Tryptophan hydroxylase 2 aggregates through disulfide cross-linking upon oxidation: possible link to serotonin deficits and non-motor symptoms in Parkinson's disease.

Parkinson's disease (PD) is a progressive neurodegenerative disorder characterized by the loss of dopamine neurons of the nigrostriatal system, resulting in severe motor disturbances. Although much less appreciated, non-motor symptoms are also very common in PD and many can be traced to serotonin neuronal deficits. Tryptophan hydroxylase (TPH) 2, the rate-limiting enzyme in the serotonin biosynthesis, is a phenotypic marker for serotonin neurons and is known to be extremely labile to oxidation. Therefore, the oxidative processes that prevail in PD could cause TPH2 misfolding and modify serotonin neuronal function much as is seen in dopamine neurons. Oxidation of TPH2 inhibits enzyme activity and leads to the formation of high molecular weight aggregates in a dithiothreitol-reversible manner. Cysteine-scanning mutagenesis shows that as long as a single cysteine residue (out of a total of 13 per monomer) remains in TPH2, it cross-links upon oxidation and only cysteine-less mutants are resistant to this effect. The effects of oxidants on TPH2 catalytic function and cross-linking are also observed in intact TPH2-expressing HEK293 cells. Oxidation shifts TPH2 from the soluble compartment into membrane fractions and large inclusion bodies. Sequential non-reducing/reducing 2-dimensional sodium dodecyl sulfate-polyacrylamide gel electrophoresis and immunoblotting confirmed that TPH2 was one of a small number of cytosolic proteins that form disulfide-bonded aggregates. The propensity of TPH2 to misfold upon oxidation of its cysteine residues is responsible for its catalytic lability and may be related to loss of serotonin neuronal function in PD and the emergence of non-motor (psychiatric) symptoms.

Role of the primary motor cortex in L-Dopa-induced dyskinesia and its modulation by 5-HT1A receptor stimulation.

While serotonin 5-HT1A receptor (5-HT1AR) agonists reduce L-DOPA-induced dyskinesias (LID) by normalizing activity in the basal ganglia neurocircuitry, recent evidence suggests putative 5-HT1AR within the primary motor cortex (M1) may also contribute. To better characterize this possible mechanism, c-fos immunohistochemistry was first used to determine the effects of systemic administration of the full 5-HT1AR agonist ±8-OH-DPAT on L-Dopa-induced immediate early gene expression within M1 and the prefrontal cortex (PFC) of rats with unilateral medial forebrain bundle (MFB) dopamine (DA) lesions. Next, in order to determine if direct stimulation of 5-HT1AR within M1 attenuates the onset of LID, rats with MFB lesions were tested for L-Dopa-induced abnormal involuntary movements (AIMs) and rotations following M1 microinfusions of ±8-OH-DPAT with or without coadministration of the 5-HT1AR antagonist WAY100635. Finally, ±8-OH-DPAT was infused into M1 at peak dyskinesia to determine if 5-HT1AR stimulation attenuates established L-Dopa-induced AIMs and rotations. While no treatment effects were seen within the PFC, systemic ±8-OH-DPAT suppressed L-Dopa-induced c-fos within M1. Intra-M1 5-HT1AR stimulation diminished the onset of AIMs and this effect was reversed by WAY100635 indicating receptor specific effects. Finally, continuous infusion of ±8-OH-DPAT into M1 at peak dyskinesia alleviated L-Dopa-induced AIMs. Collectively, these findings support an integral role for M1 in LID and its modulation by local 5-HT1AR.

Serotonin 1B receptor stimulation reduces D1 receptor agonist-induced dyskinesia.

Dopamine replacement therapy for the treatment of Parkinson's disease leads to deleterious abnormal involuntary movements (AIMs), known as L-3,4-dihydroxyphenylalanine (L-DOPA)-induced dyskinesia, which parallels enhanced striatal dopamine D1 receptor-mediated signaling. Recent evidence suggests stimulation of striatal serotonin (5-HT) 1B receptors may reduce D1-mediated signaling. Given this potential antidyskinetic mechanism, male hemiparkinsonian Sprague-Dawley rats received treatments of D1 receptor agonist, SKF81297, (0.8 mg/kg) or L-DOPA (12 mg/kg, subcutaneous injection). Dyskinetic movements were rated using the AIMs scale. Rats were then administered vehicle (100% dimethyl sulfoxide) or the 5-HT1B receptor agonist, CP94253, (1.5 or 3.0 mg/kg, subcutaneous injection), followed by SKF81297 or L-DOPA and rated for AIMs. Results indicate that CP94253 mitigates both L-DOPA and D1 receptor agonist-induced dyskinesia. These findings suggest that 5-HT1B receptor stimulation directly diminishes D1 receptor-mediated dyskinesia, implicating an important target for the treatment of L-DOPA-induced dyskinesia.