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Department and Section of Physiology, College of Veterinary Medicine and Division of Biological Sciences, Cornell University, Ithaca, New York 14853
Address all correspondence and requests for reprints to: Joanne E. Fortune, Ph.D., Cornell University, 923 VRT, Ithaca, New York 14853. E-mail: jf11{at}cornell.edu
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Abstract |
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 Top Abstract IntroductionMaterials and MethodsResultsDiscussionReferences |
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-hydroxylase (17-OH) were
localized in frozen sections from each follicle. The expression of mRNA
for LH receptor in granulosa cells was always at or near background and
was not different between days or follicle types (P
= 0.63). In contrast, the expression of mRNA for LH receptor in theca
cells of the same sections was readily detectable; there was no
difference between follicle types on the second day of the follicular
wave, but by the third day expression in the subordinate follicles had
decreased (P < 0.05). The expression of mRNA for
FSH receptor was highest in granulosa cells of dominant follicles
collected on day 3 of the follicular wave (P <
0.05) and was not different between dominant and subordinate follicles
on day 2 of the wave (P > 0.05). The expression of
mRNA for aromatase in granulosa cells was similar
(P > 0.05) between the dominant follicles on days
2 and 3 and the largest subordinate follicle on day 2 of the follicular
wave and was much lower in the remaining follicles
(P < 0.01). On day 2 of the wave, the expression
of mRNA for 17-OH was not different between the dominant and
subordinate follicles, but by day 3 the dominant follicles had more
mRNA for 17-OH than the subordinate follicles (P
< 0.05). These data show that the dominant follicle had been selected by the second day of the follicular wave (based on diameter and estradiol secretion) and that selection occurred in the absence of detectable levels of mRNA for LH receptor in the granulosa cells or differences between dominant and subordinate follicles in mRNA for LH receptor in theca cells or FSH receptor in granulosa cells. However, the divergent pattern of growth between dominant and subordinate follicles (after follicle selection) was associated with higher levels of mRNA for gonadotropin receptors and steroidogenic enzymes in dominant compared with subordinate follicles. Therefore, selection of the dominant follicle in cattle does not appear to involve the regulation of expression of mRNA for gonadotropin receptors, although such regulation may be important at other stages of differentiation of the dominant follicle.
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Introduction |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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Transient increases in FSH secretion precede the emergence of follicular waves in cattle (reviewed in Refs. 5, 6) and a subsequent decrease in circulating FSH concentrations is temporally associated with selection of the dominant follicle (7) as has been observed in primates (see 8). Hence, it is assumed that it is the ability of follicles to respond to the endocrine environment on an individual basis that dictates their particular patterns of growth and development (9).
Granulosa cells acquire FSH receptors, and theca cells acquire LH receptors before follicular recruitment (wave emergence) and selection of the dominant follicle (see Ref. 10). During the later stages of follicle development, the granulosa cells of large healthy, estrogen-secreting follicles also acquire LH receptors (rats, reviewed in Ref. 10 ; mice, Ref. 11 ; pigs, Refs. 12, 13, 14 ; sheep, Refs. 15, 16, 17, 18, 19 ; cattle, Refs. 20, 21, 22, 23, 24, 25, 26, 27). During the bovine estrous cycle, granulosa cells of large healthy antral follicles have more LH receptors than smaller atretic follicles (20, 21, 22, 25) and levels of messenger RNA (mRNA) for LH receptor in granulosa cells of dominant follicles increase as follicular development progresses (27). It has been suggested that the acquisition of LH receptors on granulosa cells plays an important role in the selection of dominant follicles (9, 26, 27, 28), but comparisons of gonadotropin receptor expression among dominant and subordinate follicles of the same follicular cohort have not been made.
The aim of the present study was to test the hypothesis that differences in the pattern of expression of mRNA for LH and FSH receptors are associated with the process of selection of dominant follicles in cattle. This was achieved by collecting dominant and subordinate follicles on days 2 and 3 of the first follicular wave of the cycle (around the time of dominant follicle selection), determining their steroidogenic capabilities, and measuring mRNA for the steroidogenic enzymes and gonadotropin receptors.
