Efficient oxytocic for hippocampus in late pregnancy and oxytocic method of efficient oxytocic

Abstract

The invention discloses an efficient oxytocic for hippocampus in the late pregnancy period and an oxytocic method of the efficient oxytocic. Research verifies that the odinagogue taking prostaglandin E2 as a main active component can rapidly induce the hippocampus to give birth in the late pregnancy (take effect within 60 minutes after odinagogue), and the odinagogue time is remarkably shortened. The juvenile fish produced by the method has independent viability, has no significant adverse effect on the sea horse with incomplete embryonic development, can effectively improve the mature fry acquisition rate when the embryonic development condition is unknown, reduces the fry loss in the culture process, and is beneficial to sea horse resource protection and industrial layout optimization.

Description

Technical Field

[0001] The present invention relates to the field of biotechnology, and in particular to a highly effective oxytocin precisely targeting the hippocampus in late pregnancy and a method for inducing oxytocin. Background Art

[0002] Seahorses (Hippocampus) are a valuable marine fish used for both medicinal and edible purposes, possessing extremely high economic value. However, due to factors such as overfishing and ecological changes, seahorse resources in natural waters are nearly depleted.

[0003] A major factor limiting my country's seahorse supply is bottlenecks in artificial breeding technology, particularly the high loss rate of fry. Furthermore, seahorses typically give birth at dawn. In high-density breeding environments, if fry are not promptly harvested and individually reared in the early morning, they are easily devoured by adult seahorses, further exacerbating fry losses. Therefore, developing an effective method for centralized seahorse spawning is crucial to improving fry recovery rates.

[0004] Currently, Arginine vasotocin (AVT) is the only hormone known to effectively stimulate pregnant seahorses to give birth in a relatively short period of time, and its biological effects are quite significant. However, this hormone has the following defects: on the one hand, its mechanism of action is unrelated to the stage of embryonic development. If used when the embryo is immature, it may induce birth prematurely, causing the expulsion of incompletely developed embryos and resulting in the loss of effective seedlings. In conventional breeding environments, the use of such non-selective oxytocin is risky due to the difficulty in real-time monitoring of the embryonic development status in the male seahorse's brood pouch. On the other hand, the time required for induction of labor is still relatively long. There is a 6-hour window from the induction treatment to the effectiveness of the induction, which is not conducive to the time arrangement of the breeders and also increases the risk of seedling loss. In view of this, there is an urgent need for an oxytocin and induction method that is selective and more efficient for seahorse embryos. Summary of the Invention

[0005] In view of the shortcomings of existing hippocampal induced labor technology, especially the lack of a special oxytocin that can accurately match embryo maturity and has a highly effective induced labor effect, the present invention aims to provide a highly effective oxytocin for hippocampus in late pregnancy and a corresponding induced labor method.

[0006] In a first aspect, the present invention provides the use of prostaglandin E2 in the preparation of a highly effective oxytocin for the hippocampus in late pregnancy.

[0007] Preferably, in the above application, the preparation of the highly effective oxytocic agent comprises the following steps: dissolving prostaglandin E2 in acetone, and then adding NaCl solution for dilution to obtain the product.

[0008] Preferably, in the above application, the concentration of the NaCl solution is 0.65% by mass.

[0009] Preferably, in the above application, the hippocampus includes hippocampus striatus.

[0010] Preferably, in the above application, the dosage of prostaglandin E2 is 1 μg prostaglandin E2 per gram of hippocampal body weight.

[0011] In a second aspect, the present invention provides a highly effective method for inducing labor in late-pregnancy hippocampus, comprising the following steps:

[0012] (1) Preparation of an oxytocic agent: dissolve prostaglandin E2 in acetone and dilute with 0.65% NaCl solution;

[0013] (2) Remove the pregnant seahorse from the temporary holding tank and gently move it to a wet towel. Use a 1 mL sterile medical syringe to uniformly inject the oxytocin into the softer area of ​​the abdomen above the seahorse's cloaca, with the needle inserted to a depth of 2 to 3 mm.

[0014] Preferably, in the above method, the injection volume of the oxytocic agent is 100 μL per hippocampus.

[0015] Preferably, in the above method, the dosage of the oxytocic agent is 1 μg of prostaglandin E2 per gram of hippocampal body weight.

[0016] Preferably, in the above method, the hippocampus includes hippocampus striatum.

[0017] Male seahorses incubate embryos in their closed pouches, a characteristic that makes it difficult to track the development of embryos within individual seahorse pouches during captivity. The only current seahorse oxytocin, oxytocin, is not selective for embryonic developmental stage, which can lead to premature expulsion of immature fry. Furthermore, the time window from induction treatment until induced fertilization is effective is relatively long.

