MOLECULAR AND CELLULAR NEUROSCIENCRS 3,171-178 (19%)

LH and FSH Subunit mRNA Concentrations during the Progesterone- Induced Gonadotropin Surge in Ovariectomized

Estrogen-Primed Immature Rats DARRELL BRANN, JAMES O’CONNER, MARLENE WADE, AND VIRENDRA MAHESH

Department of Physiology and Endocrinology, Medical CoUege of Georgia, Augusta, Georgin 30912

Received for publication October 24, 1991

Progesterone is able to bring about an LH and FSH surge in the estrogen-primed ovariectomixed rat while dexamethasone brings about selective FSH release. The purpose of this study was to determine if progesterone- and dexametbasone-induced gonadotropin secretion is accompanied by changes in LH@ and FSH@ mRNA levels. Gonadotropin a-subunit, LII&subunit, and FSH&subunit mRNA levels in the pituitary of ovariectomixed rats were suppressed by estrogen treatment and dexamethasone brought about a significant increase in FM&!? mRNA within 1 h. Progesterone treatment (0900 h) led to a surge in serum LH levels, with peak values at 1400 h. LIfa mRNA levels were slightly elevated by progesterone at 1400 h. However, an elevation of LH@ at 1400 h was also observed in the dexamethasone group which did not show an increase in serum LH. Serum FSH levels were elevated at 1400 and 1600 h in the progesterone group and at 1600 h in the dexamethasone group after an initial fall at 1000 h. No correlation was observed between in- creases in serum FSH during these times with FSI&3 mRNA levels. In conclusion, the ability of progesterone to induce LH and FSH surges in the estrogen-primed ovariectomised rat was not associated with any clear correlative changes in the mRNAs for these hormones. On the other hand, dexamethasone did increase FSW mRNA levels prior to elevating serum levels of FSH. Nevertheless, as a whole, steroid effects on the temporal secretory pattern of LH and/or FSH in the estrogen- primed ovariectomized rat were not mirrored by correl- ative changes in the mRNA levels for these hormones. Q 1992 Academic Preus. Inc.

INTRODUCTION

During the estrous cycle in the rat, the key event that precedes and initiates ovulation is the precisely timed surge of gonadotropins, luteinizing hormone (LH), and follicle-stimulating hormone (FSH) [(l), for review]. The preovulatory surge of gonadotropins has been thought to be triggered by the gonadal steroid estradiol (l-3). How-

ever, recent work by our laboratory and others has pro- vided substantial evidence suggesting that the gonadal steroid progesterone also plays an important role in en- hancing the preovulatory gonadotropin surge and ensur- ing a full ovulation (4-8).

Progesterone is well known to be a potent inducer of gonadotropin surges in estrogen-primed ovariectomized rats (1, 9) and in pregnant mare serum gonadotropin- primed intact immature rats, in which it also facilitates ovulation (1, 10). This gonadotropin-releasing effect of progesterone appears to be physiologically important in proestrus in the rat. This is evidenced by studies showing that blockade of progesterone action via administration of progesterone antagonists or progesterone synthesis in- hibitors results in a significant attenuation of the proes- trous gonadotropin surge and ovulation (4-8, 11).

The recent focus of our laboratory has been on deter- mining the underlying mechanisms and site(s) of action involved in progesterone induction of gonadotropin se- cretion. Along these lines, we have provided evidence of hypothalamic catecholamine (12), neuropeptide y (13), and excitatory amino acid (14) neurotransmission in- volvement in progesterone induction of GnRH and go- nadotropin release. However, one interesting and unre- solved question is whether progesterone’s ability to induce the gonadotropin surge is coupled with or independent of parallel synthesis of LH/3- and FSHfl-subunit mRNA.

Therefore, the purpose of the present study was to de- termine whether the progesterone-induced surge of serum gonadotropins in the estrogen-primed ovariectomized immature rat was accompanied by correlative changes in the LHB- or FSH&subunit mRNA in the anterior pitu- itary. While chronic progesterone treatment has been found either to be inhibitory or to have no effect on go- nadotropin subunit mRNA (E-17), there has been no correlative study examining the effect of progesterone on LH/3 and FSH/3 mRNA during the time in which it induces the gonadotropin surge. Such a study is important because the pituitary stores substantial quantities of LH and FSH and their release may or may not be dependent upon new

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172 BRANN ET AL.

synthesis of these hormones. Since there have been many reports that glucocorticoids can cause selective release of FSH in vitro and in uivo [(18), for review], a second ob- jective of the study was to examine the effect of dexa- methasone on serum FSH levels and FSHP-subunit mRNA levels in the anterior pituitary.

