LC-MS/MS Quantification of Neuroestrogens in Song Sparrow Brain Reveals Seasonal Regulation Independent of Aggression

Neuroestrogens regulate social behavior and cognition and are synthesized in the brain. In the song sparrow (Melospiza melodia), 17β-estradiol (17β-E2) promotes aggression, even during the non-breeding season, when circulating 17β-E2 levels are non-detectable. Measuring estrogen is challenging due to its low concentrations and the limited sensitivity of many existing assays. Moreover, many types of estrogen, other than 17β-E2, are often overlooked. Responding to this challenge, University of British Columbia (Vancouver, Canada) researchers developed a liquid chromatography tandem mass spectrometry (LC-MS/MS) assay for the simultaneous measurement of 11 types of estrogen using derivatization with 1,2-dimethylimidazole-5-sulfonyl-chloride (DMIS) to enhance sensitivity. A paper based on their work has been published in PLOS One (1).

A well-established songbird model for studying the steroid regulation of aggression under natural conditions, song sparrows in the Pacific Northwest breed from March through August, with males exhibiting robust territorial aggression during both the breeding and non-breeding seasons, despite the lack of circulating androgens and estrogens in the non-breeding season (2,3). Estrogen plays a critical role in their physiology and behavior, with 17β-estradiol (17β-E₂), the most widely studied estrogen, being traditionally associated with female reproductive behavior, but also the modulation of memory, attention, executive functions, and multiple social behaviors in both sexes (4-10).

The researchers conducted their analysis on free-living adult male song sparrows during both the breeding season (May 8–24, 2019) and non-breeding season (November 2-December 2, 2019). A total of 45 individuals were used: 5 for assay development and validation and 40 for method application. The 11 types of estrogen studied (E₁, 17β-E₂, 17α-E₂, E₃, 4OH-E₂, 2Me-E₂, 4Me-E₂, 16α-OH-E₁, 2OH-E₁, 2Me-E₁, and 4Me-E₁) were extracted from brain tissue or blood samples. As expected, there was large regional variation in neuroestrogen levels and very low estrogen levels in blood. There was also large seasonal variation, and estrogen levels were lower in the non-breeding season. Despite robust aggression to simulated territorial intrusion (STI), estrogen levels did not differ between STI and control subjects in either season (1).

The authors state that their approach “improves upon previous methods, by correcting for matrix effects and incorporating additional analytes to the panel. Application of the method to free-living male song sparrows exposed to an acute social challenge did not show changes in neuroestrogen levels, despite a strong behavioral response. Our refined method offers a powerful tool for quantifying estrogens in brain, with potential applicability across diverse vertebrate systems” (1).

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References

1. Jalabert, C.; Liu, M. Q.; Soma, K. K. Estrogen Profiling in Blood and Brain: Effects of Season and an Aggressive Interaction in a Songbird. PLoS One 2025, 20 (12), e0326727. DOI: 10.1371/journal.pone.0326727
2. Arcese, P. Territory Acquisition and Loss in Male Song Sparrows. Anim. Behav. 1989, 37, 45–55. DOI: 10.1016/0003-3472(89)90006-7
3. Jalabert, C.; Gray, S. L.; Soma, K. K. An Aggressive Interaction Rapidly Increases Brain Androgens in a Male Songbird during the Non-breeding Season. J. Neurosci. 2024, 44 (23), e1095232024. DOI: 10.1523/JNEUROSCI.1095-23.2024
4. Shanmugan, S.; Epperson, C. N. Estrogen and the Prefrontal Cortex: Towards a New Understanding of Estrogen’s Effects on Executive Functions in the Menopause Transition. Hum. Brain Mapp. 2014, 35 (3), 847–865. DOI: 10.1002/hbm.22218
5. Gervais, N. J.; Remage-Healey, L.; Starrett, J. R. et al. Adverse Effects of Aromatase Inhibition on the Brain and Behavior in a Nonhuman Primate. J. Neurosci. 2019, 39 (5), 918–928. DOI: 10.1523/JNEUROSCI.0353-18.2018
6. Tinkler, G. P.; Voytko, M. L. Estrogen Modulates Cognitive and Cholinergic Processes in Surgically Menopausal Monkeys. Prog. Neuropsychopharmacol Biol. Psychiatry 2005, 29 (3), 423–431. DOI: 10.1016/j.pnpbp.2004.12.016
7. Rosvall, K. A.; Bentz, A. B.; George, E. M. How Research on Female Vertebrates Contributes to an Expanded Challenge Hypothesis. Horm. Behav. 2020, 123, 104565. DOI: 10.1016/j.yhbeh.2019.104565
8. Quintana, L.; Jalabert, C.; Fokidis, H. B. et al. Neuroendocrine Mechanisms Underlying Non-breeding Aggression: Common Strategies Between Birds and Fish. Front. Neural Circuits 2021, 15, 716605. DOI: 10.3389/fncir.2021.716605
9. Jalabert, C.; Munley, K. M.; Demas, G. E. et al. Aggressive Behavior; in Encyclopedia of Reproduction. Elsevier. 2018, pp. 242–247. DOI: 10.1016/b978-0-12-801238-3.64591-9
10. Plumier, J. P.; Jalabert, C.; Munley, K. M. et al. Aggressive Behavior; in Reference Module in Biomedical Sciences. Elsevier. 2024. DOI: 10.1016/b978-0-443-21477-6.00303-5