Detection and localization of an estrogen receptor beta splice variant protein (ERβ2) in the adult female rat forebrain and midbrain regions

Estrogens regulate neural processes such as neuronal development, reproductive behavior, and hormone secretion, and signal through estrogen receptor (ER) α and ERβ (here called ERβ1). Recent studies have found variations in ERα and ERβ1 mRNA splicing in rodents and humans. Functional reporter gene assays suggest that these splicing variations alter ER-mediated transcriptional regulation. Estrogen receptor beta 2 (ERβ2), an ERβ1 splice variant containing an 18 amino acid (AA) insert in the ligand binding domain, binds estradiol with ≈ 10-fold lower affinity than ERβ1, suggesting that it may serve as a low-affinity ER. Moreover, ERβ2 reportedly acts in a dominant-negative fashion when heterodimerized with ERβ1 or ERα. To explore the function of ERβ2 in brain, an antiserum (TwoβER.1) targeting the 18 AA insert was developed and characterized. Western blot analysis and transient expression of ERβ2 in cell lines demonstrated that TwoβER.1 recognizes ERβ2. In the adult female rat brain, ERβ2 immunoreactivity is localized in the cell nucleus and is expressed with a distribution similar to that of ERβ1 mRNA. ERβ2 immunoreactive cell numbers were high in, for example, piriform cortex, paraventricular nucleus, supraoptic nucleus, arcuate nucleus, and hippocampal CA regions, whereas it was low in the dentate gyrus. Moreover, ERβ2 is coexpressed in gonadotropin-releasing hormone and oxytocin neurons. These studies demonstrate ERβ splice variant proteins in brain and support the hypothesis that ER signaling diversity depends not only on ligand or coregulatory proteins, but also on regional and phenotypic selectivity of ER splice variant proteins.