Fig. 1.
The experimental paradigm of the quantitative mass spectrometry we employed to investigate the neuropeptide level changes in response to food intake. Rats fed on a regular diet or starved were all sacrificed 2.5 h after the feeding time. Rat brains were immediately dissected following decapitation and the dorsal striatum, hippocampus, hypothalamus and nucleus accumbens regions were collected as tissue punches, followed by rapid heating by DenatorTM to minimize postmortem degradation. The tissue samples were pooled as an aliquot to minimize individual variation and increase the amount of neuropeptides contained in samples. Four punches were pooled for NAc and hippo, three punches were pooled for HT and two punches were pooled for DS as an aliquot. The sample aliquots were then processed individually, and the endogenous neuropeptides were analyzed by an Orbitrap LC-MS/MS. The high accuracy MS and MS/MS data yielded by Orbitrap were searched against database for identification, whereas peak areas calculated using the extracted ion chromatograms of each identified peptide are employed for relative quantitation. The locations of the tissue punches we collected are shown on a sagittal slice of the rat brain.