New role of P2X7 receptor in an Alzheimer's disease mouse model.

Extracellular aggregates of amyloid ß (Aß) peptides, which are characteristic of Alzheimer's disease (AD), act as an essential trigger for glial cell activation and the release of ATP, leading to the stimulation of purinergic receptors, especially the P2X7 receptor (P2X7R). However, the involvement of P2X7R in the development of AD is still ill-defined regarding the dual properties of this receptor. Particularly, P2X7R activates the NLRP3 inflammasome leading to the release of the pro-inflammatory cytokine, IL-1ß; however, P2X7R also induces cleavage of the amyloid precursor protein generating Aß peptides or the neuroprotective fragment sAPPα. We thus explored in detail the functions of P2X7R in AD transgenic mice. Here, we show that P2X7R deficiency reduced Aß lesions, rescued cognitive deficits and improved synaptic plasticity in AD mice. However, the lack of P2X7R did not significantly affect the release of IL-1ß or the levels of non-amyloidogenic fragment, sAPPα, in AD mice. Instead, our results show that P2X7R plays a critical role in Aß peptide-mediated release of chemokines, particularly CCL3, which is associated with pathogenic CD8+ T cell recruitment. In conclusion, our study highlights a novel detrimental function of P2X7R in chemokine release and supports the notion that P2X7R may be a promising therapeutic target for AD.

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.

New 6-Aminoquinoxaline Derivatives with Neuroprotective Effect on Dopaminergic Neurons in Cellular and Animal Parkinson Disease Models.

Parkinson's disease (PD) is a neurodegenerative disorder of aging characterized by motor symptoms that result from the loss of midbrain dopamine neurons and the disruption of dopamine-mediated neurotransmission. There is currently no curative treatment for this disorder. To discover druggable neuroprotective compounds for dopamine neurons, we have designed and synthesized a second-generation of quinoxaline-derived molecules based on structure-activity relationship studies, which led previously to the discovery of our first neuroprotective brain penetrant hit compound MPAQ (5c). Neuroprotection assessment in PD cellular models of our newly synthesized quinoxaline-derived compounds has led to the selection of a better hit compound, PAQ (4c). Extensive in vitro characterization of 4c showed that its neuroprotective action is partially attributable to the activation of reticulum endoplasmic ryanodine receptor channels. Most interestingly, 4c was able to attenuate neurodegeneration in a mouse model of PD, making this compound an interesting drug candidate for the treatment of this disorder.

CX3CR1 Disruption Differentially Influences Dopaminergic Neuron Degeneration in Parkinsonian Mice Depending on the Neurotoxin and Route of Administration.

Parkinson's disease (PD) is characterized by progressive degeneration of dopaminergic neurons accompanied by an inflammatory reaction. The neuron-derived chemokine fractalkine (CX3CL1) is an exclusive ligand for the receptor CX3CR1 expressed on microglia. The CX3CL1/CX3CR1 signaling is important for sustaining microglial activity. Using a recently developed PD model, in which the 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) toxin is delivered intranasally, we hypothesized that CX3CR1 could play a role in neurotoxicity and glial activation. For this, we used CX3CR1 knock-in mice and compared results with those obtained using the classical PD models through intraperitonal MPTP or intrastriatal 6-hydroxydopamine (6-OHDA). The striatum from all genotypes (CX3CR1(+/+), CX3CR1(+/GFP) and CX3CR1-deficient mice) showed a significant dopaminergic depletion after intranasal MPTP inoculation. In contrast to that, we could not see differences in the number of dopaminergic neurons in the substantia nigra of CX3CR1-deficient animals. Similarly, after 6-OHDA infusion, the CX3CR1 deletion decreased the amphetamine-induced turning behavior observed in CX3CR1(+/GFP) mice. After the 6-OHDA inoculation, a minor dopaminergic neuronal loss was observed in the substantia nigra from CX3CR1-deficient mice. Distinctly, a more extensive neuronal cell loss was observed in the substantia nigra after the intraperitoneal MPTP injection in CX3CR1 disrupted animals, corroborating previous results. Intranasal and intraperitoneal MPTP inoculation induced a similar microgliosis in CX3CR1-deficient mice but a dissimilar change in the astrocyte proliferation in the substantia nigra. Nigral astrocyte proliferation was observed only after intraperitoneal MPTP inoculation. In conclusion, intranasal MPTP and 6-OHDA lesion in CX3CR1-deficient mice yield no nigral dopaminergic neuron loss, linked to the absence of astroglial proliferation.

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.

Evaluation of nigrostriatal neurodegeneration and neuroinflammation following repeated intranasal 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) administration in mice, an experimental model of Parkinson's disease.

