Hypothalamic GHRH and pituitary GH genes expression levels in neonatal inhibin-

immunoneutralized female rats

Al-Qadisiya UniversityAl-Qadisiya UniversityCollege of PharmacyCollege of PharmacyIRAQIRAQ

Prof. Jabbar A.A.Al-Sa'aidiProf. Jabbar A.A.Al-Sa'aidiBVMS, MS, PhD, Endocrinol. & Reprod. Physiol.BVMS, MS, PhD, Endocrinol. & Reprod. Physiol.

hypothalamic-pituitary-gonadal axis

Mammalian ovary (follicles) Inhibins & Activins Immunization against inhibin alpha subunit

Growth hormone & IGFs

IntroductionIntroduction

Ovarian GH receptors Is GH influence fertility

The composition of inhibins & The composition of inhibins & ActivinsActivins

Hypothesis of Hypothesis of studystudy

Inhibin immunoneutralization will

increase GH gene expression level in

pituitary gland along with the increment

of FSH secretion.

Aim of studyAim of study

The present study aims to investigate the role of passive immunization against

inhibin alpha in different postnatal stages of female rats until development of puberty on hypothalamic and pituitary

genes responsible for GH secretion as well as their reproductive performance

after puberty.

P-8 P-15 P-20 P-8 P-15 P-20 P-23 P-23 P-30 P-30 P-45 P-45 Arrival Arrival Antiserum/salineAntiserum/saline SacrificedSacrificed SacrificedSacrificed SacrificedSacrificed

schematic diagram showing time schematic diagram showing time course of treatment and sample course of treatment and sample

collections .collections .

Experimental designExperimental design

48 immature female rats (aged 15 48 immature female rats (aged 15 days)days)

Treated (No.24) injected Treated (No.24) injected with 1μg of inhibin α with 1μg of inhibin α subunit antiserum in subunit antiserum in

days 15, 16 , 17 ,and 20days 15, 16 , 17 ,and 20

Control (No.24) Control (No.24) injected with 100μl of injected with 100μl of normal saline in days normal saline in days 15, 16 , 17 , and 2015, 16 , 17 , and 20

Body weight of all rats throughout Body weight of all rats throughout the experimentthe experiment

TreatmenTreatment t

ControControl l

Group 1 Group 1 (No.8) (No.8) Sacrificed Sacrificed in day 23 in day 23 of age.of age. Group 3 Group 3

(No.8) (No.8) Sacrificed Sacrificed in day 45 in day 45 of age.of age.

Group 2 Group 2 (No.8) (No.8) Sacrificed Sacrificed in day 30 in day 30 of age.of age.

Group 2 Group 2 (No.8) (No.8) Sacrificed Sacrificed in day 30 in day 30 of age.of age.Group 3 Group 3

(No.8) (No.8) Sacrificed Sacrificed in day 45 in day 45 of age.of age.

Group 1 Group 1 (No.8) (No.8) Sacrificed Sacrificed in day 23 in day 23 of age.of age.

Experimental design (cont.)

After sacrification

Obtaining of pituitary and hypothalamus for detection of GH and

GHRH genes, respectively

GH, GHRH, and GABDH mRNA

analysis usingreal time RT-PCR

ResulResultsts

0

200

400

600

800

1000 ControlTreatm ent

Periods

RNA conc. (ng/ml)

PituitaPituitaryry

0

200

400

600

800

1000 ControlTreatment

Periods

RNA conc. (ng/ml)

HypothalamHypothalamusus

Total RNA concentration in Total RNA concentration in pituitary and hypothalamus tissues pituitary and hypothalamus tissues of control and inhibin-α immunized of control and inhibin-α immunized female rats at 23, 30, and 45 day female rats at 23, 30, and 45 day old.old.

** ** ** **

**

23 d. 23 d. oldold

30 d. 30 d. oldold

45 d. 45 d. oldold

Fold changes of pituitary Fold changes of pituitary GHGH of control female of control female rats (blue arrows) and female rats passively rats (blue arrows) and female rats passively immunized against inhibin-α (red arrows) at 23, immunized against inhibin-α (red arrows) at 23, 30, and 45 days old.30, and 45 days old.

Fold changes of hypothalamus Fold changes of hypothalamus GHRHGHRH of control of control female rats (blue arrows) and female rats female rats (blue arrows) and female rats passively immunized against inhibin-α (red passively immunized against inhibin-α (red arrows) at 23, 30, and 45 days old.arrows) at 23, 30, and 45 days old.

23 d. 23 d. oldold

30 d. 30 d. oldold

45 d. 45 d. oldold

0

1

2

3

4

5 ControlTreatm ent

Hypothalam us

G roups

Fold ch

ange

s of

GHR

H gen

e

0246101214161820

ControlTreatm ent

Pituitary

G roups

Fold cha

nge ofGH gen

e

Fold changes of hypothalamus Fold changes of hypothalamus GHRHGHRH and pituitary and pituitary GHGH of control female rats (blue arrows) and female of control female rats (blue arrows) and female rats passively immunized against inhibin-α (red rats passively immunized against inhibin-α (red arrows) at 23, 30, and 45 days old.arrows) at 23, 30, and 45 days old.

ConclusiConclusionsonsIn conclusion, the present over expression In conclusion, the present over expression

of hypothalamus of hypothalamus GHRHGHRH and pituitary and pituitary GHGH genes genes may indicates a significant potency of pre-may indicates a significant potency of pre-

pubertal immune-neutralization of endogenous pubertal immune-neutralization of endogenous inhibin on female reproductive function. inhibin on female reproductive function.

This finding may suggest the strong This finding may suggest the strong correlation between correlation between GHRHGHRH and and GHGH genes genes expression levels and ovary growth and expression levels and ovary growth and

development.development.

This elucidation may lead to the development This elucidation may lead to the development of novel strategies aimed to support of novel strategies aimed to support

reproduction and its future performance, and reproduction and its future performance, and may play a significant potent role in may play a significant potent role in

reproduction applications.reproduction applications.

Thank YouThank You