Supplementary Material for: Kainate Receptors Mediate Regulated Exocytosis of Secretory Phospholipase A2 in SH-SY5Y Neuroblastoma Cells

Secretory phospholipase A₂ (sPLA₂) isoforms are widely expressed in the brain and spinal cord. Group IIA sPLA₂ (sPLA₂-IIA) has been shown to stimulate exocytosis and release of neurotransmitters in neuroendocrine PC12 cells and neurons, suggesting a role of the enzyme in neuronal signaling and synaptic transmission. However, the mechanisms by which sPLA₂ is itself released, and a possible relation between glutamate receptors and sPLA₂ exocytosis, are unknown. This study was carried out to elucidate the effects of glutamate receptor agonists on exocytosis of sPLA₂-IIA in transfected SH-SY5Y neuroblastoma cells. sPLA₂-IIA enzyme was packaged in fusion-competent vesicles and released constitutively or upon stimulation, suggesting regulated secretion. The signal peptide of sPLA₂-IIA is required for its vesicular localization and exocytosis. External application of α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) and kainate (KA) induced vesicular exocytosis and release of sPLA₂-IIA. UBP 302, a GluR5-specific KA receptor antagonist, abolished the effect of KA, confirming the role of KA receptors in mediating sPLA₂-IIA secretion. Moreover, KA-induced sPLA₂-IIA secretion is dependent on Ca²⁺ and protein kinase C. Together, these findings provide evidence of a link between glutamate receptors and regulated sPLA₂ secretion in neurons that may play an important role in synaptic plasticity, pain transmission and neurodegenerative diseases.