Signaling Mechanisms in the Nitric Oxide Donor- and Amphetamine-Induced Dopamine Release in Mesencephalic Primary Cultured Neurons.

Previous research has shown that nitric oxide (NO) synthase inhibitors prevent rodents' sensorimotor gating impairments induced by dopamine releasing drugs, such as amphetamine (Amph) and methylphenidate. The mechanisms of this effect have not been entirely understood. In the present work, we investigated some possible mechanisms by which the NO donor, NOC-12 (3-ethyl-3-(ethylaminoethyl)-1-hydroxy-2-oxo-1-triazene), influence spontaneous and Amph-induced dopamine release, using rat mesencephalic primary cultured neurons preparations. Our results showed that NOC-12 increased dopamine release in a concentration-dependent manner and potentiated the Amph-induced one. Dopamine release induced by NOC-12 was disrupted by N-acetyl-L-cystein (NAC-a free radical scavenger) and MK-801, a NMDA (N-methyl-D-aspartate) non-competitive antagonist, and was concentration dependently affected by oxadiazolo[4,3]quinoxalin-1-one, an inhibitor of the soluble guanylate cyclase (sGC). In contrast, dopamine released by Amph was facilitated by NAC and by MK-801 and not affected by nifedipine (a L-type-Ca(+2) channel blocker), which enhanced NOC-12-induced dopamine release. The present work demonstrates that DA release induced by NOC-12 is partially dependent on sGC and on NMDA activation, and is modulated by L-type Ca(+2) channel and the antioxidant NAC. This mechanism differs from the Amph-induced one, which appears not to depend on L-type Ca(+2) channel and seems to be facilitated by NMDA channel blocking and by NAC. These results suggest that Amph and NOC-12 induce dopamine release through complementary pathways, which may explain the potentiation of Amph-induced dopamine release by NOC-12. These findings contribute to understand the involvement of NO in dopamine-related neuropsychiatric and neurodegenerative diseases.

Neuroprotective effects of a brain permeant 6-aminoquinoxaline derivative in cell culture conditions that model the loss of dopaminergic neurons in Parkinson disease.

Parkinson disease is a neurodegenerative disorder of aging, characterized by disabling motor symptoms resulting from the loss of midbrain dopaminergic neurons and the decrease of dopamine in the striatum. Current therapies are directed at treating the symptoms but there is presently no cure for the disease. In order to discover neuroprotective compounds with a therapeutical potential, our research team has established original and highly regioselective methods for the synthesis of 2,3-disubstituted 6-aminoquinoxalines. To evaluate the neuroprotective activity of these molecules, we used midbrain cultures and various experimental conditions that promote dopaminergic cell loss. Among a series of 11 molecules, only compound MPAQ (2-methyl-3-phenyl-6-aminoquinoxaline) afforded substantial protection in a paradigm where dopaminergic neurons die spontaneously and progressively as they mature. Prediction of blood-brain barrier permeation by Quantitative Structure-Activity Relationship studies (QSARs) suggested that MPAQ was able to reach the brain parenchyma with sufficient efficacy. HPLC-MS/MS quantification in brain homogenates and MALDI-TOF mass spectrometry imaging on brain tissue sections performed in MPAQ-treated mice allowed us to confirm this prediction and to demonstrate, by MALDI-TOF mass spectrometry imaging, that MPAQ was localized in areas containing vulnerable neurons and/or their terminals. Of interest, MPAQ also rescued dopaminergic neurons, which (i) acquired dependency on the trophic peptide GDNF for their survival or (ii) underwent oxidative stress-mediated insults mediated by catalytically active iron. In summary, MPAQ possesses an interesting pharmacological profile as it penetrates the brain parenchyma and counteracts mechanisms possibly contributive to dopaminergic cell death in Parkinson disease.

Developmental co-expression and functional redundancy of tyrosine phosphatases with neurotrophin receptors in developing sensory neurons.

Receptor-type protein tyrosine phosphatases (RPTPs) have been implicated as direct or indirect regulators of neurotrophin receptors (TRKs). It remains less clear if and how such RPTPs might regulate TRK proteins in vivo during development. Here we present a comparative expression profile of RPTP genes and Trk genes during early stages of murine, dorsal root ganglion maturation. We find little if any specific, temporal mRNA co-regulation between individual RPTP and Ntrk genes between E12.5 and E14.5. Moreover, a double fluorescent in-situ hybridization and immunofluorescence study of seven Rptp genes with Ntrks revealed widespread co-expression of RPTPs in individual neurons, but no tight correlation with Trk expression profiles. No Rptp is expressed in 100% of Ntrk1-expressing neurons, whereas at least 6 RPTPs are expressed in 100% of Ntrk2- and Ntrk3-expressing neurons. An exception is Ptpro, which showed very selective expression. Short hairpin RNA suppression of Ptprf, Ptprs or Ptpro in primary, E13.5 DRG neurons did not alter TRK signalling. We therefore propose that TRK signalling may not be simply dependent on rate-limiting regulation by individual RPTP subtypes during sensory neuron development. Instead, TRK signalling has the potential to be buffered by concurrent inputs from several RPTPs in individual neurons.

