Estrogen Receptor β Activation Rapidly Modulates Male Sexual Motivation through the Transactivation of Metabotropic Glutamate Receptor 1a

Abstract

In addition to the transcriptional activity of their liganded nuclear receptors, estrogens, such as estradiol (E2), modulate cell functions, and consequently physiology and behavior, within minutes through membrane-initiated events. The membrane-associated receptors (mERs) underlying the acute effects of estrogens on behavior have mostly been documented in females where active estrogens are thought to be of ovarian origin. We determined here, by acute intracerebroventricular injections of specific agonists and antagonists, the type(s) of mERs that modulate rapid effects of brain-derived estrogens on sexual motivation in male Japanese quail. Brain aromatase blockade acutely inhibited sexual motivation. Diarylpropionitrile (DPN), an estrogen receptor β (ERβ)-specific agonist, and to a lesser extent 17α-estradiol, possibly acting through ER-X, prevented this effect. In contrast, drugs targeting ERα (PPT and MPP), GPR30 (G1 and G15), and the Gq-mER (STX) did not affect sexual motivation. The mGluR1a antagonist LY367385 significantly inhibited sexual motivation but mGluR2/3 and mGluR5 antagonists were ineffective. LY367385 also blocked the behavioral restoration induced by E2 or DPN, providing functional evidence that ERβ interacts with metabotropic glutamate receptor 1a (mGluR1a) signaling to acutely regulate male sexual motivation. Together these results show that ERβ plays a key role in sexual behavior regulation and the recently uncovered cooperation between mERs and mGluRs is functional in males where it mediates the acute effects of estrogens produced centrally in response to social stimuli. The presence of an ER-mGluR interaction in birds suggests that this mechanism emerged relatively early in vertebrate history and is well conserved. Significance statement: The membrane-associated receptors underlying the acute effects of estrogens on behavior have mostly been documented in females, where active estrogens are thought to be of ovarian origin. Using acute intracerebroventricular injections of specific agonists and antagonists following blockade of brain aromatase, we show here that brain-derived estrogens acutely facilitate male sexual motivation through the activation of estrogen receptor β interacting with the metabotropic glutamate receptor 1a. This behavioral effect occurring within minutes provides a mechanistic explanation of how an estrogen receptor not intrinsically coupled to intracellular effectors can signal from the membrane to govern behavior in a very rapid fashion. It suggests that different subtypes of estrogen receptors could regulate the motivation versus performance aspects of behavior.

Keywords: ERβ; estrogens; membrane-initiated effects; sexual motivation.

Figure 1.

Time course of aromatase inhibition and acute E₂ action on male sexual motivation. A, Blockade of local estrogen synthesis by the aromatase inhibitor VOR (50 μg) inhibits RCSM frequency within 30–240 min. B, Experimental design used to test the time course of acute E₂ action (50 μg) on behavior acutely inhibited by aromatase blockade. Black arrows indicate how long E₂ was injected before 15 min, 2 h, or 4 h testing conditions; the gray arrow indicates how long E₂ was injected before the 5 min test condition. C–F, The behavioral inhibition resulting from acute estrogen deprivation is prevented by E₂ (50 μg) injected 15 min (D) before the test, but not 5 (C), 120 (E), and 240 min (F) before the test. The Pre and Post black bars provide reference behavior frequencies after vehicle intracerebroventricular injections performed before and after the acute treatments, but these data are not included in the statistical analyses. Δp < 0.05, ΔΔp < 0.01, and ΔΔΔp < 0.001 versus vehicle (Veh) and ##p < 0.01 versus VOR 30 min by Newman–Keuls post hoc tests after identification of a significant overall treatment effect by two-way ANOVA (repeated measure; n = 13). H, The behavioral inhibition resulting from acute neuroestrogen depletion is prevented by E₂ injected 15 min but not after 2 or 4 h before. **p < 0.01 by Newman–Keuls post hoc tests following a significant treatment effect by one-way ANOVA (repeated measure; n = 12) on data expressed as percentage of VOR condition. G, Representation of individual values comparing the effect of E₂ after 15 min to the positive (Veh) and negative (VOR) controls .