MATERIALS AND METHODS

Animal Model

The animal model used in these studies has been de- scribed previously (9). Briefly, immature female Holtzman virus-free rats (Harlan Co., Madison, WI) were bilaterally ovariectomized under ether anesthesia at 26 days of age. They were maintained in air-conditioned rooms with a 14-h light/lo-h dark cycle (lights on at 0500 h, off at 1900 h) and were given water and rat chow ad libitum. At 1700 h on Days 27 and 28, the animals received an injection of estradiol (E2) or vehicle (Sigma, St. Louis, MO; 2 pg/ rat, subcutaneously; 25% ethanol-saline vehicle, 0.2 ml). At 0900 h the following day (29 days of age), the animals received either vehicle, progesterone, or dexamethasone (Dex) (all steroids: 1 mg/kg body wt; 25% ethanol-saline vehicle, 0.2 ml; Sigma, St. Louis, MO). Six animals were used per group and two pituitaries were pooled for RNA extraction to give replication in triplicate.

Tissue Preparation

The animals were sacrificed by decapitation at 2-h in- tervals starting 1 h after the vehicle or steroid treatment, trunk blood was collected for serum LH and FSH mea- surements, and the anterior pituitary was rapidly removed and the posterior pituitary was dissected and discarded. Once isolated the anterior pituitaries were homogenized in RNAzol for subsequent RNA extractions.

RNA Extraction

The detailed RNA extraction procedure as employed has been described previously (19). Briefly, total anterior pituitary RNA was extracted from vehicle- or steroid- treated ovariectomized rats at 29 days of age utilizing RNAzol (Biotecx Laboratories, Houston, TX), the com- mercial product (20) derived from the acid guanidinium thiocyanate phenol chloroform procedure first described by Chomczynski and Sacchi (21). The final RNA pellet was solubilized in 0.5% SDS in DEPC-treated water (21) and stored at -70°C; the RNA concentration was esti- mated by OD at 260 nm readings.

Northern Blotting

The Northern blotting procedure as utilized in these experiments has been previously described (22). Briefly, total RNA (5 pg/lane) was glyoxal denatured (Clontech 8021-l) and electrophoresed (1.5% agarose; 0.01 M sodium

phosphate, pH 7.5) as previously described (23). Also in- cluded was an RNA marker lane (5 pug; BRL 5620SA). Xylene cyan01 (Bio-Rad 161-0423) and bromphenol blue (Bio-Rad 161-0404) were utilized as tracking dyes. At the completion of the run, marker lanes were removed while that portion of the gel intended for probing was alkaline transferred (24) by capillarity to BioTrans + membrane (ICN BNBZF3050). The membrane was then baked under vacuum; prehybridization and hybridization were in so- lutions as described by Davis et al. (25) with the addition of 0.5% SDS. After hybridization for 24 h at 42°C the blots were washed as previously described (22) and ex- posed to Kodak X-OMAT film utilizing two Fisher Bio- tech L Plus intensifying screens (FBIS-1417). Exposures of 24-48 h duration typically yielded autoradiograms that were suitable for scanning in an LKB/Pharmacia UltroScan laser densitometer; data were expressed as ar- bitrary densitometric units (ADU). Figure 1 presents a representative example of 28 S rRNA stained with Stains All; these particular bands were derived from those sam- ples probed for the a-subunit mRNA at 1500 h (vehicle only and Ez + Dex; Fig. 1, lanes l-3 and 4-6, respectively) and 1700 h (vehicle only and Ez + Dex; Fig. 1, lanes 7-9 and 10-12, respectively). A representative Northern blot and densitometric scan as utilized to generate the gonad- otropin subunit mRNA data in Figs. 3-5 is depicted in Fig. 2. To take into account potential differences in gel loading, the relative abundance of 28 S rRNA in each lane of the gel was determined utilizing a cationic car- bocyanine dye (Stains All; Bio-Rad 161-0413) according to a recently published protocol (19). The relative staining intensity of the 28 S rRNA band in each lane as deter- mined by an LKB/Pharmacia UltroScan was then utilized to adjust the data for observed variability in gel loading.

