Analyses of physiological factors regulating oocyte maturation and early development of a hermaphrodite pond snail Lymnaea stagnalis

(1)钉螺从受污染的死水转移到干净的曝气水中，可在3h内刺激其产卵。MALDI-MS分析表明，移植后30min，尾状背角细胞激素(CDCH)从大脑连合(COM)释放到血淋巴(HL)。COM背体粗提物(DB)能诱导卵子-消化腺复合体的体外配子释放(卵母细胞和精子)。(2)排出的卵母细胞很快在精管的远端出现、移动和受精。将DSO卵母细胞浸泡在蜗牛生理盐水、HL和蒸馏水中可诱导其减数分裂和卵裂，但在体内对其有严格的抑制作用，直至产卵。用卵膜+卵周液孵化的卵母细胞在低渗甘露醇溶液中可出现极体形成和分裂，而在高渗溶液中则不能。浸泡实验表明，大多数低相对分子质量的渗透物质直接进入囊膜，在胶体内渗透压作用下，卵周液始终保持高于HL的高渗态。这似乎是产卵前卵母细胞减数分裂不被允许的主要原因。(3)产卵螺用10^&lt；-3&gt；~10^&lt；-1&gt；Mβ-蜕皮激素溶液浸泡，卵囊体积略有增加。(4)从成熟钉螺生殖道提取液中未扩增出蜕皮激素受体的互补DNA。(5)高效液相色谱-酶免疫分析显示，HL中β-蜕皮激素水平在卵母细胞包装早期(蛋白腺体分泌期)达到峰值，随后迅速下降。α-蜕皮激素水平逐渐升高，并在产卵期前后达到峰值。因此，β-蜕皮激素可能通过卵黄周液的形成在阻止减数分裂中发挥重要作用。

(1) Snails could be stimulated to oviposit egg-mass within 3h by their transfer from polluted stagnant water into clean aerated one. MALDI-MS analysis revealed that caudo-dorsal cell hormone (CDCH) were released from cerebral commissure(COM) into haemolymph (HL) within 30 min after transfer. The crude extracts of COM with dorsal bodies (DB) could induce in vitro gamete-release(oocytes and sperm) from the dissected ovotesis-digestive gland complex.(2) Ovulated oocytes soon appeared, moved and fertilized in the distal part of spermoviduct (DSO). Meiosis and cleavage could be induced by immersing DSO oocytes in snail saline, HL and distilled water, while they were strictly inhibited in vivo till spawning. Isolated oocytes just spawned with capsule (egg membrane + perivitelline fluid) could show polar body formation and cleavage in hypotonic solution of mannitol but in hypertonic ones they could not. Immersion test showed that most osmolytes with low molecular weight immediately penetrate the capsule, which always keep perivitelline fluid hypertonic than HL by inner colloid osmotic pressure. This seems to be the major reason why oocyte meiosis can never be allowed before spawning.(3) Immersion of spawning snails in 10^<-3>〜10^<-1>M solution of β-ecdysone slightly increased the whole volume of egg capsule. Formation of perivitelline fluid may be stimulated by this steroid.(4) Complementary DNA of ecdysone receptor was not obtained by RT-PCR from the extract of the reproductive tract of mature snails.(5) HPLC-EIA demonstrated that β-ecdysone level in HL reached the peak at the early phase of oocyte-packaging (at the secretion of albumen gland) and rapidly decreased. Alpha-ecdysone level gradually increased and showed the peak around spawning time. Then β-ecdysone may play an important role in preventing meiosis through the formation of perivitelline fluid.