[0018] The present invention has developed an efficient oxytocin and its oxytocin method that is precisely targeted at late-pregnant seahorses. With prostaglandin E2 as the main active ingredient, it can quickly induce late-pregnant seahorses to give birth (effective within 60 minutes after induced labor), significantly shortening the induced labor time. The fry produced by this method have the ability to survive independently, have no significant adverse effects on seahorses with incomplete embryonic development, and can effectively increase the acquisition rate of mature seedlings when the embryonic development status is unclear, reducing seedling losses during the breeding process. This technology, disclosed for the first time in the present invention, provides a new solution for seahorse breeding, which is not only efficient in operation, but also contributes to the protection of seahorse resources and the optimization of the breeding industry. BRIEF DESCRIPTION OF THE DRAWINGS

[0019] Figure 1These are seahorse fry induced by oxytocin.

[0020] Figure 2 Schematic diagram of the induced labor experiment. DETAILED DESCRIPTION

[0021] The following examples are provided to further illustrate the present invention, but are not intended to limit the present invention.

[0022] Unless otherwise specified, the experimental methods used in the following examples are conventional methods, and the reagents and materials used are all commercially available unless otherwise specified.

[0023] In the following examples, the 0.65% NaCl solution refers to a NaCl solution with a mass fraction of 0.65%.

[0024] Example 1

[0025] Taking Hippocampus erectus as an example, the induction effect of the present invention is described through a control experiment. The specific implementation scheme is as follows:

[0026] (1) Preparation of materials for induced labor test

[0027] (1) Reagents

[0028] Prostaglandin E2 (PGE2) dry powder 1 mg; acetone 1 mL; 0.65% NaCl solution 50 mL.

[0029] (2) Equipment

[0030] 4 fish tanks; several 1mL sterile medical syringes; absorbent paper; clean towels.

[0031] (3) Experimental fish

[0032] There are 53 healthy seahorses with swollen abdomens and similar body size and weight.

[0033] (2) Experimental steps for inducing labor in pregnant seahorses

[0034] (1) Prepare four marine fish tanks one day in advance and divide them into a control group and an induced spawning group. Each group includes a temporary holding tank and an induced spawning tank. Keep the environmental conditions of all fish tanks consistent, with the water temperature controlled at 24-28°C and the salinity maintained at 28-32‰. Ensure a gentle water flow. Wrap gauze around the water filter of the induced spawning tank to prevent the fry from being sucked away by the water flow.

[0035] (2) The seahorses were wiped clean of seawater with absorbent paper and weighed. The seahorses were randomly placed in a temporary holding tank in the control group (18) and the induced fertilization group (35). The developmental stage of the embryos in the pouch of all seahorses was unknown before dissection.

[0036] (3) Add 100 μL of acetone to 1 mg of PGE2 dry powder. Dissolve thoroughly, then add 900 μL of 0.65% NaCl solution to prepare a 1 mg / mL oxytocic solution. This preparation should be performed on ice to ensure the stability and effectiveness of PGE2.

[0037] (4) Based on the average weight of the hippocampus to be induced to labor, a dose of 1 μg PGE2 per gram of body weight is administered. The oxytocic solution is mixed with a 0.65% NaCl solution to prepare an induced labor working solution. The preparation process must be carried out on ice, and the completed working solution should also be kept on ice. The volume of working solution injected into each hippocampus is fixed at 100 μL. The required volumes of oxytocic solution and 0.65% NaCl solution are calculated according to the following formula:

[0038] V 原液 =m×n(μL),V NaCl =100n-V 原液

[0039] Where, m: average weight of hippocampus to be induced (unit: g); n: total number of hippocampus to be induced (unit: tail); V 原液 : Volume of oxytocic solution (unit: μL); V NaCl : Volume of 0.65% NaCl solution (unit: μL).

[0040] The preparation process must be carried out on ice, and the prepared working solution should also be stored on ice.

[0041] In this experiment, the average weight of hippocampus in the oxytocin group was about 10 g, with a decimal of 35. The required volume of oxytocin stock solution was calculated to be 350 μL, and the volume of 0.65% NaCl solution was 3150 μL. After mixing the two, 3500 μL of oxytocin working solution was obtained.

[0042] In this step, since the oxytocin may be easily lost during the induction process, it is recommended to reserve enough for 2 to 3 seahorses. When calculating, replace n in the formula with the actual number of seahorses prepared.

[0043] (5) 18 mL of 0.65% NaCl solution containing 1% acetone was prepared according to the volume ratio of acetone to 0.65% NaCl solution = 1:99, and used as the injection for the control group.

[0044] (6) Gently transfer the seahorses from the holding tank to a moist towel. Using a 1 mL sterile medical syringe, uniformly inject 100 μL of solution into the softer area of ​​the seahorse's abdomen above the cloaca, with the needle penetrating approximately 2 to 3 mm. The control group's seahorses were injected with a 0.65% NaCl solution containing 1% acetone, while the oxytocin working solution was injected into the seahorses in the oxytocin group. After the injections were completed, the seahorses were immediately placed in the corresponding oxytocin tanks.