Probes

The preparation of the cDNA probes for the gonado- tropin subunit mRNAs has been previously described (22). Briefly, the 350-bp cDNA probe for the LHP-subunit

123 4 56 7 8 9 10 11 12

FIG. 1. Representative example of 28 S rRNA stained with Stains All utilized for assessment of variability in gel loading. Total anterior pituitary RNA was glyoxal denatured and run on 1.5% agarose gel; the 28 S rRNA band was then stained according to a previously published protocol (19) utilizing Stains All. The gel was photographed and the photograph was scanned with an LKB/Pharmacia UltroScan laser den- sitometer to assess and correct for variability in gel loading. Figure 1 presents the 28 S rRNA hands observed in those samples probed for the a-subunit mRNA at 1400 h (vehicle only and E2 + Dex; lanes l-3 and 4-6, respectively) and 1600 h (vehicle only and E2 + Dex; lanes 7- 9 and 10-12, respectively).

STEROID EFFECT ON LH AND FSH mRNA 173

2.5

FIG. 2. Representative Northern blot and densitometric scan of anterior pituitary FSH&subunit mRNA from 1000 h time point. Fol- lowing administration of vehicle, e&radio1 (E,), E2 + Dex, or Ez + P, (lanes 1-4, respectively) at 0900 h, anterior pituitaries were collected at 1000 h for RNA extraction. Total RNA (5 ag) was glyoxal denatured and electrophoresed on 1.5% agarose gel followed by alkaline transfer to BioTrans + membrane. The membrane was then hybridized to the [32P]dCTP-labeled cDNA probe for the FSH@-subunit mRNA. Following exposure to Kodak X-OMAT film, the autoradiogram was scanned in an LKB/Pharmacia UltroScan laser densitometer. Data are expressed as arbitrary densitometric units (ADU).

mRNA and the 495bp cDNA probe for the a-subunit mRNA were generous gifts from Dr. William W. Chin. The 880-bp cDNA probe for the FSH@-subunit mRNA was a generous gift from Dr. Richard A. Maurer. Following amplification in HB-101, plasmid isolation was according to the Promega polyethylene glycol procedure (Promega Biological Research Products, Madison, WI). Following restriction, the cDNA inserts were isolated in low-melting- point agarose and subsequently labeled to a specific ac- tivity of high lo8 to low 10’ dpm/pg utilizing the random primer method (Pharmacia 27-9250-01). Labeled probes were used at 1 X lo6 dpm/ml hybridization solution.

RIA of LH and FSH

The concentrations of LH and FSH in serum samples were analyzed by a double-antibody RIA method as de- scribed by Rao and Mahesh (5). The purified hormones and standards and the first antibodies for LH [NIDDK- rLH-S-10 (rabbit)] and FSH [NIDDK-rFSH-Sll (rab- bit)] were obtained from NIDDK, NIH. The purified hor- mones were iodinated with lz51 (Amersham, Arlington Heights, IL) by the chloramine-T method. The second antibody was purchased from Arnell, Inc. (Brooklyn, NY). A 25% binding was obtained at 1:46,825 and 1:25,000 di- lutions for LH and FSH antisera, respectively. The assay was linear at 4-128 rig/tube for LH and 32-512 rig/tube for FSH. The intra- and interassay variabilities as deter- mined by analysis of replicate serum pool samples were 8 and 12.0% for LH and 6 and 10% for FSH. Hormone levels are expressed in terms of NIDDK-RP-1 standards for LH and FSH.

Statistical Analysis

The results given in the text are expressed as means + SEM. The differences between the experimental groups were analyzed using one-way analysis of variance, and comparisons between treatment means were made by a Student-Newman-Keuls multirange test. Where appro- priate, when comparing two groups only, the Student t test was employed. P < 0.05 was considered significant.

RESULTS

The results of a preliminary experiment in which ste- roid effects on (Y-, LH@-, and FSH&subunit mRNA and LH and FSH serum levels were determined at 1000 h (1 h after progesterone or dexamethasone treatment) are presented in Fig. 3. E&radio1 treatment appeared to lower a-subunit mRNA compared to vehicle-treated ovariec- tomized controls but this effect was not statistically sig- nificant. Estradiol treatment significantly suppressed LHP- and FSH&subunit mRNA without a corresponding decrease in serum levels of LH or FSH.

Dexamethasone treatment of estrogen-primed rats ap- peared to increase a-subunit mRNA compared to the es- trogen alone control, but this effect was not statistically significant. LH/3-subunit mRNA and LH serum levels in dexamethasone-treated rats were not different from those of the estrogen-treated control. Interestingly, dexameth- asone induced a significant elevation of FSH@ mRNA compared to estrogen-treated control rats, while serum levels of FSH were reduced by dexamethasone.

