Excitotoxic profiles of novel, low-affinity kainate receptor agonists in primary cultures of murine cerebellar granule cells

The involvement of low-affinity kainate (KA) receptors in neuronal injury w as investigated by employing a variety of agonists active at GluR5-7. Their excitotoxic profiles were determined in primary cultures of cerebellar gra nule cells, which abundantly expressed low-affinity KA receptors, and in th e absence of any AMPA receptor- mediated neurotoxicity. Neurotoxicity induc ed by these compounds was analysed by phase contrast microscopy, a cell via bility assay, the TUNEL technique (apoptosis), and by employing propidium i odide (PI; necrosis). All agonists induced concentration-dependent neurotox icity, with rank order (EC50 values; muM): (S)-iodowillardiine (IW) 0.2 > ( 2S,4R)-4-methylglutamate (4-MG) 36 > (2S,4R,6E)-2-amino-4-carboxy-7-(2-naph thyl)hept-6-enoic acid (LY339434) 46 > KA 74 > (RS)-2-amino-3-(hydroxy-5-te rt-butylisoxazol-4yl)propanoic acid (ATPA) 88. IW exposure resulted in apop tosis at lower concentrations (< 30 muM) and necrosis at higher concentrati ons, both of which were attenuated by CNQX (50 muM), but not MK-801 (10 muM ). ATPA-mediated neurotoxicity was purely apoptotic and was attenuated by t he non-NMDA receptor antagonists. Both IW and ATPA induced injury with the morphological characteristics of apoptosis shown by the presence of TUNEL-p ositive neurones. LY339434-mediated neuronal injury was only attenuated by MK-801 and was necrotic in nature. Similarly, 4-MG (> 30 muM) exposure caus ed necrosis that was partially attenuated by MK-801 (10 PM) and CNQX (50 mu M). The patterns of neurotoxicity possessed a complex pharmacological profi le, demonstrated an apoptotic-necrotic continuum and were inconsistent with past findings, further outlining the importance of characterizing novel co mpounds at native receptors. ATPA and to a lesser extent IW appear to be su itable drugs for low-affinity KA receptors. Since toxicity-mediated by low- affinity KA receptors seem likely to contribute to neurodegenerative condit ions, our study importantly examines the excitotoxic profile of these novel agonists. (C) 2001 Published by Elsevier Science Ltd.