ABSTRACT
L-Serine-O-phosphate (L-SOP), the precursor of l-serine, is an agonist at group III metabotropic glutamate receptors. Despite the interest in L-SOP, very few articles have reported its brain levels. Here we report a convenient and reproducible method for simultaneous analysis of L-SOP and several other important amino acids in brain tissue using high-performance liquid chromatography (HPLC) with fluorimetric detection after derivatization with o-phthaldialdehyde and N-isobutyl-L-cysteine. Analyses were carried out in rat whole brain and cerebellum and in mouse whole brain, forebrain, amygdala, and prefrontal cortex. The method should be useful for future comprehensive neurochemical and pharmacological studies on neuropsychiatric disorders.
Subject(s)
Brain Chemistry , Chromatography, High Pressure Liquid/methods , Fluorometry/methods , Phosphoserine/analysis , Animals , Brain/metabolism , Cerebellum , Mice , Mice, Inbred C57BL , Prefrontal Cortex/chemistry , Rats , Rats, Sprague-Dawley , Serine/metabolismABSTRACT
Phenelzine (PLZ), a nonselective irreversible inhibitor of monoamine oxidase (MAO), also inhibits GABA-transaminase (GABA-T), markedly increasing brain GABA levels. PLZ is also a substrate for MAO, and studies suggest that a metabolite formed by the action of this enzyme on PLZ may be responsible for the increase in GABA observed. We have recently found that PLZ also elevates brain ornithine (ORN), an amino acid precursor to both glutamate (and GABA) and the polyamines, and have conducted dose- and time-response studies on this effect. Rats were treated with vehicle or PLZ doses (7.5, 15 or 30 mg/kg i.p.), and brains were collected 3 h later. In the time-response study, animals were treated with vehicle or PLZ (15 mg/kg i.p.) and brains were collected 1-24 h later. To determine whether a metabolite formed by the action of MAO on PLZ may be responsible for the elevation in brain ORN observed, animals were pretreated with vehicle or the MAO inhibitor tranylcypromine (TCP) before vehicle or PLZ (15 mg/kg), and brains collected 3 h later. ORN levels (measured by an HPLC procedure) were dose- and time-dependently increased in PLZ-treated animals, with levels reaching approximately 650% of control at 6 and 12 h. Pretreatment with TCP completely abolished the PLZ-induced increase in brain ORN, suggesting, as with GABA, that a metabolite of PLZ formed by the action of MAO is responsible for the elevation of brain ORN observed. The possible contribution of increased ORN to therapeutic and/or neuroprotective properties of PLZ is discussed.
Subject(s)
Brain Chemistry/drug effects , Monoamine Oxidase Inhibitors/pharmacology , Ornithine/metabolism , Phenelzine/antagonists & inhibitors , Phenelzine/pharmacology , Animals , Dose-Response Relationship, Drug , Male , Rats , Rats, Sprague-Dawley , Time Factors , Tranylcypromine/pharmacology , gamma-Aminobutyric Acid/metabolismABSTRACT
A simple and versatile methodology using high-performance liquid chromatography (HPLC) with fluorimetric detection was developed to simultaneously determine d-serine along with other metabolically related neuroactive amino acids in the glutamatergic system: L-serine, L-glutamate, L-glutamine, and glycine. On-column sensitivity was in the lower picomole range. Of two chiral thiol reagents investigated, amino acid derivatives of o-phthaldialdehyde (OPA) in combination with N-isobutyryl-L-cysteine were found to have consistently higher responses than their corresponding N-tert-butyloxycarbonyl-L-cysteine derivatives. This methodology was applied to the quantitative detection of amino acids in human plasma and lays the foundation for further investigations of the role of neuroactive amino acids in the pathophysiology and treatment of neurological and psychiatric disorders.
