Erythropoietin receptor-mediated inhibition of exocytotic glutamate release confers neuroprotection during chemical ischemia

Erythropoietin (EPO) reduced Ca2+-induced glutamate (Glu) release from cult ured cerebellar granule neurons. Inhibition was also produced by EPO mimeti c peptide 1 (EMP1), a small synthetic peptide agonist of EPO receptor (EPO- R), but not by iEMP1, an inactive analogue of EMP1. EPO and EMP1 induced au tophosphorylation of Janus kinase 2 (JAK2), a tyrosine kinase that associat es with EPO-R. Furthermore, genistein, but not genistin, antagonized both t he phosphorylation of JAK2 and the suppression of Glu release induced by EP O and EMP1. During chemical ischemia, substantial amounts of Glu were relea sed from cultured cerebellar and hippocampal neurons by at least two distin ct mechanisms. In the early phase, Glu release occurred by exocytosis of sy naptic vesicle contents, because it was abolished by botulinum type B neuro toxin (BoNT/B). In contrast, the later phase of Glu release mainly involved a BoNT/B-insensitive non-exocytotic pathway. EMP1 inhibited Glu release on ly during the early exocytotic phase. A 20-min exposure of hippocampal slic es to chemical ischemia induced neuronal cell death, especially in the CA1 region and the dentate gyrus, which was suppressed by EMP1 but not iEMP1. H owever, EMP1 did not attenuate neuronal cell death induced by exogenously a pplied Glu. These results suggest that activation of EPO-R suppresses ische mic cell death by inhibiting the exocytosis of Glu.