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Materials and Methods |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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(25 mg Lutalyse, im; Upjohn,
Kalamazoo, MI) to induce luteolysis, a follicular phase, and ovulation.
Dominant and subordinate follicles were obtained at specific times
during the first follicular wave of the next estrous cycle. Starting
the day before PGF2 injection, the ovaries of each heifer were
examined daily by transrectal ultrasonography using a 7.5 MHz linear
array transducer and a B-Mode scanner (Aloka 500V, Corometrics Medical
Systems Inc, Wallingford, CT) to monitor changes in ovarian follicular
growth, as we have described previously (2). Animals were checked for
estrus once daily by observation of their behavior during the outdoor
exercise period; heifers allowing others to mount were considered to be
in estrus. The day of estrus was designated as Day 0 of the cycle.
The day of emergence of the first follicular wave was designated as day
0 of the wave and was retrospectively identified as the last day on
which the dominant follicle was 4 or 5 mm in diameter. Heifers were
randomly assigned to be ovariectomized on day 2 or 3 of the first
follicular wave (n = 3 per group). These days were chosen based on
a retrospective analysis of ultrasonographic data from 177 waves of
follicular development (from 74 animals) previously recorded by members
(n = 5) of our laboratory (2, 29–32 and unpublished
observations). The analysis showed that the dominant follicle was
present as the largest follicle of the cohort 79% of the time on day 2
and 93% of the time on day 3 of the follicular wave. In only 3% and
2% of waves was the diameter of a subordinate follicle greater than
that of the dominant follicle on day 2 and 3 of the wave, respectively.
In the remaining cases, the subordinate follicle(s) were similar in
size to the dominant follicle. These data are in agreement with those
of Ginther et al. (3), who reported that divergence in
diameter between dominant and subordinate follicles occurred by day 2
of the follicular wave. In the present experiment, ovaries were
collected on day 2 of the wave from animals in which one follicle was 1
mm larger (ultrasonography measurement) than any other follicle and on
day 3 when a clear dominant follicle was present (
2 mm
difference).
Ovariectomies were performed via colpotomy under local anesthesia (2% procaine, epidural), and the ovaries were transported to the laboratory within 15 min of collection in ice-cold MEM containing 25 mM HEPES buffer supplemented with penicillin (50 U/ml) and streptomycin (50 µg/ml; Life Technologies, Grand Island, NY). All animal procedures were approved by the Cornell University Animal Care and Use Committee (protocol no. 86–214-95).
Follicle isolation and tissue culture
The dominant follicle in each animal was identified as the
largest follicle of the cohort on the day of ovariectomy; this was
later confirmed by follicular fluid estradiol concentrations. Follicles
identified as the dominant and the two largest subordinate follicles
were dissected from the ovaries and measured to the nearest 0.5 mm as
described previously (33). Follicular fluid was then aspirated and
stored at -20 C for later measurement of steroids. Each follicle was
cut open and a portion frozen in embedding medium (Tissue-Tek, Miles
Inc., Elkhart, IN) and stored at -80 C for later use for in
situ hybridization. The remainder of the follicle was prepared for
tissue culture; the theca interna and adherent granulosa cells
(i.e. the follicle wall) were peeled away from
the theca externa/stroma and cut into equal-sized pieces (about
2.5 x 2.5 x 0.25 mm) ranging from 2–5% of the total
tissue, depending on the size of the follicle.
The pieces of follicle wall were transferred into 24-well cluster dishes (three pieces per well; Costar, Cambridge, MA) and cultured in 0.5 ml medium. The medium was MEM supplemented with penicillin, streptomycin, glutamine, nonessential amino acids (50 U/ml, 50 µg/ml, 2 mM, 0.1 mM, respectively; Life Technologies), insulin, transferrin (1 µg/ml, 5 µg/ml, respectively; Collaborative Research Inc., Waltham, MA) and cortisol (40 ng/ml; Sigma Chemical Co., St. Louis, MO). Pieces of follicle wall were cultured in duplicate wells per follicle with either medium alone or medium plus testosterone (0.5 µM) for 24 h in 5% CO2, 95% air at 37 C. At the end of culture, medium was collected and stored at -20 C for later measurement of steroids and the pieces of follicle wall were weighed. Steroid production in vitro was expressed as nanograms secreted per 24 h per mg tissue.
