Category Archives: Hormone

Gonadotropin-Releasing Hormone: DISCUSSION(7)

DISCUSSION(7)

Alternatively, other signals that could affect selectively the a subunit gene expression may overcome the blocking effect of cetrorelix on the GnRH-regulated a subunit gene expression during sexual maturation.

In summary, this study shows that sexual development in male rats is accompanied by a progressive and concerted activation of GnRH-R and gonadotropin subunit genes in the anterior pituitary through the infantile and juvenile periods to puberty.

Gonadotropin-Releasing Hormone: DISCUSSION(6)

Also, the dramatic inhibition of FSHp mRNA levels by cetrorelix suggests that pituitary and gonadal activin is not a major contributor to the activation of the FSHp gene expression in the prepubertal period. However, a caveat to this conclusion must be introduced because significant evidence indicates that GnRH can indirectly regulate the FSHp gene through induction of activin and follistatin genes. In agreement with the blocking effect on gonadotropin p subunit gene expression, cetrorelix also abated serum LH and FSH concentrations and aborted the initial rise of serum total and free testosterone levels.

Gonadotropin-Releasing Hormone: DISCUSSION(5)

DISCUSSION(5)

Also, it has been reported that chronic cetrorelix treatment of female rats from Day 25 to Day 36 down-regulates (by 50%) the expression of the GnRH-R gene measured by quantitative reverse transcription-polymerase chain reaction (RT-PCR) assay. However, another report, also using RT-PCR, showed that GnRH-R as well as gonadotropin p subunit mRNAs were not affected by treatment of infantile female rats with a GnRH antagonist. These different responses can be attributed to sexual dimorphism and to different experimental protocols, such us different antagonists and different developmental periods of treatment.

Gonadotropin-Releasing Hormone: DISCUSSION(4)

Inasmuch as pulsatile GnRH is the major positive regulatory signal for GnRH-R gene expression, presumably most of the increment of GnRH-R mRNA levels throughout development is due to the progressive increase of endogenous GnRH secretion as young male rats approach sexual maturity. Our data show that the specific GnRH antagonist cetrorelix completely prevents the rise of levels of both GnRH-R gene transcripts during the infantile and juvenile periods.

Gonadotropin-Releasing Hormone: DISCUSSION(3)

DISCUSSION(3)

Pituitary weight increases 3- to 4-fold from infancy to puberty. Literature about the relative contribution of different cell populations to total pituitary cellular mass during ontogeny is scarce, but some studies pointed out that GnRH is mitogenic for gonadotrophs in vitro. Therefore, it is possible that populations of gonadotropes are relatively more represented in the anterior pituitary during puberty, contributing, at least in part, to the dramatic elevation of the gonadotrope mRNAs observed in the present work. In fact, anterior pituitaries of male rats treated with cetrorelix weigh ~15% less than controls (data not shown).

Gonadotropin-Releasing Hormone: DISCUSSION(2)

The ontogenetic patterns of gonadotropin p subunit gene expression are quite similar to those of the GnRH-R gene. However, the juvenile acceleration started several days later than the GnRH-R mRNA. This fact is not surprising taking into account that appropriate expression of GnRH receptors in gonadotropes may allow the subsequent expression of gonadotropin subunits in response to GnRH. Again, the progressive increase of serum testosterone, acting as negative feedback on GnRH secretion, may explain the decline of gonadotropin p subunit mRNAs that occurred after Day 35.

Gonadotropin-Releasing Hormone: DISCUSSION(1)

DISCUSSION(1)

To our knowledge, the present work shows, for the first time, the developmental changes in GnRH-R and gonadotropin subunit mRNA levels in male rats from Day 5 to sexual maturity. The GnRH-R mRNA levels increased progressively and markedly through the infantile and juvenile periods until the time of puberty. This increase is slow through the first part of the infantile period and accelerates during the second part of the infantile and during the juvenile period. This supports the idea that pubertal activation of GnRH-R gene expression is the major mechanism by which the gonadotrope cell increases the number of GnRH-R during sexual development.

