET), dimethylformamide or dimethylacetamide plus 90% diluent comprising 80% seawater and 20% fetal bovine serum (FBS) showed higher percentages of post-thaw motility than those cryopreserved with 10% dimethylsulfoxide or glycerol. When spermatozoa were cryopreserved with various concentrations (0-20%) of MET and 100-80% diluent, 10% MET showed the highest percentages of post-thaw motility. When spermatozoa were cryopreserved with 10% MET and 90% diluent comprising various concentrations (0-100%) of FBS or Ringer solution mixed with seawater, the percentages of post-thaw motility peaked at 20% FBS or Ringer solution, and were significantly higher for 20% FBS than for 20% Ringer solution. The percentages of post-thaw motility increased with increasing dilution ratios from 2.5- to 50-fold. Spermatozoa cooled to -50 degrees C and then immersed in liquid nitrogen (LN) showed higher post-thaw motility than those cooled to -30 degrees C or -40 degrees C. When spermatozoa were cryopreserved to -50 degrees C at various cooling rates by changing the sample height above the LN surface, the post-thaw motilities of spermatozoa cooled at 10 cm (cooling rate: -21.3 degrees C/min) and 12.5 cm (-15.6 degrees C/min) from the LN surface were higher than those at 5, 7.5 or 15 cm. These results indicate that 10% MET plus 90% diluent comprising 80% seawater and 20% FBS is a suitable extender for cryopreservation of Japanese pearl oyster spermatozoa and that samples should be cooled to -50 degrees C at a cooling rate between -15 and -20 degrees C/min for efficient storage.
authors.affiliation: Department of Fisheries, Graduate School of Agriculture, Kinki University, Nara 631-8505, Japan.
authors.forename: Takayuki
authors.initials: T
authors.lastname: Kawamoto
authors.forename: Teruyoshi
authors.initials: T
authors.lastname: Narita
authors.forename: Kiyoshi
authors.initials: K
authors.lastname: Isowa
authors.forename: Hideo
authors.initials: H
authors.lastname: Aoki
authors.forename: Masahiro
authors.initials: M
authors.lastname: Hayashi
authors.forename: Akira
authors.initials: A
authors.lastname: Komaru
authors.forename: Hiromi
authors.initials: H
authors.lastname: Ohta
mesh_terms: D000818:Animals; D015925:Cryopreservation / Q000379:methods*; D003451:Cryoprotective Agents / Q000494:pharmacology; D008297:Male; D049911:Pinctada / Q000502:physiology*; D012662:Semen Preservation / Q000379:methods; D013081:Sperm Motility; D013094:Spermatozoa / Q000187:drug effects / Q000502:physiology*; D013696:Temperature; D013997:Time Factors
chemical_list: D003451:Cryoprotective Agents