TY - JOUR
T1 - Gas chromatography/tandem mass spectrometry detection of extracellular kynurenine and related metabolites in normal and lesioned rat brain
AU - Notarangelo, Francesca M.
AU - Wu, Hui Qiu
AU - Macherone, Anthony
AU - Graham, David R.
AU - Schwarcz, Robert
N1 - Funding Information:
This study was supported in part by National Institutes of Health (NIH) grants AG022074, NS057715 and MH085554 . We thank Kyle Horning for excellent technical assistance.
PY - 2012/2/15
Y1 - 2012/2/15
N2 - We describe here a gas chromatography-tandem mass spectrometry (GC/MS/MS) method for the sensitive and concurrent determination of extracellular tryptophan and the kynurenine pathway metabolites kynurenine, 3-hydroxykynurenine (3-HK), and quinolinic acid (QUIN) in rat brain. This metabolic cascade is increasingly linked to the pathophysiology of several neurological and psychiatric diseases. Methodological refinements, including optimization of MS conditions and the addition of deuterated standards, resulted in assay linearity to the low nanomolar range. Measured in samples obtained by striatal microdialysis in vivo, basal levels of tryptophan, kynurenine, and QUIN were 415, 89, and 8 nM, respectively, but 3-HK levels were below the limit of detection (<2 nM). Systemic injection of kynurenine (100 mg/kg, i.p.) did not affect extracellular tryptophan but produced detectable levels of extracellular 3-HK (peak after 2-3 h: ∼50 nM) and raised extracellular QUIN levels (peak after 2 h: ∼105 nM). The effect of this treatment on QUIN, but not on 3-HK, was potentiated in the N-methyl-D-aspartate (NMDA)-lesioned striatum. Our results indicate that the novel methodology, which allowed the measurement of extracellular kynurenine and 3-HK in the brain in vivo, will facilitate studies of brain kynurenines and of the interplay between peripheral and central kynurenine pathway functions under physiological and pathological conditions.
AB - We describe here a gas chromatography-tandem mass spectrometry (GC/MS/MS) method for the sensitive and concurrent determination of extracellular tryptophan and the kynurenine pathway metabolites kynurenine, 3-hydroxykynurenine (3-HK), and quinolinic acid (QUIN) in rat brain. This metabolic cascade is increasingly linked to the pathophysiology of several neurological and psychiatric diseases. Methodological refinements, including optimization of MS conditions and the addition of deuterated standards, resulted in assay linearity to the low nanomolar range. Measured in samples obtained by striatal microdialysis in vivo, basal levels of tryptophan, kynurenine, and QUIN were 415, 89, and 8 nM, respectively, but 3-HK levels were below the limit of detection (<2 nM). Systemic injection of kynurenine (100 mg/kg, i.p.) did not affect extracellular tryptophan but produced detectable levels of extracellular 3-HK (peak after 2-3 h: ∼50 nM) and raised extracellular QUIN levels (peak after 2 h: ∼105 nM). The effect of this treatment on QUIN, but not on 3-HK, was potentiated in the N-methyl-D-aspartate (NMDA)-lesioned striatum. Our results indicate that the novel methodology, which allowed the measurement of extracellular kynurenine and 3-HK in the brain in vivo, will facilitate studies of brain kynurenines and of the interplay between peripheral and central kynurenine pathway functions under physiological and pathological conditions.
KW - 3-Hydroxykynurenine
KW - Microdialysis
KW - Quinolinic acid
KW - Selective reaction monitoring
KW - Tandem mass spectrometry
KW - Tryptophan
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U2 - 10.1016/j.ab.2011.12.032
DO - 10.1016/j.ab.2011.12.032
M3 - Article
C2 - 22239963
AN - SCOPUS:84862785742
SN - 0003-2697
VL - 421
SP - 573
EP - 581
JO - Analytical biochemistry
JF - Analytical biochemistry
IS - 2
ER -