Gonadotropin-Releasing Hormone: RESULTS(2)

Effect of the GnRH Antagonist Cetrorelix on the GnRH-R Gene Expression

To investigate whether maturational activation of the GnRH-R gene expression is GnRH dependent, developing male rats were treated either with vehicle or the GnRH antagonist cetrorelix during the infantile and juvenile stages of prepubertal development. As shown in Figure 4, cetro-relix treatment completely abolished the rise of the two major species of the GnRH-R mRNA during the two periods of the study. flovent inhalers

Effect of the GnRH Antagonist Cetrorelix on the Gonadotropin Subunit Gene Expression

In a similar way as occurred with the GnRH-R mRNA levels, cetrorelix treatment also prevented the rise of levels of FSHp and LHp subunit mRNAs (Fig. 5) during both the infantile and juvenile periods (FSHp mRNA, infantile period: control Day 5 = 4.30 ± 0.11 arbitrary densitometric units [ADU], control Day 20 = 9.70 ± 0.30 ADU, cetro-relix-treated = 0.5 ± 0.01 ADU, P < 0.001 vs. control Day 20; juvenile period: control Day 20 = 4.50 ± 0.34 ADU, control Day 35 = 12.5 ± 0.8 ADU, cetrorelix treated = 1.10 ± 0.10 ADU, P < 0.001 vs. control Day 35; and LHp mRNA, infantile period: control Day 5 = 4.1 ± 0.30 ADU, control Day 20 = 8.2 ± 0.28 ADU, cetrorelix treated = 0.5 ± 0.14 ADU, P < 0.001 vs. control Day 20; juvenile period: control Day 20 = 6.2 ± 0.10 ADU, control Day 35 = 13.50 ± 0.80, cetrorelix treated = 3.70 ± 0.10 ADU, P < 0.001 vs. control Day 35).

Gonadotropin-Releasing Hormone: RESULTS(1)

RESULTS(1)GnRH-R mRNA Levels

The two major species of GnRH-R mRNA, 5.0 and 4.5 kb, were present in the pituitaries of 5-day-old male pups. As shown in Figure 1, the levels of both transcripts increased very slowly through the infantile period. A more rapid rise occurred through the juvenile period to peak at Day 35. Thereafter, the levels of both transcripts declined slightly until Day 55. The other two much less abundant transcripts, 2.5 and 1.3 kb, were almost undetectable and therefore their levels were not quantitated.

Gonadotropin Subunit mRNA Levels

As shown in Figure 2, the a subunit mRNA was easily detected at Day 5 and its level increased slowly until Day 15, when it incremented faster up to Day 25; then a mild nonstatistically significant increase occurred to Day 35. Thereafter, the levels fell to Day 55. FSHp and LHp mRNA levels increased slowly until Day 25, when the levels of both mRNAs rose sharply to peak at Day 35 (Fig. 2). Then the levels of both mRNAs declined to Day 55.

Gonadotropin-Releasing Hormone: MATERIALS AND METHODS(4)

Serum Gonadotropin and Testosterone

Serum LH and FSH were determined by RIA using reagents generously supplied by Dr A.F. Parlow (National Hormone and Pituitary Program). LH-RP3 and FSH-RP2 were used as reference preparations. LH and FSH were iodinated by the chloramine T method, and specific RIAs were performed according to the procedure recommended by the National Institutes of Health. The intraassay variations were 8% and 7% for LH and FSH, respectively. Serum total and free testosterone were measured by RIA using commercial kits obtained from Diagnostic Products Corporation DPC (Los Angeles, CA). The intraassay variations for total testosterone assay were 5% for total testosterone concentrations of 100-400 ng/dl, 10% for concentrations of 50-100 ng/dl, and 15% for concentrations of 10-20 ng/dl.

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