Chaveiro A, Liu J, Mullen S, Woelders H, Critser J K
Institute for Animal Science and Health (ID-Lelystad), P.O. Box 65, 8200 AB Lelystad, The Netherlands.
Cryobiology. 2004 Feb;48(1):72-80. doi: 10.1016/j.cryobiol.2003.12.005.
The objective of this study was to determine the membrane permeability characteristics of bovine spermatozoa. These included the hydraulic conductivity (Lp), the permeability coefficients (Ps) of four common cryoprotective agents (CPAs) and the associated reflection coefficients (sigma). Stopped-flow fluorometry was applied in order to capture rapid cell volume changes under anisosmotic conditions in the absence or presence of permeant solutes (CPAs). This technique utilizes a concentration-dependent self-quenching entrapped fluorophore. The resulting cell volume changes were used in three-parameter fitting calculations to compute Lp in the absence of permeant solutes and Ps and Lp in the presence of permeating solutes (CPAs) at 22 degrees C. The hydraulic conductivity in the absence of permeating solutes was estimated to be 0.68+/-0.05 microm/min/atm (mean+/-SEM). Hydraulic conductivity (Lp) in the presence of CPAs was 0.91+/-0.27 (mean+/-SEM), 0.29+/-0.04, 0.42+/-0.05, and 0.39+/-0.03 microm/min/atm in the presence of dimethylsulfoxide (Me(2)SO), glycerol, propylene glycol (PG), and ethylene glycol (EG), respectively. The values for Ps were estimated to be 1.72+/-0.36, 1.75+/-0.03, 2.47+/-0.24, and 1.49+/-0.33 x 10(-3)cm/min for Me(2)SO, glycerol, PG, and EG, respectively. The data were used to simulate volume excursions during addition and removal of CPA, to predict the different effects of the four CPAs.
本研究的目的是确定牛精子的膜通透性特征。这些特征包括水力传导率（Lp）、四种常见冷冻保护剂（CPA）的渗透系数（Ps）以及相关的反射系数（sigma）。采用停流荧光法，以捕捉在等渗条件下不存在或存在渗透性溶质（CPA）时细胞体积的快速变化。该技术利用浓度依赖性自猝灭包埋荧光团。在22℃下，将所得细胞体积变化用于三参数拟合计算，以计算不存在渗透性溶质时的Lp以及存在渗透性溶质（CPA）时的Ps和Lp。不存在渗透性溶质时的水力传导率估计为0.68±0.05微米/分钟/大气压（平均值±标准误）。在存在二甲基亚砜（Me(2)SO）、甘油、丙二醇（PG）和乙二醇（EG）的情况下，CPA存在时的水力传导率（Lp）分别为0.91±0.27、0.29±0.04、0.42±0.05和0.39±0.03微米/分钟/大气压。Me(2)SO、甘油、PG和EG的Ps值估计分别为1.72±0.36、1.75±0.03、2.47±0.24和1.49±0.33×10(-3)厘米/分钟。这些数据用于模拟添加和去除CPA期间的体积变化，以预测四种CPA的不同效果。