Parkinson's disease (PD) is the second most common neurodegenerative disorder affecting approximately 1% of the population older than 60 years. The administration of the proneurotoxin 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) in mice is the most widely used approach to elucidate the mechanisms of cell death involved in PD. However, the magnitude of the PD-like neurodegeneration induced by MPTP depends on many variables, including the regimen of its administration. It has been demonstrated that intranasal (i.n.) administration of MPTP constitutes a new route of toxin delivery to the brain that mimics environmental exposure to neurotoxins. Previous data showed that mice submitted to chronic and acute i.n. MPTP treatment displayed a robust (~80%) and moderate (~55%) loss of striatal dopamine, respectively. However, little is known about the neurodegenerative and neuroinflammatory processes following a subacute i.n. MPTP administration in mice. Here, the C57BL/6 mice were infused intranasally with MPTP (1 mg/nostril/day) during 4 consecutive days. At 7 and 28 days after the last administration, the subacute i.n. MPTP regime decreased the tyrosine hydroxylase (TH)-labeling in the striatum (40-50%) and substantia nigra (25-30%) and increased the astrogliosis in such brain areas at both time points. Taken together, our data showed that the subacute administration of MPTP into the nasal cavity of C57BL/6 mice induces long-lasting neurodegeneration and neuroinflammation in the nigrostriatal pathway, thus representing a valuable animal model for the investigation of neuroprotective strategies in PD.

MALDI mass spectrometry imaging of 1-methyl-4-phenylpyridinium (MPP+) in mouse brain.

Parkinson's disease (PD) is the second most common neurodegenerative disorder affecting ~1% of the population older than 60 years. The administration of the proneurotoxin 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) in mice is one of the most widely used approach to elucidate the mechanisms of cell death involved in PD. Its toxicity is attributed to its active metabolite 1-methyl-4-phenylpyridinium (MPP(+)). However, the magnitude of the PD-like neurodegeneration induced by MPTP depends on many variables, including the route of administration. Different groups, including us, demonstrated that intranasal (i.n.) administration of MPTP constitutes a new route of toxin delivery to the brain that mimics environmental exposure to neurotoxins. In particular, our previous data showed that mice submitted to acute i.n. MPTP administration displayed a significant decrease of striatal dopamine (DA) and a loss of dopaminergic (DA) neurons in the substantia nigra pars compacta. However, little is known about the timing and the anatomical distribution of MPP(+) after i.n. MPTP administration in mice. In the present study, C57BL/6J mice received one dose of i.n. MPTP (1 mg/nostril) and were sacrificed at two different times after the administration. Using matrix-assisted laser desorption-ionization mass spectrometry imaging, a new technique for the detection of endogenous unlabeled molecules in tissue sections, we showed for the first time the MPP(+) anatomical distribution in different brain regions. We demonstrated that the toxin first reached almost all the brain areas; however, in a second time MPP(+) remained highly concentrated in the olfactory bulb, the basal ganglia, the ventral mesencephalon, and the locus coeruleus, regions differently affected in PD.

Six weeks of voluntary exercise don't protect C57BL/6 mice against neurotoxicity of MPTP and MPP(+).

Exercise improves the central nervous system (CNS) functions and is widely recommended for neurological patients with, e.g., Alzheimer's and Parkinson's disease (PD). However, exercise-induced neuroprotection is an open discussion. Here, the intranasal administration of the neurotoxin 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP, 65 mg/kg) caused death of dopaminergic neurons in the substantia nigra pars compacta and depletion of dopamine in the striatum of C57BL/6 mice. 1-Methyl-4-phenylpyridinium, the active metabolite of MPTP, also inhibited complex-I activity of mitochondria isolated from the CNS of mice. However, 6 weeks of exercise on voluntary running wheels did not protect against nigrostriatal neurodegeneration or mitochondrial inhibition, suggesting that benefits of exercise for PD may not be associated with neuroprotection. The literature presents other candidates, such as neurotrophins or increased antioxidant defenses.

Parkin-knockout mice did not display increased vulnerability to intranasal administration of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP).

The loss of nigral dopaminergic neurons in Parkinson's disease (PD) is believed to result from interactions between genetic susceptibility and environmental factors. Although loss-of-function mutations in the parkin gene cause early-onset familial PD, the hybrid 129Sv-C57BL/6 parkin-deficient mice did not display spontaneous degeneration of the nigrostriatal pathway or enhanced vulnerability to neurotoxicity induced by 6-hydroxydopamine (6-OHDA) or intraperitoneal 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) intoxication. We aimed to re-evaluate the role of parkin in a pure C57BL/6 background after an acute intranasal (i.n.) MPTP administration, a new route of toxin delivery to the brain that mimics environmental exposure to neurotoxins. We found that the deficiency of parkin gene modifies the D-amphetamine-induced locomotion in saline-treated animals. Intranasal MPTP induced Parkinsonism in parkin⁺/⁺ mice, through depletion of striatal dopamine, decreased number of dopaminergic neurons in the substantia nigra, and decreased D-amphetamine-induced hyperlocomotion. Additionally, the deletion of the parkin gene in a pure C57BL/6 background did not lead to increased vulnerability to i.n. MPTP-induced neurotoxicity. Moreover, the i.n. MPTP induced nigral astrogliosis predominantly in the pars reticulata in wild type and parkin⁻/⁻ mice. Taken together, these results showed that the absence of parkin did not modify the vulnerability of nigrostriatal dopaminergic pathway after i.n. MPTP intoxication, suggesting that independently of mouse strain, the endogenous parkin is not required for protection of this system. These findings also suggest that the development of familial parkin-linked PD is not associated with exposure to environmental factors that specifically affects the dopaminergic system.