RIAs
Duplicate aliquots (0.1–5 µl) of follicular fluid from each
dominant and subordinate follicle were assayed without extraction for
estradiol, androstenedione and progesterone by RIA, as described and
validated previously (34). Estradiol was also measured in medium from
follicle wall pieces cultured with and without testosterone and
androstenedione was measured in medium from cultures treated with
control medium. Samples of medium were assayed in duplicate. Inter- and
intraassay coefficients of variation ranged between 3% and 6% and the
estradiol, androstenedione, and progesterone assays had sensitivities
of 5, 12.5, and 12.5 pg/assay tube, respectively.
In situ hybridization
In situ hybridization was carried out as previously
described by our laboratory (35), except that frozen tissue was used.
In brief, 14 µm thick sections were cut and mounted on
poly-L-lysine coated slides (Sigma) and stored at -80 C
until needed. Sections were dried on a slide-warmer at 55 C for 60 min
and fixed for 5 min in freshly prepared 4% paraformaldehyde in 0.1
M sodium phosphate buffer. Sections were rinsed twice in
buffer for 15 min and in water for 5 min and then treated for 5 min in
0.1 M triethanolamine (pH 8) followed by 10 min in 0.1
M triethanolamine with 0.25% acetic anhydride. Sections
were rinsed for 5 min in double strength standard sodium citrate
solution (SSC; 1x = 150 mM sodium chloride and 15
mM sodium citrate, pH 7) and then prehybridized with 50
µl of hybridization buffer [50% formamide, 0.3 M sodium
chloride, 20 mM Tris (pH 8), 5 mM EDTA, 10
mM sodium phosphate buffer, 1x Denhardt’s solution
(0.02% each of Ficoll, polyvinylpyrrolidone, and BSA), 10
mM dithiothreitol (DTT), 500 µg/ml yeast transfer RNA]
for 2 h at 55 C in a humid chamber.
Radiolabeled sense and antisense riboprobes were generated by in
vitro transcription, as described previously (36), using
35S-labeled UTP (UTP35S; Dupont-New England
Nuclear Products, Boston, MA). The recombinant plasmid containing the
bovine P450 aromatase complementary DNA (cDNA) (36) was linearized with
BamHI or KpnI, and 137-bp sense or antisense
probes were transcribed using T7 or T3 RNA polymerase, respectively.
The cDNA for the bovine P450 17-hydroxylase (17-OH) enzyme was
generously provided by Dr. M. Waterman (Vanderbilt University, School
of Medicine, Nashville, TN; 36, 37) and after linearization with
BamHI or EcoRI, 299-bp sense or antisense
riboprobes were generated using T7 or T3 RNA polymerase, respectively.
The cDNAs for the ovine LH and bovine FSH receptors were kindly
provided by Dr. M. Smith (University of Missouri, Columbia, MO; 27).
The LH receptor cDNA was linearized with NsiI and sense (406
bp) or antisense (333 bp) riboprobes were generated using T7 or T3 RNA
polymerases, respectively. The FSH receptor cDNA was linearized with
BglII and sense (126 bp) or antisense (509 bp) riboprobes
were generated using T3 or T7 RNA polymerases, respectively. After
phenol-chloroform extraction the riboprobes were precipitated with 0.5
vol of 7.5 M ammonium acetate and 2 vol of ethanol.
