Optimization of neuropeptide extraction from the mouse hypothalamus

Sample preparation for neuropeptidomic studies is a critical issue since protein degradation can produce high levels of peptides that obscure the endogenous neuropeptides. We compared different extraction conditions for the recovery of neuropeptides and the formation of protein breakdown fragments from mouse hypothalami. Sonication and heating in water (70 degrees C for 20 min) followed by cold acid and centrifugation enabled the efficient extraction of many neuropeptides without the formation of protein degradation fragments seen with hot acid extractions. The hot water/cold acid extraction procedure resulted in the reproducible recovery of many hypothalamic peptides, including several novel peptides.