日本鰻人工催熟之研究

Abstract

為了達到日本鰻（Anguilla japonica）完全養殖之規模，相關研究雖已建立利用 Salmon Pituitary Homogenate ( SPH ）及 17 α， 20β-dihydroxy-4-pregnen-3-one ( DHP）進行人工催熟方面之研究，但仍有誘導效果不易提升、催熟週數分散、催熟時機不易掌握、種鰻催熟後自然產卵易影響卵質，以及操作成本較高等瓶頸待突破。養殖日本鰻由於須先進行誘導卵黃蓄積後，方可進一步催熟，所以本實驗根據王 ( 1999 ) ，使用 SPH 或併用甲基睪固酮（MT）的方法作為前處理後，進一步人工催熟並達產卵階段，並藉由離體培養，加以探討卵細胞最後成熟過程中，外源性 Progesterone 之效果及作用機制。 養殖3年以上之日本鰻（體重 504 士 84 公克）經個別標識後,隨機分成 SPH 或與 MT(17a-methyltestosterone）併用6週（SPH + MT6）以及全程併用（SPH + MT）三處理組，於海水20土1℃馴化兩週後，每週注射乙次作為誘導卵黃蓄積前處理。實驗結果，在 7-12 週的前處理後若種魚當週體重上升比（BWI）大於10 % ，且48小時後持續上升超過20% ，便可進行 SPH priming ，並於24 小時後施打DHP或OHP(17a- hydroxyprogesterone）進行人工催熟。而 BWI 接近但小於10％者則需進行抽卵鏡檢，若其卵組成中，透明吸水卵佔 75 ％以上而油球開始融合，則亦可進行人工催熟。 SPH 、 SPH+MT6、 SPH+MT 、對照組，分別可誘導 67 . 6 士 7.2 ％、90 . 8 士6.5 ％、 83 . 7 士 89 ％、 0 ％的種鰻進入卵黃蓄積，以及誘導 34 . 4 士 6 . 2 ％、 42.5 士 15.4 ％、56.4士35.9 ％、0％的種鰻完成卵黃蓄積。顯示雄性素的持續升高會延遲但不會抑制卵細胞進入吸水成熟階段。DHP及 OHP 皆可100％誘導排卵發生，但種鰻產卵時間， DHP 及 OHP 組分別為排卵後2-7 小時及 1-4 小時，以DHP催熟顯然會使排卵及產卵的間距上較長，若使種鰻自然產卵易造成卵質良窳參差不齊，因此魚卵排出後須馬上進行人工授精。於離體培養發現誘導 GVBD 的能力 DHP : 85.6士1. 9％、 OHP : 51.1 士 5.1 ％、20β-S: 58.9士3.9％及對照組:0%,而 DHP係透過轉錄及轉譯之層面來產生誘導卵細胞GVBD的效果。 由以上結果，人工催熟過程中若使用 SPH 併用6週MT ，可在 12 週的試驗時間內，誘導 90 ％的種鰻進行卵黃蓄積，並使42％的種鰻完成卵黃蓄積達吸水階段，吸水之種鰻並可在催熟後 100％進行排產卵。而 OHP可較 DHP 縮短排卵到產卵之間距，若使種鰻自然產卵亦不致於明顯影響卵質，有助於維持授精率及孵化率，並避免人工擠卵中造成的緊迫。由以上的技術改進，期望能使既有的人工催熟操作效率更高，更具便利性、經濟性及可預期性。

The cultivated Japanese eel (Anguilla japonica) is usually immature in reproduction, it cannot proceed to accomplish final maturation without first inducing the vitellogenesis. Artificial induction of maturation in Japanese eel has been performed by serial injections of Salmon Pituitary Homogenate (SPH) and DHP, yet still not so effectively. The induction rate of vitellogenesis is not more than 50% by SPH treatment, and eggs with good quality cannot always be spontaneously obtained finally by DHP treatment. Therefore, this experiment was designed to follow Wang (1999), using SPH cotreated with exogenous steroids to improve the induction rate of vitellogenesis, then induced eels to the final maturation. The Japanese eel (504±84 grams) were tagged and separated into three groups randomly: SPH treatment and SPH cotreated with MT (for 6 weeks only or weekly injection). The injections were performed once a week to induce vitellogenesis. In addition to in vivo study, this study also examined the effect of DHP and OHP in final maturation in vitro. Result showed that SPH cotreated with MT for 6 weeks could induce more eels to start vitellogenesis than those of SPH alone after 7—8 weeks of the treatments. However, the existence of MT during late vitellogenesis stage would delay but cannot inhibit the hydration to occur. The eels with hydrated oocytes can proceed with induction of final maturation by DHP or OHP. In morphology examination, the intra-ovarian oocytes were transparent and with oil droplets centrally located under a binocular microscope. It was suitable for those oocytes to proceed with induction of final maturation. All of the eels with hydrated oocytes could be induced to GVBD and ovulate at l4-20 hrs after DHP or OHP injection. The eels oviposied at 2-7 hrs after ovulation induced by DHP. In contrast, the eels oviposited within 4 hrs after ovulation by OHP. As the quality of eggs retained in body cavity decreased rapidly after ovulation. The time interval between ovulation and oviposition in OHP group were obviously shorter than in DHP group. Therefore, better eggs can be obtained if the spawner is induced to oviposite spontaneously by OHP.