After the prehybridization, hybridization buffer was blotted from around the sections and 1 x 106 cpm of riboprobe, diluted in hybridization buffer, was added to each slide. A coverslip was sealed over the sections and hybridization was carried out at 55 C for 14–16 h. The slides were then washed in 4x SSC for 30 sec to remove the coverslips, twice in 4x SSC plus 4 mM DTT for 15 min, in NTE buffer (0.5 M sodium chloride, 10 mM Tris [pH 8], 5 mM EDTA) at 37 C for 2 min, in Ribonuclease A (20 µg/ml; Sigma Chemical Co.) in NTE buffer at 37 C for 30 min, in NTE buffer plus 2 mM DTT at 37 C for 30 min, in 2x SSC plus 1.5 mM DTT for 20 min, in 0.1x SSC plus 1 mM DTT for 20 min, in 0.1x SSC for 5 min and dehydrated in increasing concentrations of ethanol with 0.3 M ammonium acetate. Autoradiography was performed overnight to determine the success of the hybridization, and then the sections were coated with NTB-2 emulsion (Eastman Kodak Co., New Haven, CT) and exposed at 4 C in a light-tight box. After 12–14 days, the sections were processed and lightly stained with hematoxylin.
The hybridization intensity was quantified using the NIH Image analysis system. Within a defined area, the total number of pixels was determined and the number of pixels occupied by silver grains was determined by setting a gray-scale threshold (0 to 120 units of pixel intensity). The hybridization intensity was calculated as the percentage of pixels occupied by silver grains as a result of binding of the antisense riboprobe minus the percentage occupied as a result of binding of the sense riboprobe, as described by Xu et al. (38). For each follicle, six fields were analyzed per tissue type (granulosa cells, theca cells and ovarian stroma) per section, on two sections per slide, on two slides for the antisense riboprobes, and on one slide for sense riboprobes. Ovarian stromal tissue in the same sections served as a negative tissue control for the in situ hybridization procedure.
Statistical analyses
All data were analyzed by ANOVA (General Linear Models procedure
of SAS, Cary, NC) with a nested design to evaluate the effects of day
of the follicular wave, animal (nested within day), follicle type
(dominant, or first or second subordinate), the interaction of day
x follicle type, individual follicle (nested within follicle type) and
the effect of treatment in vitro where appropriate. Before
ANOVA, each set of data was subjected to Hartley’s test for
heterogeneity of variance. When heterogeneity was present, data were
transformed before ANOVA by a method that yielded homogeneity of
variance (usually logarithmic transformation, but in one case square
root transformation was used). Post-ANOVA comparisons among means were
made using the Student-Newman-Kuel’s test (P < 0.05
and P < 0.01). All values given are the mean ±
SEM of untransformed values.
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Results |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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), both when measured
by ultrasonography (measures antral diameter only) or after dissection.
On day 2 of the follicular wave, the mean diameter of the dominant
follicle was greater (P < 0.05) than the diameter of
both subordinate follicles when measured by ultrasonography (Fig. 1A),
but only greater than the second largest subordinate follicle when
measured after dissection (Fig. 1B). Measurement of follicles after
dissection on day 2 showed that, within individual animals, the
dominant follicle was 1.0 mm, 1.0 mm, and 0.5 mm greater than the
largest subordinate follicle. As expected (39), follicle diameters
measured in vivo by ultrasonography were about 1–2 mm less
than the diameters of dissected follicles.
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). Follicular fluid
concentrations of estradiol in the first and second subordinate
follicles were greater on day 2 of the follicular wave compared with
similar subordinate follicles on day 3 (P < 0.05, Fig. 2A). Overall, androstenedione concentrations in follicular fluid were
greater on day 2 compared with day 3 of the first follicular wave
(P < 0.001). However, comparisons between means of
different follicle groups showed that only concentrations of
androstenedione in the largest subordinate follicles were greater on
day 2 vs. day 3 of the wave (P < 0.05; Fig. 2B). Progesterone concentrations in follicular fluid were not different
between follicle types or days of the wave (P > 0.45;
Fig. 2C).
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). Estradiol
secretion in control medium was highest by pieces of follicle wall from
dominant follicles on day 3 of the follicular wave and lowest by
subordinate follicles on day 3 (P < 0.05; Fig. 3B).