Progesterone treatment of estrogen-primed rats had no significant effect on (Y-, LH/?-, or FSHP-subunit mRNA levels as compared to estrogen-treated control rats even though it appeared to reduce serum LH levels (not sta- tistically significant) and serum FSH levels (P < 0.05).

Figures 4A-4D depict results from a full-time-course experiment in which the effects of progesterone in estro- gen-primed rats on LH and FSH serum levels (Figs. 4A and 4B) and P-subunit mRNA levels in the anterior pi- tuitary (Figs. 4C and 4D) are compared to estrogen plus vehicle-treated controls. Progesterone treatment pro- duced a well-defined surge of LH serum levels with peak values at 1400 h followed by a sharp fall at 1600 h to a level which was still, however, significantly elevated over that of the estrogen control (Fig. 4A). As shown in Fig. 4C, both e&radio1 plus vehicle-treated and e&radio1 plus progesterone-treated rats had LHP levels significantly lower than those of the vehicle only control. LH&subunit mRNA levels in progesterone-treated rats basically mir- rored and were no different from those of the estrogen- treated control, with the exception of 1400 h in which a small but significant elevation of LH/3-subunit mRNA was induced by progesterone.

As shown in Fig. 4B, progesterone exhibited a biphasic effect on serum levels of FSH, in that at 1000 h it sup-

174 BRANN ET AL.

Alpha Subunit mRNA, lOOOh

LH-B Subunit mRIVA, lOOOh

Serum LH Levels, lOOOh

p 60- 2 2 40-

3 20-

FSH-B Subunit tnJWA, lOOOh 25

3 20 2 f 15 .Z j 10

$5 4

0

Serum FSH Levels, ZOOOh 2000

FIG. 3. Preliminary experiment demonstrating steroid effect on gonadotropin serum levels and subunit mRNAs in the anterior pituitary in ovariectomised rats at 1000 h on Day 29, 1 h after progesterone or dexamethasone treatment. Twenty-six-day-old ovariectomixed rats received either vehicle or 2 gg of estradiol (E,) on Days 27 and 28. On Day 29, the animals received vehicle, progesterone (E, + P,) (1 mg/kg), or dexamethasone (& + Dex) (1 mg/kg) at 0900 h and were killed 1 h later for serum LH and FSH measurements and mRNA measurements. n = 6 rats per group. Groups with different subscripts are significantly different, P < 0.05.

pressed serum FSH levels while at 1400 and 1600 h it significantly elevated FSH levels compared to the estrogen alone control. In Fig. 4D, both estradiol plus vehicle- treated and estradiol plus progesterone-treated rats had FSH/3-subunit mRNA levels significantly lower than those of the vehicle only controls. The pattern of proges- terone effect on se= FSH levels was not mirrored in its effect on FSHP-subunit mRNA since progesterone-treated rats had FSH&subunit mRNA levels that were no dif- ferent from those of the estrogen alone controls (Fig. 4D).

Figures 5A-5D depict the effect of dexamethasone on LH and FSH serum levels and B-subunit mRNAs in es-

trogen-primed ovariectomized rats. As shown in Fig. 5A, dexamethasone had no significant effect on serum LH levels compared to the estrogen alone control. However, as shown in Fig. 5C, while estradiol plus vehicle-treated and estradiol plus dexamethasone-treated rats had LHB- subunit mRNA levels that were significantly lower than those of vehicle only control rats, dexamethasone treat- ment did significantly elevate LHB-subunit mRNA levels compared to estrogen alone control rats at 1400 h.

The effect of dexamethasone on serum FSH levels was similar to that observed for progesterone in that an early suppression of serum FSH levels was observed at 1000 h

STEROID EFFECT ON LH AND FSH mRNA

A SERUM LH B SERUM FSH

1000 1200 1400 1600 1000 1200 140@ 1600 Time (hours) Time (hours)

C LH-B Subunit mRNA D FSH-B Subunit mRNA 25-

s . oux + Vek

2 15- ii .m

B lo-

B - H 5- a

01 1 01 I 1 t 1000 1200 1400 1600 1000 1200 1400 1600

Time (hours) Time (hours)

FIG. 4. (A-D) Effect of progesterone on serum LH and FSH levels and LH& and FSHB-subunit mRNA levels in the anterior pituitary in estrogen-primed ovariectomised immature rats. The model is as described in Fig. 3. The Ovx t Veh control is from a 1006 h time point and is presented for comparison only. E2, estradiol; Veh, vehicle; P1, progesterone; Ovx, ovariectomized. n = 6 rate per group. *P c: 0.05 vs Ez + Veh; **P < 0.01 vs E2 + Veh.

followed by a pattern of increase that reached significance at 1600 h (Fig. 5B). As shown in Fig. 5D, dexamethasone caused a marked elevation of FSHj3-subunit mRNA levels 1 h after administration (1000 h) compared to estrogen alone, such that FSH@-subunit mRNA levels were no longer significantly different from those of the vehicle only control.

