Unique estrogenic mechanisms for unique gonadotropin-releasing hormone neurons?

The history of studies of mechanisms of estrogen action in the brain is filled with at least two periods of intense dogmatic beliefs. These include thinking that a single type of cytoplasmic, unliganded estrogen receptor (ER) bound ligand and moved to the nucleus after activation (1), to thinking that all ERs were located in cell nuclei even in the unoccupied state (2). The dogma of cytoplasmic vs. nuclear distribution began to die in the early 1990s with the recognition of a more widespread intracellular distribution (3), and the single ER concept died later in the 1990s with the discovery of ER(4). The current state of affairs was well summarized recently (5, 6) as a situation in which a plethora of proteins localized from plasma membrane to nuclei has the potential to bind estrogens and initiate diverse signal transduction pathways. A history of studies of mechanisms governing the functioning of GnRH neurons has also broken through several dogmatic stretches, including the once-held belief that GnRH neurons contain few if any hormone receptors, particularly ERs (7). Nevertheless, the importance of hormone modulation of GnRH physiology (whether direct or indirect) has long been recognized. Despite a long history of studying estrogen effects on the hypothalamic-pituitary-gonadal axis and GnRH neurons, investigations of hormone influences on GnRH neurons have remained limited by an 800-lb gorilla in the room; the considerably heterogeneous GnRH neuronal network consists of relatively few cells widely distributed throughout the forebrain, making single-cell investigations a prodigious challenge. Significant progress on teasing out how GnRH neurons work can be traced first to the development of in vitro approaches to identifying GnRH neurons (e.g. Ref. 8), and then to the development of transgenic mice in which the GnRH promoter drives the expression of enhanced green fluorescent protein (9). These advances, together with others in molecular genetics and the use of immortalized GnRH neuronal cell lines, have allowed researchers to begin investigating hormone feedback to GnRH neurons with single cell resolution, notwithstanding challenges presented by their scattered topography. The march of progress now brings the ever-expanding field of ER signaling barreling down a track toward the field of GnRH neuronal function at the single cell level with ever increasing frequency. The crash, seemingly analogous to those produced in a particle accelerator, seems to throw out new “particles” with each collision. A recent report in Endocrinology (10) used an explant model derived from primate olfactory placodes to describe rapid increases in intracellular Ca oscillations mediated by estrogens signaling through a membrane/cytoplasmic ER. This is not the first report of Ca oscillations in GnRH neurons as the authors point out; however, the new data point to the G protein-coupled receptor, GPR30, as the source of the signaling rather than ERor . In the current issue of Endocrinology, two groups present further novel data on estrogenic influences on GnRH neuron physiology using brain slices and GnRH neurons identified by their expression of GFP (11) or a genetically encoded calcium indicator, pericam, under control of the GnRH promoter (12). Use of the calcium indicator revealed rapid effects of estradiol on calcium dynamics in murine GnRH neurons. Activation of intracellular calcium transients was observed approximately 15 min after treatment with 17-estradiol through a mechanism shown to be selective for intracellular ER as compared with a membrane located ER , ER , or GPR30. Interestingly, blocking action-potential dependent synaptic transmission with tetrodotoxin was not sufficient to abolish the ER-dependent transients, whereas tetrodotoxin treatment in the presence of a -aminobutyric acid-(GABA)A receptor antagonist completely inhibited stimulation of transients. Together, these experiments revealed a nonclassical estrogenic modulation of presynaptic -aminobutyric acid-ergic terminals that then impacts a subset of GnRH neurons. In a separate study, a novel combination of knockout and transgenic mice provided evidence that classical genomic ER -based estrogen response element (ERE)-dependent signaling is necessary for the proper regulation of negative and positive estrogen feedback at the level of the GnRH neuron (11). A line of mice was bred that expressed a knock-in allele that selectively restores a modified ER that can no longer bind to EREs on an ER knockout background that also carries the transgene for GnRH promoter driven enhanced green fluorescent protein expression. Using these mice the authors were able to differentiate ERE-dependent and ERE-independent mechanisms of GnRH neuron activation. GnRH neuron firing rates were altered in brain slices from mice placed in situations of either positive or negative feedback in ER knockout mice, or in ER mice with selectively restored ERE-independent signaling. These data suggest that classical negative and positive feedback is dependent on genomic ER interactions with ERE, although the cellular locus of estrogenic signaling was not determined. Because the manipulated ER was the -form, the cellular locus is presumed to be other than GnRH neurons themselves. Together, these studies provide additional evidence that a variety of estrogenic mechanisms may influence GnRH neurons. The authors of these papers (10–12) eloquently review past history in the field that is directly relevant to their results. In this commentary we take a step back and look at the emergSee articles p. 5328 and 5335. Abbreviations: ER, Estrogen receptor; ERE, estrogen response element.