LC-MS/MS Quantification of Neurotransmitters

Tissue block containing CPu was weighed and homogenized in 80% acetonitrile containing 0.1% formic acid (5 μL) and then processed following a previous study [24 (link)]. The homogenates were centrifuged at 14,000g for 10 min at 4°C. The supernatants were collected and used to determine dopamine (DA), 3,4-dihydroxyphenylacetic acid (DOPAC), and homovanillic acid (HVA) levels as previously described [24 (link)]. LC separation was performed on an Agilent 1200 LC system (Agilent, Santa Clara, USA) using a Synergi Fusion-RP C18 column (50 mm × 3.0 mm, 4 μm) provided by Phenomenex. MS/MS detection was carried out using a 3200 QTRAP™ LC-MS/MS System (Applied Biosystems, Foster City, CA, USA). The multiple-reaction monitoring mode was used for quantification. The principal validation parameters of the LC-MS/MS are described in a previous study [24 (link)].