The addition of testosterone to the medium as a substrate for estradiol
synthesis increased estradiol secretion only by pieces of follicle wall
from dominant follicles collected on day 3 of the follicular wave
(P < 0.05; Fig. 3B). Androstenedione secretion in
control medium was not different between follicle types or days of the
follicular wave (P > 0.50, Fig. 4).
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and Fig. 6). Interestingly, on day 2
of the wave there was no difference (P > 0.05) between
dominant and subordinate follicles in the expression of mRNA for FSH
receptor (Fig. 5B). The expression of mRNA for LH receptor in granulosa
cells was not different between days or follicle types
(P = 0.63; Fig. 6 and Fig. 5A) and was at or near
background in all cases (overall mean hybridization intensity 0.9
± 0.3%). In contrast, the expression of mRNA for LH receptor in theca
cells of the same sections was readily detectable (Fig. 6). There was
no difference between follicle types in the expression of mRNA for LH
receptor in theca cells on the second day of the follicular wave (Fig. 5C), but by the third day expression in the subordinate follicles, but
not the dominant, had decreased relative to day 2 (P <
0.05, Fig. 5C).
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Expression of mRNA for steroidogenic enzymes
The expression of mRNA for aromatase in granulosa cells was
similar (P > 0.05) in the dominant follicles on days 2
and 3 and the largest subordinate follicle on day 2 of the follicular
wave (Fig. 7A). By day 3 of the follicular wave, the
expression of mRNA for aromatase in both subordinate follicles was
negligible and much lower than the dominant follicle (P
< 0.01; Figs. 7A and Fig. 8). The expression of mRNA
for 17-OH in theca cells of dominant follicles was similar on days 2
and 3 of the follicular wave (P > 0.05; Fig. 7B). On
day 2 of the wave, expression was not different between the dominant
and subordinate follicles, but by day 3 the dominant follicles had more
mRNA for 17-OH than the subordinate follicles (P <
0.05; Fig. 7B and Fig. 8).
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-OH mRNAs. |
Discussion |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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Dominant follicle selection has been defined as the time of divergence of the growth profiles of dominant and subordinate follicles (7). However, selection in a biological context is the process by which a single follicle avoids atresia and acquires the potential to ovulate. It is likely that selection of a dominant follicle occurs before differences in diameter. Cattle provide an excellent model for studies of follicular selection because regular, successive waves of follicular development occur throughout the estrous cycle. The waves appear to be equivalent in that each consists of sequential follicular recruitment, selection, and dominance and the dominant follicle of each wave can ovulate a fertile oocyte if luteolysis occurs naturally or is induced (4).
In the present study, on day 2 of the first follicular wave the mean
diameter of the dominant follicle was only slightly larger (about 1 mm)
than the largest subordinate follicle (Fig. 1
). Previous observations
by our laboratory indicated that in only 3% of waves does the largest
follicle on the second day of a follicular wave fail to develop into
the dominant follicle. Hence, in the present study there was a great
likelihood that the dominant follicle had been selected and was
identified by size on day 2 of the follicular wave. Although the
dominant follicles were only slightly larger than the largest
subordinate follicles on day 2 of the wave (Fig. 1), there was a great
difference in follicular fluid estradiol concentrations (Fig. 2A) and
estradiol secretion in vitro between these follicle types
(Fig. 3A). This confirmed their identification as dominant follicles
and is in agreement with studies showing that follicular fluid
concentrations of estradiol differed remarkably between groups of
follicles that had similar diameters (22) and that asymmetry between
ovaries in estradiol secretion into the ovarian veins occurred before
dominant follicles were recognizable in the ovaries (humans, 40;
cattle, 41).
The observation that the dominant follicle has been selected by day 2
of the follicular wave (based on individual follicle diameters and
estradiol secretion, Figs. 1 and 2), taken together with the lack of
mRNA for LH receptor in granulosa cells on days 2 or 3 of the
follicular wave (Figs. 5A and 6), suggests that the acquisition of LH
receptor on granulosa cells does not play a role in selecting the
dominant follicle. This is in general agreement with the findings of
Ireland and Roche (22), who showed that the binding of labeled hCG was
not different between estrogen-active and estrogen-inactive (presumably
dominant and subordinate) follicles on days 3 or 5 of the estrous
cycle, and also with the findings of Bodensteiner et al.
