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Different Types of Inducing Agents and their Structural & Functional

Pathway

Prepared by

Sachin sahu

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Introduction

• Argentina 1930 B. A. Houssay is the 1st start from the carp pituitary extract as a inducing agent.

• 1934 Von Ihering et al., of brazil succeeded through administration of pituitary gland extract.

• In India the 1st attempt to induce C. mrigala to spawn by the injection of mammalian pituitary extract was done by H. Khan(1937) later H. Chaudhuri(1955).

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Collection & preservation & preparation of Pituitary gland extract

Collection • Dissection and removing a portion of scalp• Through foramen magnum

• Preservation• Absolute alcohol• Acetone Freezing

• Extract preparationHomogenigation Centrifugation

• Storage of extract• In water & Glycerine• In propylene glycol

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DOSE OF PITUITARY GLAND EXTRACT

• Female:- Initially dose 2-3mg/kg body weight

After 5-6 hours • female :-2nd dose is – 10-12mg/kg • Male :- 4-6 mg/kg body weight

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Composition of pituitary extract • Pituitary gland contain gonadotropins hormone.• GtH I (FSH)• GtH II(LH)

• All gonadotropins are Glycoproteins in nature

• Ganarly prefer mature Common carp, Tilapia due to there availability is around year.

• Two types of Gonadotropins» GtH I - Vitellogenesis» GtH II Oocyte maturation, ovulation/spermiation and spawning

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GtH IProduced during Vitellogenic phase

GtH I

Thecal and Granulosa cell of ovarian follice

Estradiol

Estradiol released in blood

Estradiol acts on liver to produce Vitellogenin

Vitellogenin carried to Oocytes through circulation and incorporated into cytoplasm of Oocytes

Vitellogenin converted into Yolk Protein - Lipovitellin and Phosvitin

Diagrammatic representation of the follicle and oocyte during early vitellogenesis

Nagahama, 1983

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Gth II• Secreted during Post-Vitellogenic Phase

Gth II

Follice cells

Maturation Inducing Hormone (MIH) or 17 alpha- Progesterone

MIH stimulates the production of Maturation Promoting Factor (MPF) in the cytoplasm of oocytes

Finally Oocyte is considered as mature

1st Dose

Rupture of Oocyte envelope and release of Mature Ova -Ovulation2st Dose

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Central nervous system

HYPOTHALAMUS

GnRHInject Pituitary gland

extract

Hypophysis

Gonadotrophs

Ooplasm Oocyte envelope

Vitellogenesis

Gravidoocyte Estradiol

Oocyte envelopes

ooplasm

Mih

Mpf 1st mat div

Germinal vesicle

Mature oocyte

Vitellogenic oocyte

Ovary

GthI GthII

Dopamine

Vitellogenin

liver

Sperm

ovum

Zygote nucleus

Zygote

Embryonic devlopment

Hatching

Post vitellogenic oocyte

Ovulation

Pathway of pituitary extract

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PROBLEMS IN PITUITARY EXTRACT

Non-availability of good quality of pituitary gland (P.G).

Inadequate supply of pituitary gland.

High cost Mixture of hormones may have side effects on gametogenesis and other

function.

Variability in Pituitary Gonadotropin potency Cumbersome procedure of preparation of extract

Short shelf life (Extract can be stored only for 6-12 months)

Less Percentage of spawning success

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Synthetic compound

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OVAPRIM • Ovaprim is a potent ovulating/spermiating agent used to promote

and facilitate reproduction of many species of fish.•   • Manufactured by syndel laboratories in Canada and marketed in

India by agri.vet farma of Glaxo India Ltd.,Bombay.

• Composition :-• 1) 20 μg/ml Salmon gonadotropin releasing hormone (sGnRH) • 2) domperidone 10mg/ml Ref :- (2015 Syndel Asia Sdn Bhd)

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Species MALE (ml/kg) Female (ml/kg

Catla 0.10-0.15 0.4-0.5

Rohu 0.10-0.15 0.35

Mrigal 0.10-0.15 0.25

Silver carp 0.15-0.20 0.40

Grass carp 0.15-0.20 0.60

GnRH - StructureDomperidone structure

Doses of ovaprimIt can be administered in a single dose to brooders.

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Ovaprim is used to:• Induce maturation within a spawning season.

• Advance spawning date

• Coordinate and synchronize spawning times

• Increase milt production including increased sperm count

• Induce maturation in difficult species

• Induce spawning in the most important and difficult to spawn species

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Working principle of ovaprim

• When injected into the body cavity or muscle of receptive, mature, conditioned fish,

• The synthetic GnRH travels from the injection site through the blood to activation sites in the pituitary gland.

• Ovaprim initiates the reproductive cascade and eliminates the need for a natural trigger.

• Domperidone, the other active component of Ovaprim, helps block the inhibitory effects of dopamine.

