17β-Estradiol levels in male zebra finch brain: Combining Palkovits punch and an ultrasensitive radioimmunoassay

Local aromatization of testosterone into 17β-estradiol (E2) is often required for the physiological and behavioral actions of testosterone. In most vertebrates, aromatase is expressed in a few discrete brain regions. While many studies have measured brain aromatase mRNA or activity, very few studies have measured brain E2 levels, particularly in discrete brain regions, because of technical challenges. Here, we used the Palkovits punch technique to isolate 13 discrete brain nuclei from adult male zebra finches. Steroids were extracted via solid phase extraction. E2 was then measured with an ultrasensitive, specific and precise radioimmunoassay. Our protocol leads to high recovery of E2 (84%) and effectively removes interfering brain lipids. E2 levels were high in aromatase-rich regions such as caudal medial nidopallium and hippocampus. E2 levels were intermediate in the medial preoptic area, ventromedial nucleus of the hypothalamus, lateral and medial magnocellular nuclei of anterior nidopallium, nucleus taeniae of the amygdala, and Area X. E2 levels were largely non-detectable in the cerebellum, HVC, lateral nidopallium and optic lobes. Importantly, E2 levels were significantly lower in plasma than in the caudal medial nidopallium. This protocol allows one to measure E2 in discrete brain regions and potentially relate local E2 concentrations to aromatase activity and behavior.