Characterization of the G‐protein‐coupled membrane‐bound estrogen receptor GPR30 in the zebra finch brain reveals a sex difference in gene and protein expression

Abstract

Estradiol-induced structural dimorphisms exist in the songbird brain. However, how they arise is not clear since there is a scarce distribution of ERα and lack of ERβ in song control nuclei. This suggests that other receptors are involved. The G-protein coupled membrane-bound estrogen receptor, GPR30, is a candidate but has never been investigated in songbirds. In this study, we characterized its gene and protein in the zebra finch brain. Analysis of the putative GPR30 protein sequence revealed a strong similarity to avian and mammalian homologues. Quantitative PCR indicated that the gene was elevated in the telencephalon of both sexes from posthatching day (P) 15 to P45, with a male-biased sex difference at P21 and P30. In comparison, expression at younger posthatching ages and in adults was significantly less. At P21, GRP30 protein was widespread, nonuniform, and overlapped with song control nuclei. Of particular interest, the number of immunoreactive cells was greatest in HVC and RA, but less in LMAN and Area X. Labeling in HVC was also dimorphic; with more cells present in males than in females. In parallel with the gene, by adulthood, protein expression was reduced across most brain regions. Taken together these data suggest that GPR30 may contribute to differences in song system development by mediating dimorphic responses to estrogens. In addition, the extensive protein distribution indicates that it may also have a role in general brain development in both sexes.

Sex differences and rapid estrogen signaling: A look at songbird audition

“…Selective agonists for both classical receptors have not been able to reproduce the rapid auditory evoked effects of E2 (Remage-Healey et al , 2013), suggesting that other membrane receptors (ex: GPER1 or ER-X) could control this signaling. GPER1 expression is sexually dimorphic in zebra finch telencephalon around the critical period of song-learning; however, this sex differences disappears by adulthood (Acharya & Veney, 2012). …”

The importance of neural aromatization in the acquisition, recall, and integration of song and spatial memories in passerines

“…Rapid E 2 -dependent changes in intracellular signal transduction pathways in postsynaptic cells (McEwen and Alves, 1999;Akama and McEwen, 2003) support the idea that synaptic aromatase may be involved in use-dependent synaptic physiology and raises the possibility that synaptic E 2 may function more like a neurotransmitter than a neuromodulator Mukai et al, 2006;Saldanha et al, 2011). This mode of signal transduction is strongly supported by the localization of fixed-membrane estrogen receptors at extra-nuclear sitesincluding dendritic spines -in the songbird (Acharya and Veney, 2012) and rodent (Milner et al, A C C E P T E D M A N U S C R I P T…”

“…In zebra finches, aromatase is abundant in somata, dendritic arbors, and terminals in NCM (Saldanha et al, 1999;Saldanha et al, 2000;Peterson et al, 2005), and estrogen receptors are expressed throughout the region (Balthazart et al, 1992;Bernard et al, 1999;Metzdorf et al, 1999;Acharya and Veney, 2012). A variety of studies have examined the role of E 2 manipulation on song preference, immediate early gene activity, and/or the neurophysiological activity of NCM cells in male and female zebra finches as well as seasonally-breeding birds (reviewed in Cornil et al, 2012;Maney and Pinaud, 2012;De Groof et al, 2013).…”

“…For example, in the zebra finch hippocampus, males have more aromatase-positive presynaptic boutons and a higher proportion of synapses that express aromatase (Peterson et al, 2005), and in general there are more cells expressing membranebound G-protein coupled estrogen receptors in males than in females (Acharya and Veney, 2012). Interestingly, in tests of spatial memory, there are no sex differences in acquisition, but males perform better than females at retention trials over time as measured via latency to remember the location of a cup containing seed (Figure 2; Bailey et al, 2009).…”

“…Regions of the song control circuit contain abundant estrogen receptors (Gahr and Metzdorf, 1997;Metzdorf et al, 1999;Acharya and Veney, 2012) and all of the enzymes necessary for steroid hormone synthesis from cholesterol (Shen et al, 1994;Shen et al, 1995;Saldanha and Coomaralingam, 2005;Saldanha et al, 2000;London et al, 2006). HVC has long been described as responsive to E 2 during development and adulthood (Gurney and Konishi, 1980;Gurney, 1981;Walters and Harding, 1988;Casto and Ball, 1996;Oberlander et al, 2004).…”