Analysis of flagellar movement of golden hamster spermatozoa by measuring sliding between outer doublet microtubules

1. 在金仓鼠精子中建立了“鞭毛模型”方法，即在MgATP^<2->激活下，脱膜精子显示出与完整精子相似的鞭毛运动。再激活液中Ca^<2+>的浓度改变了精子绕其长轴的滚动运动方向和精子在复盖下的偏转运动方向以及鞭毛波的振幅和不对称性。与海胆精子不同的是，大部分完好无损的金仓鼠精子从头部看时呈逆时针方向滚动。因此，金仓鼠精子细胞内Ca^<2+>的浓度可能与海胆精子不同。用DTT和胰蛋白酶处理脱膜精子，然后加入MgATP^<2->，在暗场显微镜下观察到外双线微管从轴突上滑动。这种滑动的速度，通过测量微管的长度和时间来确定，大约是8 μ m/s。介质中Ca^<2+>浓度对滑动速度的影响不大。睾丸与附睾头尾精子的滑动速度无显著差异。这些结果证实，这些精子中的运动器官已经在睾丸中功能组装，尽管这些精子在Tyrode溶液中是不运动的。

1. The method of "flagellar model", which is demembranated spermatozoa showing the flagellar movement similar to that of intact spermatozoa when activated with MgATP^<2->, has been developed in golden hamster spermatozoa. The direction of rolling motion of a spermatozoon about its long axis and of yawing motion of spermatozoa under the coverslip as well as the amplitude and asymmetry of flagellar wave was changed by the Ca^<2+> concentration in reactivation solution. In contrast to the sea urchin spermatozoa, the majority of intact golden hamster spermatozoa rolled counterclockwise when viewed from the head. Thus, intracellular concentration of Ca^<2+> of golden hamster spermatozoa is perhaps different from that of sea urchin spermatozoa.2. When the demembranated spermatozoa were treated with DTT and trypsin, and then MgATP^<2-> was added to them, sliding of the outer doublet microtubules from the axoneme was observed under a darkfield microscope. The velocity of this sliding, which was determined by measuring the length of microtubules and time, was approximately 8 um/s. The sliding velocity did not change by the Ca^<2+> concentration in the medium so much.3. There was no significant difference in the sliding velocity among testicular and caput and cauda epididymal spermatozoa. These results confirmed that the motor apparatus in these spermatozoa is already functionally assembled in the testis, although these spermatozoa are immotile in the Tyrode's solution.