• Domperidone, therefore, is very important for induced spawning of species for which the reproductive cascade would be stopped due to stressors that lead to dopamine release, because dopamine will block GnRH activity.

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Functional pathway of ovaprim

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Advantages • Ready to use liquid, consistent potency and reliable results,

• Long shelf life, and can be stored at room temperature,

• Formulated to prevent over dosing,

Male and female can be injected only once simultaneously,

• Reduces handling and post breeding mortality,

• High percentage of eggs

• Fertilization and hatching.

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OVATIDE • Ovatide is an indigenous, cost-effective and new hormonal

formulation for induced breeding of fishes.

• Combined of GnRH analogue with dopamine antagonist pimozide

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Doses of Ovatide

Species MALE (ml/kg) Female (ml/kg

Catla 0.20-0.30 0.40-0.50Rohu 0.10-0.20 0.20-0.40Mrigal 0.10-0.20 0.20-0.40Silver carp 0.20-0.25 0.40-0.50

Grass carp 0.20-0.25 0.40-0.50

By:- Hemmo Pharmaceuticals

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The Advantages of Ovatide: • It is cost-effective hormonal preparation.• It gives high fertilisation and hatching percentage (85-95%).• It is increases egg production through complete spawning.• It produces healthy seed.• It is easy to inject due to its low viscosity.• It does not cause adverse effects on brood fish after injection.• It can be administered in a single dose to brooders.• It can be stored at room temperature.• It is quite effective even under climatic adversities and ovatide

is available in the market as 10 ml vial, which costs Rs. 300.

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Wova-FH• Composition:• Synthetic Gonadotropin Releasing Hormone

Analogue (SGnRH).• Developed by Wockhardt, one of the largest

pharmaceutical companies state of the art Research and Development Centre, in Mumbai India.

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Advantage and Dose Advantages:• High fertilization & hatching rate. • Single dose administration.• No stress to the broodstock.• Easy injectability.• Cost effective.• Stable at room temperature• long shelf life.• Easily available and ready to use.• Used In Following Countries:• India• Iran

Species Sex ml/kg body weight

Rohu Male 0.10 - 0.30 Female 0.30 - 0.50Catla Male 0.10 - 0.30 Female 0.30 - 0.50Mrigal Male 0.10 - 0.30 Female 0.30 - 0.50Grass carp Male 0.10 - 0.30 Female 0.40 - 0.80Silver carp Male 0.10 - 0.30 Female 0.40 - 0.80Cat Fishes Male 0.20 - 0.40 Female 0.60 - 0.90

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OVAPEL • Ovapel developed by university of Godollo in Hungary, Combined of

mammalian GnRH analogue and dopamine receptor antagonist, Lactosum,

• Recommended dose 1-2 pellet/kg of fish in Catla and Rohu.

• Use• Weight of brood stock should be measured before propagation. • The required amount of ovapel is 1 pellet for 1 kg of brood stock .• When, fish are ready for spawning, pellet should be powdered and

dissolved in salt solution (0.65% NaCl) 12 hour before planned ovulation then injected in to the abdomen fish.

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Structure of ovapel

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Puberogen• Puberogen consists of 63% follicle stimulating hormone (FSH)

and 34% Leutinizing hormone (LH).

Dosage

• Female • 1st 50–100 IU/kg of fish. • 2nd 100-200 IU/kg (48 hours after the first injection)• Male 20–50 IU/kg of fish.• The fish will normally spawn within 12–15 hours after the

second injection.

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Reference

1. Hoar W.S. ,Fish physiology(valume-9B)(2014), ;Induced final maturation , ovulation and Spermiation in cultured fish.pp-351-352

2. Sharad R Surnar, AD Kamble, NS Walse, OP Sharma, VP Saini, ; (2015), Hormone administration with induced spawning of Indian major carp, International Journal of Fisheries and Aquatic Studies.

3. Sharma charula (2008) Assessment of Syn"ergistic Efficacy of Pituitary Extract with Ovaprim and Ovatide on Breeding Perforrnance and Survival of Spawn of Cirrhinus mrigaJa (Ham)

4. L.B. Tiro, Jr. B.J. Pudadera, Jr., and I.O. Potesta. ; Biology and culture of sea bass (Lates calcarifer) Aquaculture Department, Southeast Asian Fisheries Development Center network of aquaculture centres in asia.

5. S. Chantarasri, Hanung Santosa, Hardoto and Sumbodo Kresno Yuwono, induced spawning and larval rearing of seabass (lates calcarifer bloch) in captivity

6. www.Wockhardt.com7. MahapatraB.K., (2004) Conservation of the Asiatic catfish, Clarias batrachus through artificial

propagation and larval rearing technique in India8. U. K. Sarkar1, R. S. Negi, P. K. Deepak, S.P. Singh, S.M. Srivastava and Dipak Roy*, Captive

breeding of vulnerable Indian carp Cirrhinus reba with Ovaprim for conservation of wild populations

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Thank you