J. Neurochem. (2010) 114, 1745–1755. Abstract Catechol‐O‐methyltransferase (COMT) plays an active role in the metabolism of dopamine (DA) in the prefrontal cortex (PFC). Because of low levels of dopamine transporter (DAT), it is proposed that the majority of released DA is taken up by either norepinephrine transporter (NET) and subsequently metabolized by monoamine oxidize (MAO) or by uptake2 (to glial cells and post‐synaptic neurons) and metabolized by COMT. However, a comprehensive in vivo study of rating the mechanisms involved in DA clearance in the PFC has not been done. Here, we employ two types of microdialysis to study these pathways using DAT, NET and MAO blockers in conscious mice, with or without Comt gene disruption. In quantitative no‐net‐flux microdialysis, DA levels were increased by 60% in the PFC of COMT‐knockout (ko) mice, but not in the striatum and nucleus accumbens. In conventional microdialysis studies, we showed that selective NET and MAO inhibition increased DA levels in the PFC of wild‐type mice by two‐ to fourfold, an effect that was still doubled in COMT‐ko mice. Inhibition of DAT had no effect on DA levels in either genotype. Therefore, we conclude that in the mouse, PFC COMT contributes about one half of the total DA clearance.

A liquid chromatographic-electrospray/tandem mass spectrometric (LC-ESI-MS/MS) method was developed for the analysis of dopamine and its phase I and phase II metabolites from brain microdialysis samples. The method provides for the first time the analysis of intact dopamine glucuronide and sulfate without hydrolysis. The paper describes also an enzymatic synthesis method using rat liver microsomes as biocatalysts and characterization of dopamine glucuronide as a reference compound. The method was validated for quantitative analysis by determining limits of detection and quantitation, linearity,repeatability, and specificity. Dopamine glucuronide was found for the first time in rat and mouse brain microdialysis samples. The concentrations of dopamine and its glucuronide in the microdialysates collected from the striatum of rat brains were approximately equal (2 nM).Dopamine sulfate was not detected in the microdialysates(limit of detection 0.8 nM). The main metabolites of dopamine were dihydroxyphenylacetic acid (DOPAC,1200 nM) and homovanillic acid (HVA, 700 nM).

Background and purpose: Catechol-O-methyltransferase (COMT) metabolizes compounds containing catechol structures and has two forms: soluble (S-COMT) and membrane-bound (MB-COMT). Here we report the generation of a mouse line that expresses MB-COMT but not S-COMT. We compared the effects of deleting S-COMT only or both COMT forms on the pharmacokinetics of oral L-DOPA. Experimental approach: L-DOPA (10 mg·kg ) were given to mice by gastric tube, and samples were taken at various times. HPLC was used to measure L-DOPA in plasma and tissue samples, and dopamine and its metabolites in brain. Immunohistochemistry and Western blotting were used to characterize the distribution of COMT protein isoforms. Key results: Lack of S-COMT did not affect the levels of L-DOPA in plasma or peripheral tissues, whereas in the full COMT-knock-out mice, these levels were increased. The levels of 3-O-methyldopa were significantly decreased in the S-COMT-deficient mice. In the brain, L-DOPA levels were not significantly increased, and dopamine was increased only in females. The total COMT activity in the S-COMT-deficient mice was 22-47% of that in the wild-type mice. In peripheral tissues, female mice had lower COMT activity than the males. Conclusions and implications:In S-COMT-deficient mice, MB-COMT in the liver and the duodenum is able to O-methylate about one-half of exogenous L-DOPA. Sexual dimorphism and activity of the two COMT isoforms seems to be tissue specific and more prominent in peripheral tissues than in the brain.

A lack of S-COMT has a notable, albeit small, brain-area and sex-dependent effect on the O-methylation of dopamine and 3,4-dihydroxyphenylacetic acid in the mouse brain. It also induces subtle changes in mouse social interaction behaviors and nociception.

Neuropathic pain is caused by damage or malfunctioning of the nervous system. It is fairly common and more resistant to treatment than other types of pain. Since nitecapone, an inhibitor of catechol-O-methyl-transferase (COMT), has decreased neuropathic symptoms in diabetic rats, we studied its effects in another model of neuropathic pain, the spinal nerve ligation (SNL) model. Spinal nerves L5-6 were ligated in male Wistar rats under anaesthesia to produce the SNL model of neuropathic pain. Nitecapone (30 mg/kg, i.p.) or vehicle was administered once daily starting either 1h before or 2 days after surgery and continued for 14-19 days. Threshold for mechanical allodynia was measured with the digital von Frey test and responses to cold stimuli with the acetone test, before surgery and every other day after it 1h before drug administration. Mechanical and cold allodynia developed in all study groups. Both nitecapone treatments significantly reduced mechanical allodynia and withdrawal thresholds were 80-95% higher compared with the control group. In the acetone test, both nitecapone groups also showed less signs of cold allodynia than the control groups. In nitecapone-naïve animals a single dose of nitecapone also reduced mechanical allodynia on the 14th day after the surgery. Nitecapone reduced the symptoms of neuropathic pain after the SNL, which is in line with the earlier study. Our results suggest that nitecapone and other COMT inhibitors should be studied further in the treatment of neuropathic pain.