[0045] (7) Within 60 minutes after injection, remove any seahorses that exhibit calving behavior and place them back in a temporary holding tank for statistical analysis and dissection, and to record the embryonic development in their pouches.

[0046] (8) Remove the juvenile fish and observe and evaluate their development using a stereomicroscope.

[0047] (9) All experimental seahorses were dissected, the embryonic development in the pouch was counted, and the effects of oxytocin on seahorses at different gestational stages were analyzed.

[0048] (3) Results

[0049] 1. The anatomical evaluation of the seahorses used in the experiment was carried out and they were divided into the following four categories based on the morphological characteristics of the embryonic development in the pouch:

[0050] Table 1: Criteria for the classification of pregnant hippocampus

[0051]

[0052] 2. The number of seahorse tails at different stages of pregnancy in the control and induced labor groups was counted, as well as the number of tails that showed signs of giving birth after injection of the solution (see Table 2). The results showed that none of the seahorses in the control group showed signs of giving birth, indicating that needle puncture and intraperitoneal injection of the control solution did not induce male seahorses to give birth. In the induced labor group, seahorses in the non-pregnant and early pregnant stages did not respond to the oxytocin, indicating that the oxytocin did not induce seahorses in these two stages to give birth; only 2 of the 9 mid-pregnant seahorses showed signs of giving birth, while all 6 late-pregnant seahorses gave birth under the action of the oxytocin.

[0053] Table 2: Statistics of hippocampal tails showing farrowing behavior within 60 minutes after injection

[0054]

[0055] 3. Observe the development of the seahorse seedlings collected in step (8) under a stereomicroscope (see Figure 1 The results showed that the snout, tail, dorsal and anal fins of the induced seedlings were fully formed, with pigment spots visible on the body surface. The yolk of most seedlings had completely disappeared, indicating that their overall development was complete.

[0056] In summary, the oxytocin and oxytocin method of the present invention can effectively induce pregnant seahorses to give birth, and the seahorses will show puerperal behavior within 60 minutes after treatment. And the oxytocin effect is closely related to the stage of embryonic development: it is effective on late-pregnant seahorses, effective on some mid-pregnant seahorses, and has no obvious effect on non-pregnant and early-pregnant seahorses. Microscopic observation showed that the vast majority of induced puerperal seedlings are fully developed and have the ability to live freely. This shows that the present invention selectively and quickly induces late-pregnant seahorses to give birth and produces mature seedlings, providing a reliable technical reference for artificial induced puerperal birth in seahorses.

[0057] Example 2

[0058] Referring to the method for inducing labor in Example 1, prostaglandin F2a was used instead of prostaglandin E2 to observe the effect of prostaglandin F2a. The results showed that prostaglandin F2a had no inducing effect on hippocampus at different pregnancy stages. The results are shown in Table 3.

[0059] Table 3: Statistics of hippocampal tails showing farrowing behavior within 60 minutes after injection

[0060]

[0061] The above detailed description is a specific description of the possible embodiments of the present invention. The embodiment is not intended to limit the patent scope of the present invention. Any equivalent implementation or modification that does not depart from the present invention should be included in the patent scope of this case.

Claims

1. Application of prostaglandin E2 in the preparation of a highly effective oxytocin for the hippocampus in late pregnancy.

2. The use according to claim 1, characterized in that The preparation of the high-efficiency oxytocic agent comprises the following steps: dissolving prostaglandin E2 with acetone, and then adding NaCl solution for dilution.

3. The use according to claim 2, characterized in that The concentration of the NaCl solution is 0.65% by mass.

4. The use according to claim 2, characterized in that The seahorse includes the hippocampus linearis.

5. The use according to claim 1, characterized in that The dosage of prostaglandin E2 is 1 μg of prostaglandin E2 per gram of hippocampal body weight.

6. An efficient method for inducing labor in late-pregnancy hippocampus, characterized in that: The following steps are involved: (1) Preparation of an oxytocic agent: dissolve prostaglandin E2 in acetone and dilute with 0.65% NaCl solution; (2) Remove the pregnant seahorse from the temporary holding tank and gently move it to a wet towel. Use a 1 mL sterile medical syringe to uniformly inject the oxytocin into the softer area of ​​the abdomen above the seahorse's cloaca, with the needle inserted to a depth of 2 to 3 mm.

7. The method according to claim 6, characterized in that The injection volume of the oxytocic agent was 100 μL per hippocampus.

8. The method according to claim 6, characterized in that The dosage of the oxytocic agent is 1 μg of prostaglandin E2 per gram of hippocampal body weight.

9. The method according to any one of claims 6 to 8, characterized in that: The seahorse includes the hippocampus linearis.

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