DETECTION OF MESSENGER-RNA FOR INHIBIN ALPHA-SUBUNITS AND BETA(A)-SUBUNITS IN BOVINE OVARIAN TISSUES AND THE EFFECT OF IN-VIVO ADMINISTRATION OF GNRH

The aims of these studies were to determine which types of bovine ovarian tissue contain mRNA for inhibin/activin subunits and whether administration of GnRH influences concentration of these mRNAs. In experiment (exp.) one, cows in the luteal phase of the estrous cycle were given prostaglandin F2alpha (PGF2alpha) to induce luteal regression and injected after 40 hr with saline (n=5) or 100 mug GnRH (n=6). Ovaries were removed 6 hr later. In exp. two, unilaterally ovariectomized (OVX) heifers (n=33) in the luteal phase of their estrous cycle were given PGF2alpha to induce luteal regression. Twelve heifers were OVX without injection of GnRH at 24 (n=6) or 40 hr (n=6) after PGF2alpha. The remaining heifers (n=21) were given 100mug GnRH at 40 hr after PGF2alpha injection and OVX 8 (n=4), 16 (n=5), 24 (n=6) or 48 (n=6) hr after GnRH injection. Total cellular RNA was isolated from large follicles (exp. one and two), small-medium follicles and stromal tissue (SMS) and corpora lutea (CL; exp. one) tissues and analyzed by dot blot and Northern blot techniques by hybridizing with cDNA probes for human inhibin/activin alpha- and beta(A)-subunits. Large follicles were classified as steroidogenically active (EA) if follicular fluid (FF) concentration of estradiol-17beta (E2) was greater than progesterone (P4), or if P4 and E2 concentrations in FF were greater than 100ng/ml, and estrogen inactive (EI) if FF concentration of E2 and P4 did not satisfy these criteria. In exp. one, mRNA for the alpha-subunit was primarily expressed in EA follicles, and detectable in EI follicles, SMS, and CL while beta(A)-subunit mRNA was detected only in large EA follicles and a few SMS samples. The mRNA (x +/- SEM fmoles/mg DNA) for both subunits of inhibin/activin was higher (P<.05) in EA follicles from GnRH-treated cows (alpha = 210.2 +/- 38.6; beta(A) = 376.9 +/- 41.0) than in EA follicles from control cows (alpha = 102.5 +/- 28.6; beta(A) = 170.8 +/- 57.6). Concentration of mRNA for the alpha-subunit of inhibin in other ovarian tissues was not different (P>.10) between saline and GnRH treatments. In exp. two, the mRNA (fmol/mg DNA +/- SEM) for the alpha-subunit decreased slightly 24 to 40 hr after PGF2a injection from 112 +/- 41 to 84 +/- 15, increased (P<.05) transientally to 158 +/- 24 at 8 hr after GnRH, and declined (P<.01) to 55 +/- 16, 40 +/- 4 and 42 +/- 14 at 16, 24 or 48 hr after GnRH, respectively. Similarly, the beta(A)-subunit decreased slightly from 242 +/- 52 to 189 +/-48 between 24 to 40 hr after PGF2alpha. Following injection of GnRH mean concentrations decreased (P<.10) further from 15 5 +/- 41 to 53 +/- 24, 64 +/- 33 or 10 +/- 7 at 8, 16, 24 or 48 hr, respectively. In summary, mRNA for the a-subunit of inhibin shows a transient rise in response to the preovulatory gonadotropin surges, but declines rapidly (3 fold decrease in 8 hr) following the transient rise and remains at this decreased level through ovulation. The mRNA for the beta(A)-subunit of inhibin/activin rises (exp. one) or remains at a steady state (exp. two) early after the preovulatory gonadotropin surges and then declines by 16 hr post GnRH injection and remains low through ovulation.