Wayman W R, Thomas R G, Tiersch T R
School of Forestry, Wildlife, and Fisheries, Louisiana Agricultural Experiment Station Louisiana State University Agricultural Center, Baton Rouge, LA, USA.
Theriogenology. 1997 Jun;47(8):1519-29. doi: 10.1016/s0093-691x(97)00158-1.
Procedures were developed for the collection, refrigerated storage and cryopreservation of black drum spermatozoa. Sperm samples were collected by removing and slicing the testis, and suspending the spermatozoa in Hanks' balanced salt solution (HBSS) at 200 mOsm/kg. Threshold activation (10%) of black drum spermatozoa occurred at 370 mOsm/kg, and complete activation occurred at 580 mOsm/kg in HBSS. Sperm cells activated in artificial seawater had higher motility than those activated in HBSS at osmolalities from 350 to 500 mOsm/kg. Spermatozoa stored at 4 degrees C in HBSS or artificial seawater at osmolalities from 202 to 290 mOsm/kg retained motility longer than did those stored at other osmolalities Dilution rate had no effect on sperm storage time at 4 degrees C. Four chemicals were evaluated as cryoprotectants: dimethyl sulfoxide (DMSO), n,n-dimethyl acetamide (DMA), methanol, and glycerol. Glycerol and DMA at concentrations of 10% significantly reduced motility within 52 min. Spermatozoa were cryopreserved at 3 freezing rates (-27, -30, or -45 degrees C/min) in a nitrogen vapor shipping dewar or a computer-controlled freezer. Spermatozoa frozen using 10% DMSO had the highest post-thaw motility at a freezing rate of -27 or -30 degrees C/min. Spermatozoa frozen using 5% glycerol, 5% DMSO, or 10% DMSO had the highest post-thaw motility at a freezing rate of -45 degrees C/min.
已制定了关于黑鼓鱼精子采集、冷藏保存和冷冻保存的程序。通过取出并切片睾丸来采集精子样本，然后将精子悬浮于200 mOsm/kg的汉克斯平衡盐溶液（HBSS）中。黑鼓鱼精子的阈值激活（10%）发生在370 mOsm/kg，在HBSS中580 mOsm/kg时发生完全激活。在350至500 mOsm/kg的渗透压下，在人工海水中激活的精子细胞比在HBSS中激活的精子细胞具有更高的活力。精子在202至290 mOsm/kg渗透压的HBSS或人工海水中于4℃保存时，其活力保持时间比在其他渗透压下保存的精子更长。稀释率对4℃下精子的保存时间没有影响。评估了四种化学物质作为冷冻保护剂：二甲基亚砜（DMSO）、N,N - 二甲基乙酰胺（DMA）、甲醇和甘油。浓度为10%的甘油和DMA在52分钟内显著降低了精子活力。精子在氮气蒸汽运输杜瓦瓶或计算机控制的冰箱中以三种冷冻速率（-27、-30或-45℃/分钟）进行冷冻保存。使用10% DMSO冷冻的精子在-27或-30℃/分钟的冷冻速率下解冻后活力最高。使用5%甘油、5% DMSO或10% DMSO冷冻的精子在-45℃/分钟的冷冻速率下解冻后活力最高。
