Pyrylium based derivatization imaging mass spectrometer revealed the localization of L-DOPA

Abstract

Simultaneous imaging of L-dihydroxyphenylalanine (L-DOPA), dopamine (DA) and norepinephrine (NE) in the catecholamine metabolic pathway is particularly useful because L-DOPA is a neurophysiologically important metabolic intermediate. In this study, we found that 2,4,6-trimethylpyrillium tetrafluoroborate (TMPy) can selectively and efficiently react with target catecholamine molecules. Specifically, simultaneous visualization of DA and NE as metabolites of L-DOPA with high steric hinderance was achieved by derivatized-imaging mass spectrometry (IMS). Interestingly, L-DOPA showed strong localization in the brainstem, in contrast to the pattern of DA and NE, which co-localized with tyrosine hydroxylase (TH). In addition, to identify whether the detected molecules were endogenous or exogenous L-DOPA, mice were injected with L-DOPA deuterated in three positions (D3-L-DOPA), which was identifiable by a mass shift of 3Da. TMPy-labeled L-DOPA, DA and NE were detected at m/z 302.1, 258.1 and 274.1, while their D3 versions were detected at 305.0, 261.1 and 277.1 in mouse brain, respectively. L-DOPA and D3-L-DOPA were localized in the BS. DA and NE, and D3-DA and D3-NE, all of which are metabolites of L-DOPA and D3-L-DOPA, were localized in the striatum (STR) and locus coeruleus (LC). These findings suggest a mechanism in the brainstem that allows L-DOPA to accumulate without being metabolized to monoamines downstream of the metabolic pathway.