DISCUSSION

During the estrous cycle in the rat, the preovulatory surge of serum LH and FSH levels is known to be par- alleled by a corresponding increase in the P-subunit mRNA for these hormones (26, 27). However, at other times in the cycle there are instances of increased gonad- otropin b-subunit mRNA levels without increased serum gonadotropin levels (26, 27). Since it is well-known that the gonadal steroids estradiol and progesterone are the principal agents responsible for the induction of the proestrous surge of serum gonadotropins in the rat [(l),

for review], the salient question arises of whether it is estradiol, progesterone, or both that are responsible for the changes in gonadotropin subunit mRNA seen on proestrus. Previous studies addressing this question using the ovariectomized adult rat in which estradiol induces a distinct LH surge found that estradiol caused only a slight, nonsignificant elevation of LHP-subunit mRNA imme- diately preceding the estradiol-induced surge of serum LH (28). A similar lack of a significant effect of estradiol on LH/?-subunit mRNA during the estradiol-induced surge of serum gonadotropins has also been reported in the anestrous ewe (29). Due to the lack of a significant effect of estradiol on LH&subunit in the above studies, the pur- pose of our present study was to examine the effect of progesterone on LH@ and FSH@-subunit mRNA in an estrogen-primed immature ovariectomized rat model de- veloped in our laboratory in which the estrogen is ad- ministered in ethanol-saline and a distinct surge of FSH and LH can be induced by the administration of proges- terone (9). The results presented in this study show that

176 BRANN ET AL.

SERUM LH

I - E2+Veh - J%2+Dex I

1000 1200 1400 1600 Time (hours)

LIZ-B Subunit mRNA D FSH-% Subunit mRNA 25 00.x + Veh

1000 1200 1400 1600 Time (hours)

B SERUM FSH 2000

1 .

01 loo0 1200 1400 1600

Time (hours)

Veh

l *

0-c I I 1 1000 1200 1400 1600

Time (hours)

FIG. 5. (A-D) Effect of dexamethasone on serum LH and FSH levels and LHfi- and FSHP-subunit mRNA levels in the anterior pituitary in estrogen-primed ovariectomized immature rats. The model is as described in Fig. 3. The Ovx + Veh control is from a 1000 h time point and is presented for comparison only. ES, estradiol; Veh, vehicle; Dex, dexamethasone; Ovx, ovariectomized. n = 6 rats per group. *P < 0.05 vs El + Veh; **P < 0.01 vs Es + Veh.

progesterone induced a distinct surge of serum gonado- tropins compared to the estradiol-treated control animals, whose serum LH and FSH values were only slightly el- evated at 1400 h. We have previously shown that our es- tradiol treatment in this model is sufficient to induce pro- gesterone receptors and sensitivity while having only a minor or no effect on serum gonadotropins (9). Therefore, this model is an excellent one for examining the mecha- nisms of progesterone-induced gonadotropin secretion. At the time of peak serum LH levels induced by progesterone (1400 h), a small but significant increase in LHB-subunit mRNA was noted in progesterone-treated rats compared to the estradiol control rats. There was no other significant difference noted in the LH/3-subunit mRNA levels be- tween the progesterone-treated rats and the estradiol control rats at any other time point. The specificity of the small elevation of LH/3-subunit mRNA by progester- one at 1400 h is questionable, since a similar small, yet significant, elevation of LH&subunit mRNA at 1400 h was also observed in dexamethasone-treated rats that did

not show a surge in serum LH levels. Even though pro- gesterone produced a surge in serum FSH levels at 1400 and 1600 h after an initial suppression at 1000 h compared to the estrogen alone controls, FSH@subunit mRNA lev- els were unchanged by progesterone compared to estrogen alone controls, except for a slight nonsignificant increase at 1200 h noted in the progesterone-treated animals.

