Elsevier

Neuroscience

Volume 154, Issue 4, 17 July 2008, Pages 1173-1177
Neuroscience

Rapid report
Estrogen receptor α is expressed on the cell-surface of embryonic hypothalamic neurons

https://doi.org/10.1016/j.neuroscience.2008.05.001Get rights and content

Abstract

Although the biological activity of estrogen is generally mediated through nuclear estrogen receptors, a large body of evidence indicates that estrogen may also affect target cells upon binding to putative membrane estrogen receptors (mER) coupled to intracellular signaling cascades; however, no agreement has been reached on the nature and precise location of the putative estrogen receptor (ER) responsible for these rapid effects. In the present report we show that the expression of ERα is associated with the plasma membrane fraction of rat hypothalamic tissue at embryonic day 16. Moreover, our experiments extend these results to rat hypothalamic neurons in vitro showing that ERα can be detected from the cell exterior as a biotinylated cell-surface protein. We have also shown that the mERα is under regulation of estradiol, and the ERα agonist, 4,4′,4″-(4-propyl-[1H]-pyrazole-1,3,5-triyl)trisphenol, induced extracellular-signal-regulated kinase signaling in a dose-dependent manner and in a time-course not compatible with genomic actions, supporting the notion of a membrane-initiated phenomenon.

Section snippets

Experimental procedures

All experimental procedures were approved by the local Committee on Bioethics and followed the NIH Guidelines for the Care and Use of Laboratory Animals. All efforts were made to minimize both the suffering and the number of animals used. Unless otherwise specified, all drugs were from Sigma-Aldrich, St. Louis, MO, USA.

Identification of membrane ER from hypothalamic tissue

To determine the presence of ERs at the plasma membrane of E16 hypothalamic tissue we used sub-cellular fractionation and immunoblotting. The plasma membrane fraction was isolated using Percoll gradient centrifugation; absence of nuclear and cytosolic contamination was corroborated with anti-histone H1 and anti-LIMK-1 antibodies, respectively. As membrane purification control, incubation with a specific anti-TrkB revealed, as expected, a 145 kDa band corresponding to membrane receptor. Results

Conclusion

In summary, we have identified an ERα variant in the plasmalemmal fraction of hypothalamic tissue and on the cell-surface of hypothalamic neurons in cultures. We have also shown that the mERα is under E2-regulation and mediates E2 induced-ERK phosphorylation in a dose-dependent manner and in a time-course compatible with a membrane effect rather than a genomic action. Although the mechanisms of E2 translocation to the neuronal cell-surface will have to be addressed, the evidence presented in

Acknowledgments

This work was supported by grants from CONICET-PIP 6238 (M.J.C.) and ANPCyT-PICT 05-26331 (M.J.C.) and PICT 05-1429 (A.G.L.). S.V.G. is a CONICET fellow; A.G.L. and M.J.C. are career members of CONICET. We thank Dr. L. Heredia for technical advice on biotinylation experiments. M.J.C. is also indebted to Dr. H. F. Carrer for helpful discussions and for critical reading of the manuscript.

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