Identification of VHY/Dusp15 as a regulator of oligodendrocyte differentiation through a systematic genomics approach.

Multiple sclerosis (MS) is a neuroinflammatory disease characterized by a progressive loss of myelin and a failure of oligodendrocyte (OL)-mediated remyelination, particularly in the progressive phases of the disease. An improved understanding of the signaling mechanisms that control differentiation of OL precursors may lead to the identification of new therapeutic targets for remyelination in MS. About 100 mammalian Protein Tyrosine Phosphatases (PTPs) are known, many of which are involved in signaling both in health and disease. We have undertaken a systematic genomic approach to evaluate PTP gene activity in multiple sclerosis autopsies and in related in vivo and in vitro models of the disease. This effort led to the identification of Dusp15/VHY, a PTP previously believed to be expressed only in testis, as being transcriptionally regulated during OL differentiation and in MS lesions. Subsequent RNA interference studies revealed that Dusp15/VHY is a key regulator of OL differentiation. Finally, we identified PDGFR-beta and SNX6 as novel and specific Dusp15 substrates, providing an indication as to how this PTP might exert control over OL differentiation.

Tryptamine-derived alkaloids from Annonaceae exerting neurotrophin-like properties on primary dopaminergic neurons.

N-fatty acyl tryptamines constitute a scarce group of natural compounds mainly encountered in Annonaceous plants. No biological activity was reported so far for these rare molecules. This study investigated the neurotrophic properties of these natural tryptaminic derivatives on dopaminergic (DA) neurons in primary mesencephalic cultures. A structure-activity relationships study led us to precise the role of a nitrogen atom into the aliphatic chain conferring to the compounds a combined neuroprotective and neuritogenic activity in the nanomolar range. The potent antioxidant activity of these natural products seems to be involved in part of their mechanism of action. This study provides the first description of natural neurotrophin mimetics present in Annonaceae extracts, and led to the biological characterization of compounds, which present a potential interest in neurodegenerative diseases such as Parkinson's disease.

Chemicals possessing a neurotrophin-like activity on dopaminergic neurons in primary culture.

BACKGROUND: Neurotrophic factors have been shown to possess strong neuroprotective and neurorestaurative properties in Parkinson's disease patients. However the issues to control their delivery into the interest areas of the brain and their surgical administration linked to their unability to cross the blood brain barrier are many drawbacks responsible of undesirable side effects limiting their clinical use. A strategy implying the use of neurotrophic small molecules could provide an interesting alternative avoiding neurotrophin administration and side effects. In an attempt to develop drugs mimicking neurotrophic factors, we have designed and synthesized low molecular weight molecules that exhibit neuroprotective and neuritogenic potential for dopaminergic neurons. PRINCIPAL FINDINGS: A cell-based screening of an in-house quinoline-derived compound collection led to the characterization of compounds exhibiting both activities in the nanomolar range on mesencephalic dopaminergic neurons in spontaneous or 1-methyl-4-phenylpyridinium (MPP(+))-induced neurodegeneration. This study provides evidence that rescued neurons possess a functional dopamine transporter and underlines the involvement of the extracellular signal-regulated kinase 1/2 signaling pathway in these processes. CONCLUSION: Cell-based screening led to the discovery of a potent neurotrophic compound possessing expected physico-chemical properties for blood brain barrier penetration as a serious candidate for therapeutic use in Parkinson disease.

Quinoline compounds decrease in vitro spontaneous proliferation of peripheral blood mononuclear cells (PBMC) from human T-cell lymphotropic virus (HTLV) type-1-infected patients.

In vitro spontaneous proliferation is the immunological hallmark of peripheral blood mononuclear cells (PBMC) from HTLV-1-infected individuals. Quinoline compounds down regulate in vitro cell proliferation of HTLV-1 transformed cell lines. In the present study we assessed the capacity of quinolines to inhibit spontaneous cell proliferation of PBMC from HTLV-1-infected individuals. Twenty-two quinolines were evaluated. Toxicity was first assessed on PBMC from healthy donors by using both the Trypan blue technique and Tetrazolium Salt (XTT) method and then the antiproliferative effect was measured by a classic lymphoproliferative assay on PBMC from three HTLV-1-infected individuals, in the presence of decreasing concentrations of quinolines (from 100microM to 0.8microM), after 5 days of culture. We found that 14 out of 22 compounds were non-toxic to PBMC from uninfected individuals at 100, 50 and 10microM. Four compounds presented a capacity to inhibit more than 80% of the spontaneous proliferation: 7 at 25microM and 10, 20 and 23 at 100microM. Our results indicate that some quinolines block spontaneous proliferation of PBMC from HTLV-1-infected individuals.