Inhibition of phosphodiesterase - 10A by Papaverine protects human cortical neurons from quinolinic acid induced oxidative stress and synaptic proteins alterations

Phosphodi esterase-10A (PDE10A) hydrolyse the secondary messengers cGMP and cAMP which play critical role in neurodevelopment and brain functions. PDE10A is linked to progression of neurodegenerative diseases like Alzheimer’s, Parkinson’s, Huntington’s diseases etc and a critical role in cognitive functions. The present study was undertaken to determine the possible neuroprotective effects and the associated mechanism of papaverine (PAP) against quinolinic acid (QUIN) induced excitotoxicity using human primary cortical neurons. Cytotoxicity potential of PAP was analysed using MTS assay. Reactive oxygen species (ROS) and mitochondrial membrane potential were measured by DCF-DA and JC10 staining, respectively. Caspase 3/7 and cAMP levels using ELISA kits. Effect of PAP on the CREB, BNDF and synaptic proteins such as SAP-97, synaptophysin, synapsin-I, PSD-95 expression was analysed by Western blotting technique. Pre-treatment with PAP increased intracellular cAMP and nicotinamide adenine dinucleotide (NAD+) levels, restored mitochondrial membrane potential (ΔΨm), and decreased ROS and caspase3/7 content in QUIN exposed neurons. PAP up-regulated CREB and BDNF, and synaptic proteins expression. In summary, these data indicate that PDE10A involves in QUIN mediated neurotoxicity and its inhibition can elicit neuroprotection by reducing the oxidative stress and protecting synaptic proteins via upregulation of cAMP signalling cascade. ### Competing Interest Statement The authors have declared no competing interest. * AD : Alzheimer’s diseases ALS : Amyotrophic lateral sclerosis ANOVA : Analysis of variance Aβ : Amyloid beta ARE : Antioxidant response element ATP : Adenosine triphosphate BDNF : Brain-derived neurotrophic factor cAMP : Cyclic adenosine monophosphate cGMP : Cyclic guanosine monophosphate CREB : cAMP response element-binding protein Cyt c : Cytochrome c DCFDA : 2□, 7□-Dichlorofluorescin Diacetate EDTA : Ethylenediaminetetraacetic acid HEPES : 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid HD : Huntington’s disease JC-10 : 5,5,6,6’-tetrachloro-1,1’,3,3’ tetraethylbenzimidazoylcarbocyanine iodide KP : Kynurenine pathway MS : Multiple sclerosis MTS : 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide NAD : Nicotinamide adenine dinucleotide NMDA : N-methyl-D-aspartate NF-Kb : Nuclear factor kappa-light-chain-enhancer of activated B cells nNOS : Neuronal nitric oxide synthase Nrf2 : Nuclear erythroid 2-related factor 2 PAP : Papaverine PARP : Poly (ADP-ribose) polymerase PBS : Phosphate-buffered saline PD : Parkinson’s disease PDE : Phosphodiesterase PDE10A : Phosphodiesterase-10A PKA : Protein kinase A pNpp : para-Nitrophenyl phosphate PPARy : Peroxisome proliferator-activated receptor gamma PSD-95 : Post synaptic density protein-95 QUIN : Quinolinic acid RIPA : Radioimmunoprecipitation assay ROF : Roflumilast ROS : Reactive oxygen species SAP 97 : Synapse-associated protein 97 SDS : Sodium dodecyl sulphate SYN1 : Synapsin-I TBST : Tris-Buffered Saline and Tween 20 ΔΨm : mitochondrial membrane potential