(42) who, using an RRA, showed that granulosa cells from dominant and
subordinate follicles, collected at the beginning of divergence (about
day 2 of the wave) had similar numbers of LH receptors. Xu et
al. (27) found that mRNA for LH receptor was not expressed in
granulosa cells of dominant follicles until day 4 of the follicular
wave. We suggest that this was after follicle selection had occurred
(3; Fig. 1) and therefore, that the expression of mRNA for LH receptor
in granulosa cells does not play a role in dominant follicle selection
in cattle. Our results also show that there was no difference between
follicle types on the second day of the wave in levels of mRNA for FSH
receptor in the granulosa cells or mRNA for LH receptor in the theca
cells. This suggests that the mechanism(s) by which follicle selection
occurs in cattle is not dependent on the regulation of gonadotropin
receptor expression. However, we cannot rule out potential differences
in expression of FSH receptor in granulosa cells or LH receptor in
theca cells of follicles destined to be dominant vs.
subordinate at an earlier stage of follicular recruitment, before they
diverge in size.
After follicle selection has taken place, the subsequent divergent
pattern of dominant and subordinate follicle growth may depend on the
regulation of gonadotropin receptor expression because differences
among follicle types in the expression of mRNA for FSH receptor in
granulosa cells and mRNA for LH receptor in theca cells were observed
on day 3 of the wave (Fig. 5). These data are in agreement with
previous observations that healthy large follicles contain more
gonadotropin receptors than smaller follicles in pigs (14, 43), sheep
(15, 17, 18) and cattle (20, 21, 22, 25). However, other reports indicated
that gonadotropin receptor binding did not differ between healthy and
atretic follicles of sheep (19) and cattle (23, 44).
The large differences between dominant and subordinate follicles in
follicular fluid concentrations of estradiol (Fig. 2A) and estradiol
secretion in vitro (Fig. 3) are in agreement with previous
studies (6, 45). However, in the current study the dominant follicles
collected on days 2 and 3 of the wave and the largest subordinate
follicles collected on day 2 of the follicular wave all had similar
levels of mRNA for aromatase (Fig. 7A). This is in contrast to their
very different follicular fluid concentrations of estradiol (Fig. 2A)
and aromatase activity in vitro (Fig. 3). This may indicate
that there may be translational regulation of mRNA for aromatase and/or
regulation of the enzyme itself during follicle development. Developing
follicles in sheep have been reported to contain an aromatase inhibitor
that has only local actions within the follicle (46). Alternatively,
the availability of insulin-like growth factors (IGFs), which are
believed to be regulated at least in part through IGF binding proteins
(IGFBPs; 47) and IGFBP proteases (48) may play an important role in the
regulation of steroid production and follicle development. On both day
2 and 3 of the follicular wave there were higher levels of IGFBP-4
protease activity in the follicular fluid from the dominant
vs. subordinate follicles used in the current study (48
and Chandrasekher, Y. A., A. C. O. Evans, L. C. Guidice, and J. E.
Fortune, unpublished observations).
The decrease in mRNA for LH receptor in theca cells of the subordinate
follicles (compared with dominant follicles) collected on day 3 of the
follicular wave (Fig. 5C) may explain the lower levels of mRNA for
17-OH (Fig. 7) and follicular fluid androstenedione concentrations
(Fig. 2B). However, the concomitant decrease in estradiol secretion
in vitro by the subordinate follicles cannot be explained by
a lack of androgen substrate since the addition of testosterone to the
cultures did not increase estradiol secretion. This reduction may best
be explained by a severe decrease between day 2 and 3 in the expression
of mRNA for the aromatase enzyme (Fig. 7A).
