Estradiol induction of spermatogenesis is mediated via an estrogen receptor-{alpha} mechanism involving neuroendocrine activation of follicle-stimulating hormone secretion

Abstract

Both testosterone and its nonaromatizable metabolite dihydrotestosterone (DHT) induce spermatogenesis in gonadotropin-deficient hpg mice. Surprisingly, because aromatization is not required, estradiol (E2) also induces spermatogenesis and increases circulating FSH in hpg mice, but the mechanism remains unclear. We studied E2-induced spermatogenesis in hpg mice on an estrogen receptor (ER)-alpha (hpg/alphaERKO) or ERbeta (hpg/betaERKO) knockout or wild-type ER (hpg/WT) background treated with subdermal E2 or DHT implants for 6 wk. In hpg/WT and hpg/betaERKO, but not hpg/alphaERKO mice, E2 increased testis and epididymal weight, whereas DHT-induced increases were unaffected by ERalpha or ERbeta inactivation. E2 but not DHT treatment increased serum FSH (but not LH) in hpg/WT and hpg/betaERKO but not hpg/alphaERKO hpg mice. DHT or E2 alone increased (premeiotic) spermatogonia and (meiotic) spermatocytes without significant change in Sertoli cell numbers. DHT alone increased postmeiotic spermatids, regardless of ER presence, compared with variable ERalpha-dependent E2 postmeiotic responses. An ERalpha-mediated effect was confirmed by treating hpg mice for 6 wk by subdermal selective ER-alpha (16alpha-LE(2)) or ERbeta (8beta-VE(2)) agonist implants. ERalpha (but not ERbeta) agonist increased testis and epididymal weight, Sertoli cell, spermatogonia, meiotic, and postmeiotic germ cell numbers. Only ERalpha agonist markedly increased serum FSH, whereas either agonist induced small rises in serum LH. Administration of ERalpha agonist or E2 in the presence of functional ERalpha induced prominent gene expression of specific Sertoli (Eppin, Rhox5) and Leydig cell (Cyp11a1, Hsd3b1) markers. We conclude that E2-induced spermatogenesis in hpg mice involves an ERalpha-dependent neuroendocrine mechanism increasing blood FSH and Sertoli cell function.

Figure 1

Testis (upper left), seminal vesicle (upper middle), and anterior prostate lobe (upper right) weights together with pituitary (lower left), serum FSH (lower middle), and LH (lower right) panels in hpg mice treated with low or high doses of a selective ERα (black bars) or selective ERβ (gray bars) agonist or untreated (white bars). Data expressed as mean and sem. For further details, see text.

Figure 2

Sertoli (SC) and germ cell population of the testis in age-matched littermate hpg mice that were untreated (upper panel) or treated with a selective ERα agonist (16α-LE₂, 45 μg/kg · d for 6 wk; middle panel) or a selective ERβ agonist (8β-VE₂, 450 μg/kg · d for 6 wk; right panel) for 6 wk. Data expressed as mean and sem. At the right is representative testicular histology from age-matched littermate hpg mice that were untreated (upper panel) or treated with the selective ERα agonist (middle panel) or the selective ERβ agonist (right panel). Note elongated spermatids (Sp) present in tubules of hpg mice treated with selective ERα but not ERβ agonist. All photomicrographs are at the same magnification. For further details, see text.

Figure 3

Testis (upper left), epididymis (upper middle), and seminal vesicles (upper right) together with serum FSH (lower left) and LH (lower right) in hpg mice on wild-type (hpg/WT, white bars), ERα knockout (hpg/αERKO, black bars), or ERβ knockout (hpg/βERKO, gray bars) genetic backgrounds. Data expressed as mean and sem. For further details, see text.

Figure 4

Testicular histology from age-matched littermate hpg mice on different genetic backgrounds comprising WT (left panels), αERKO (middle panels), and βERKO (right panels). The top row depicts the untreated, the middle row treated with E2, and the bottom row treated with DHT for 6 wk. Note elongated spermatids (Sp) indicated by arrows present in tubules of hpg mice treated with DHT or E2 for the mice on WT and βERKO but not αERKO background. Well-formed seminiferous tubular lumen (indicated by L) is present in all mice treated with DHT but not E2 or untreated. All photomicrographs are at the same magnification. For further details, see text.

Figure 5

Total testis Rhox5, Eppin, Hsd3b1, and Cyp11a1 mRNA expression levels. Data are shown as mean and sem for treatments (n = 4–10/group) including untreated (−) and ERβ and ERα agonists (left panel, study 1) in hpg mice and untreated (−), E2, or DHT in hpg/WT, hpg/αERKO, and hpg/βERKO mice (right three panels, study 2) along the x-axis. Gene expression levels (y-axis) were measured for two Sertoli cell markers Rhox5 and Eppin (upper two panels) and two Leydig cell markers Cyp11a1 and Hsd3b1 (lower two panels) by real-time qPCR analysis using whole-testis cDNA. The measured mRNA transcript levels are adjusted for Wbscr1 (housekeeping gene as loading control) and for testis weight (to account for dilution by germ cells) as detailed in text.

Figure 6

Magnitude of DHT effect in reproductive and nonreproductive organs in hpg mice on βERKO vs. WT (i.e. functional ERα and ERβ) genetic background. The magnitude of the DHT effect, defined as the increase in organ or tissue weight of the DHT-treated compared with the untreated controls on the same genetic background, with the DHT effect on βERKO background expressed as a percentage of DHT effect on the WT background (set at 100%). Data expressed as mean and sem. For further details, see text.