ABSTRACT
Parkinson's disease (PD) involves an initial loss of striatal dopaminergic terminals evolving into a degeneration of dopaminergic neurons in the substantia nigra (SN), which can be modeled by 6-hydroxydopamine (6-OHDA) administration. Since ATP is a danger signal acting through its P2X7 receptors (P2X7R), we now tested if a blood-brain barrier-permeable P2X7R antagonist, Brilliant Blue G (BBG), controlled the 6-OHDA-induced PD-like features in rats. BBG (45 mg/kg) attenuated the 6-OHDA-induced: 1) increase of contralateral rotations in the apomorphine test, an effect mimicked by another P2X7R antagonist A438079 applied intra-cerebroventricularly; 2) short-term memory impairment in the passive avoidance and cued version of the Morris Water maze; 3) reduction of dopamine content in the striatum and SN; 4) microgliosis and astrogliosis in the striatum. To grasp the mechanism of action of BBG, we used in vitro models exploring synaptotoxicity (striatal synaptosomes) and neurotoxicity (dopamine-differentiated neuroblastoma SH-SY5Y cells). P2X7R were present in striatal dopaminergic terminals, and BBG (100 nM) prevented the 6-OHDA-induced synaptosomal dysfunction. P2X7R were also co-localized with tyrosine hydroxylase in SH-SY5Y cells, where BBG (100 nM) attenuated the 6-OHDA-induced neurotoxicity. This suggests that P2X7R contribute to PD pathogenesis through a triple impact on synaptotoxicity, gliosis and neurotoxicity, highlighting the therapeutic potential of P2X7R antagonists in PD.
Subject(s)
Functional Laterality/drug effects , Motor Activity/drug effects , Neurotoxicity Syndromes/drug therapy , Parkinsonian Disorders/drug therapy , Purinergic P2X Receptor Antagonists/therapeutic use , Rosaniline Dyes/therapeutic use , Animals , Avoidance Learning/drug effects , Brain/drug effects , Brain/metabolism , Cell Line, Tumor , Disease Models, Animal , Exploratory Behavior/drug effects , Gliosis/drug therapy , Male , Maze Learning/drug effects , Memory Disorders/drug therapy , Memory Disorders/etiology , Neurotoxicity Syndromes/etiology , Oxidopamine/toxicity , Parkinsonian Disorders/chemically induced , Parkinsonian Disorders/physiopathology , Purinergic P2X Receptor Antagonists/pharmacology , Rats , Rats, Wistar , Reaction Time/drug effects , Rosaniline Dyes/pharmacologyABSTRACT
ATP is a pleiotropic cell-to-cell signaling molecule in the brain that functions through activation of the P2 receptors (P2R), encompassing ionotropic P2XR or metabotropic P2YR. Noxious brain insults increase the extracellular levels of ATP and previous studies have implicated different P2R, namely P2Y1R, in the control of ischemic brain damage, but it remains to be defined if P2Y1R antagonists also alleviate the behavioral impairments associated with brain ischemia. Furthermore, as P2Y1R can control neuronal and glial functions, we explored if P2Y1R antagonist-mediated protection would mainly involve neuronal and/or glial processes. Adult male mice subject to permanent middle cerebral artery occlusion (pMCAO) displayed an infarcted cortical area (2,3,5-triphenyltetrazolium chloride staining), decreased neurological score with decreased working and reference memory performance (Y-maze, object recognition and aversive memory), accompanied by neuronal damage (FluoroJade C), astrogliosis (glial fibrillary acidic protein) and microgliosis (CD11b). All of these changes were attenuated by intracerebroventricular pre-treatment (10 min before pMCAO) with the generic P2R antagonist 4-[(E)-{4-formyl-5-hydroxy-6-methyl-3-[(phosphono-oxy)methyl]pyridin-2-yl}diazenyl]benzene-1,3-disulfonic acid (PPADS, 0.5-1.0 nmol/µL). In contrast, the selective P2Y1R antagonist (1R*,2S*)-4-[2-Iodo-6-(methylamino)-9H-purin-9-yl]-2-(phosphono-oxy)bicycle[3.1.0] hexane-1-methanol dihydrogen phosphate ester (MRS2500, 1.0-2.0 nmol/µL) afforded equivalent behavioral benefits but only prevented neuronal damage but not astrogliosis or microgliosis upon pMCAO. These results indicated that P2Y1R-associated neuroprotection mainly occurred through neuronal mechanisms, whereas other P2R were also involved in the control of astrocytic reactivity upon brain injury.