Aromatization is enhanced by FSH (49, 50). The expression of FSH
receptors in granulosa cells is regulated by FSH and estradiol may have
a synergistic role (51, 52). It has been shown that the combined
actions of estradiol and FSH stimulate the induction of LH receptors on
granulosa cells both in vivo (51, 53, 54) and in
vitro (52, 55, 56) and that there is a correlation between
follicular fluid concentrations of estradiol and LH receptors on bovine
granulosa cells (26, 57). Hence, based on information in the literature
and the present results, we suggest the following series of events
associated with dominant follicle selection and growth. As small antral
follicles grow and develop, they increase estradiol secretion. The
cause or consequence of dominant follicle selection (mechanism unknown)
is an increase in estradiol secretion by the dominant follicle compared
with the subordinate follicles. To this point, the future dominant and
subordinate follicles are similar in size, LH receptors on theca cells
and FSH receptors on granulosa cells, and in their lack of LH receptors
on granulosa cells (Fig. 7A). After selection has occurred, the
dominant follicle continues to increase in diameter and increases
estradiol secretion. FSH receptors on the granulosa cells of the
dominant follicle also increase, LH receptors on the theca cells of the
subordinate follicles decrease (Fig. 5) and estradiol secretion from
the subordinate follicles decreases (Fig. 2A and Fig. 3). The combined
effects of increased estradiol production and FSH action on the
granulosa cells of dominant follicles may stimulate the expression of
LH receptors in granulosa cells, enabling dominant follicles to
continue developing in an environment of basal gonadotropins and to
respond to the preovulatory LH surge.
In summary, in the present study the dominant follicle was selected by the second day of the follicular wave, and this occurred before the expression of mRNA for LH receptor in granulosa cells and before differences between follicle types in mRNA for FSH receptor in granulosa cells or LH receptor in theca cells. However, the divergent pattern of growth between dominant and subordinate follicles, after follicle selection (third day of the follicular wave), was associated with an increase in mRNA for FSH receptor in granulosa cells of dominant follicles and a decrease in mRNA for LH receptor in theca cells of subordinate follicles. Although the acquisition of LH receptors by granulosa cells may be very important at later stages of development of dominant follicles, the current results suggest that dominant follicle selection in cattle occurs before the development of LH receptors on granulosa cells.
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Acknowledgments |
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-hydroxylase and Dr. M. F. Smith for
cDNA probes for the ovine LH receptor and bovine FSH receptor. |
Footnotes |
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2 Current address: Department of Animal Science and Production,
University College Dublin, Ireland.
Received December 11, 1996.
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References |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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-induced luteolysis in cattle. Biol
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Endocrinology 136:5102–5110[Abstract]
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Regulation of gene expression by ACTH and elucidation of primary
sequence. J Biol Chem 261:2475–2482-hydroxylase, and cytochrome P450 aromatase in bovine follicles
during the first follicular wave. Endocrinology 136:981–989[Abstract]
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 M. Mihm, P.J. Baker, J.L.H. Ireland, G.W. Smith, P.M. Coussens, A.C.O. Evans, and J.J. Ireland Molecular Evidence That Growth of Dominant Follicles Involves a Reduction in Follicle-Stimulating Hormone Dependence and an Increase in Luteinizing Hormone Dependence in Cattle Biol Reprod, June 1, 2006; 74(6): 1051 - 1059. [Abstract] [Full Text] [PDF]
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K. Ndiaye, T. Fayad, D. W. Silversides, J. Sirois, and J. G. Lussier Identification of Downregulated Messenger RNAs in Bovine Granulosa Cells of Dominant Follicles Following Stimulation with Human Chorionic Gonadotropin Biol Reprod, August 1, 2005; 73(2): 324 - 333. [Abstract] [Full Text] [PDF]
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D. C. Woods and A.L. Johnson Regulation of Follicle-Stimulating Hormone-Receptor Messenger RNA in Hen Granulosa Cells Relative to Follicle Selection Biol Reprod, March 1, 2005; 72(3): 643 - 650. [Abstract] [Full Text] [PDF]
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