Figure 2.

The acute intracerebroventricular injection of a bolus of E₂ (50 μg, n = 7) without concurrent depletion in neuroestrogens does not affect male sexual motivation. The Pre and Post black bars provide reference behavior frequencies after vehicle intracerebroventricular injections performed before and after the acute treatments, but these data are not included in the statistical analyses.

Figure 3.

Effects of acute activation of ERα and ERβ on the frequency of RCSMs. A–E, Blockade of local estrogen synthesis by the aromatase inhibitor VOR (50 μg) inhibits RCSM frequency within 30 min. The behavioral inhibition induced by acute estrogen deprivation is prevented by the highest doses of the specific agonist of ERβ, DPN (2, 10, and 50 μg; B–D), but not its lowest dose (0.4 μg; A). Similarly, the administration of both PPT and DPN prevents the behavioral inhibition induced by VOR (E). However, the ERα agonist PPT alone has no effect at any of the doses tested (A–E). F, The ERα antagonist has no effect on behavioral frequencies. The Pre and Post black bars provide reference behavior frequencies after vehicle intracerebroventricular injections performed before and after the acute treatments, but these data are not included in the statistical analyses. Δp < 0.05, ΔΔp < 0.01, and ΔΔΔp < 0.001 versus vehicle (Veh); *p < 0.05, **p < 0.01, and ***p < 0.001 versus VOR; #p < 0.05, ##p < 0.01, and ###p 0.001 versus VOR + PPT by Newman–Keuls post hoc tests after identification of a significant treatment effect (repeated measure; A, B, D, n = 10; C, n = 12; E, n = 9; F, n = 9) by two-way ANOVA.

Figure 4.

The acute activation of GPR30, Gq-mER, and ER-X has no effect on the frequency of RCSMs. A, C, D, Blockade of local estrogen synthesis by the aromatase inhibitor VOR (50 μg) inhibits RCSM frequency within 30 min. G1, the specific agonist of GPR30 (50 μg, n = 12, A), injected 15 min after VOR, did not counteract the effect of VOR on RCSMs, nor did STX, the specific agonist of Gq-mER (50 μg, n = 11, C), and 17α-E₂, a potential agonist of ER-X (50 μg, n = 9, D). B, No change in RCSM frequency was observed 30 min after the injection of G15, the specific antagonist of GPR30 (50 μg, n = 10). The Pre and Post black bars provide reference behavior frequencies after vehicle intracerebroventricular injections performed before and after the acute treatments, but these data are not included in the statistical analyses. Δp < 0.05, ΔΔΔp < 0.001 versus vehicle (Veh) by Newman–Keuls post hoc tests after identification of a significant treatment effect (repeated measure) by two-way ANOVA.

Figure 5.

Effects of acute blockade of mGluRs on the frequency of RCSMs. A–C, The specific antagonist of mGluR1a (LY367385) inhibits RCSM frequency (100 μg, n = 12, A) within 30 min, while specific antagonists of mGluR2/3 (LY341495) and mGluR5 (MPEP) have no effect on RCSM within 30 min (100 μg, n = 9 and 8 respectively, B, C). D, The combined effect of mGluR1a blockade and E₂ was tested following acute blockade of brain aromatase using the described experimental design. E, F, Blockade of local estrogen synthesis by the aromatase inhibitor VOR (50 μg) inhibits RCSM frequency within 30 min. E₂ or DPN (50 μg; E, F) injected 15 min after VOR prevents the effect of VOR on RCSMs. LY367385 (100 μg) prevents the restoration of behavior by E₂ (E, n = 22) or DPN (F, n = 15). The Pre and Post black bars provide reference behavior frequencies after vehicle intracerebroventricular injections performed before and after the acute treatments, but these data are not included in the statistical analyses. ΔΔΔp < 0.001 versus vehicle (Veh) by Fisher's LSD post hoc test (A) and Newman–Keuls post hoc test; ***p < 0.001 versus VOR; ###p < 0.001 versus VOR + E₂ or VOR + DPN by Newman–Keuls post hoc tests after identification of a significant treatment effect (repeated measure) by two-way ANOVA.