It should be noted that in both the estradiol only con- trols and the estradiol plus progesterone-treated rata there was a nonsignificant pattern of elevation of LH&subunit mRNA from 1000 to 1200 h, which was followed by a significant fall in the LHP-subunit mRNA in both groups at 1400 h. This slight nonsignificant increase of LHP- subunit mRNA preceding the progesterone-induced LH surge is reminiscent of a similar nonsignificant elevation of LH@ mRNA preceding the estradiol-induced LH surge in the ovariectomized adult rat reported by Haisenleder et al. (28) and in the anestrous ewe reported by Landefeld et al. (29). This slight, but consistent, increase of LH/3 mRNA reported in these studies, while not statistically

STEROID EFFECT ON LH AND FSH mRNA 177

significant, still could reflect an inherent role for estradiol to prepare the animal for progesterone induction of go- nadotropin surges by increasing the mRNA for synthesis of new hormone. The newly synthesized LH could con- tribute to the pool of hormone released during the pro- gesterone-induced surge of serum gonadotropins later in the day.

Nevertheless, one cannot ignore the fact that in both the estradiol-treated and the estradiol plus progesterone- treated groups, there was a precipitous and highly signif- icant drop in the LH@ mRNA at 1400 h, the time of peak serum LH levels induced by progesterone. This would seem to strongly suggest that an acute increase in the gonadotropin P-subunit mRNA is not a prerequisite for increases in serum gonadotropin secretion involved in the expression of the progesterone-induced surges of LH and FSH in the estrogen-primed ovariectomized immature rat. A similar conclusion had been reached by Haisenleder et al. (28) with respect to the estradiol-induced surge of go- nadotropins in the adult ovariectomized rat. To our knowledge, only one study has reported an increase in gonadotropin P-subunit mRNA during a steroid-induced gonadotropin surge (30). In that study, progesterone im- plants in intact immature female rats were found to sig- nificantly elevate FSHP-subunit mRNA and FSH serum levels 5 h after administration (LHP mRNA was not ex- amined). In our study using ovariectomized immature rats primed with estradiol and administered an acute injection of progesterone, we noticed a pattern of increased FSHP- subunit mRNA 3 h after treatment (1200 h), but this effect was not statistically significant. The difference between our findings and the above findings could be due to the different animal models used (ovariectomized versus in- tact) and the different mechanism of steroid administra- tion (injection versus implants). Nevertheless, our find- ings are in agreement with other reports in the literature that fail to find an effect of steroids on gonadotropin p- subunit mRNA levels at times of the steroid-induced surge of serum gonadotropins (28, 29).

Finally, our study demonstrates that glucocorticoids can stimulate FSHfl-subunit mRNA levels in the anterior pi- tuitary of the rat, a finding in agreement with the nu- merous previous reports of selective FSH-releasing activ- ity of glucocorticoids in vitro and in uiuo (l&31-33). Pre- vious studies showing that glucocorticoids increase pituitary content of FSH both in vitro and in vivo (34- 36) had led to the suggestion that glucocorticoids could increase synthesis of FSH, as well as secretion. Our study demonstrating a highly significant increase in FSHP-sub- unit mRNA within 1 h by dexamethasone would seem to provide strong evidence supporting this suggestion. Serum FSH levels were elevated 7 h after dexamethasone treat- ment in our study, and it may be that the FSH/3 mRNA that is rapidly induced at 1000 h by dexamethasone could contribute to the increased FSH secretion later in the day. It should be noted that, similar to our finding of

increased FSHP mRNA in the pituitary of female rats, Ringstrom et al. (37) have recently shown that cortisol has a similar effect on FSHfl mRNA in male rats.

In conclusion, a strong correlation between the effect of progesterone on serum gonadotropin levels and gonad- otropin P-subunit mRNA levels in the anterior pituitary was not demonstrated by the present study. This finding suggests that an acute increase in gonadotropin P-subunit mRNA is not a major mechanism for positive progester- one feedback in the rat, but that progesterone effects may be largely on GnRH secretion (12, 13,38-40) and/or in- creasing sensitivity of the gonadotropes to GnRH (41- 43). Additionally, it suggests that progesterone acts by stimulating the release of preformed gonadotropins. Dexamethasone, on the other hand, exerts selective mod- ulatory effects on both FSHP mRNA levels and the se- cretion of FSH in the estrogen-primed ovariectomized immature rat.

ACKNOWLEDGMENTS

This investigation was supported by Research Grants HD-16666 (V.B.M.) and HD-17614 (J.O.), NICHD, National Institutes of Health, U.S. Public Health Service, and a Basic Research Support Grant (J.O.).

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