Genetic deletion of NP1 prevents hypoxic-ischemic neuronal death via reducing AMPA receptor synaptic localization in hippocampal neurons.

Trafficking of α-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid receptors (AMPARs) to excitatory synapses is critical to their synaptic functions. Previously, we have shown induction of neuronal pentraxin 1 (NP1) and its colocalization with AMPAR subunit GluR1 in hypoxic-ischemic (HI) brain injury. However, the role of NP1 in mediating GluR1 surface expression, trafficking, and clustering at synapses in HI neuronal death is unclear. Primary hippocampal neurons, isolated from wild-type (WT) and NP1-knockout (C57BL/6 background) mice at DIV 12 to 14 were exposed to 2 to 8 hours of oxygen glucose deprivation (OGD)-in vitro conditions that mimic human stroke. OGD exposure resulted in time-dependent induction of NP1 (∼4-fold), enhanced redistribution of AMAP GluR1 receptors at excitatory synapses, and increased neuronal death. We observed a significant increase in surface and synaptic GluR1 clusters that colocalized with PSD-95 on dendrites with a simultaneous decrease in internalized GluR1. Surface cross-linking with BS(3) showed enhanced membrane insertions of GluR1, and increased phosphorylation at Ser-845 further supported enhanced surface availability of GluR1 after OGD. NP1 protein colocalized with GluR1 and PSD-95, and OGD significantly increased their synaptic coclustering. Most strikingly, the genetic deletion of NP1 resulted in decreases in surface GluR1 cluster density, synaptic localization, phospho-GluR1 (Ser-845) levels, and neuronal death after OGD compared with WT neurons. AMPA (50 μmol/L) induced NP1 and significant cell death in WT but not in NP1-/- neurons. Our results indicate that NP1 plays a key role in synaptic clustering of GluR1, suggesting that targeting NP1 might be a practical approach to preventing ischemic brain damage.