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 was investigated by employing a variety of agonists active at GluR5-7. Their excitotoxic profiles were determined in primary cultures of cerebellar granule cells, which abundantly expressed low-affinity KA receptors, and in the absence of any AMPA receptor-mediated neurotoxicity. Neurotoxicity induced by these compounds was analysed by phase contrast microscopy, a cell viability assay, the TUNEL technique (apoptosis), and by employing propidium iodide (PI; necrosis). All agonists induced concentration-dependent neurotoxicity, with rank order (EC(50) values; microM): (S)-iodowillardiine (IW) 0.2>(2S,4R)-4-methylglutamate (4-MG) 36>(2S,4R,6E)-2-amino-4-carboxy-7-(2-naphthyl)hept-6-enoic acid (LY339434) 46>KA 74>(RS)-2-amino-3-(hydroxy-5-tert-butylisoxazol-4yl)propanoic acid (ATPA) 88. IW exposure resulted in apoptosis at lower concentrations (<30 microM) and necrosis at higher concentrations, both of which were attenuated by CNQX (50 microM), but not MK-801 (10 microM). ATPA-mediated neurotoxicity was purely apoptotic and was attenuated by the non-NMDA receptor antagonists. Both IW and ATPA induced injury with the morphological characteristics of apoptosis shown by the presence of TUNEL-positive neurones. LY339434-mediated neuronal injury was only attenuated by MK-801 and was necrotic in nature. Similarly, 4-MG (>30 microM) exposure caused necrosis that was partially attenuated by MK-801 (10 microM) and CNQX (50 microM). The patterns of neurotoxicity possessed a complex pharmacological profile, demonstrated an apoptotic-necrotic continuum and were inconsistent with past findings, further outlining the importance of characterizing novel compounds at native receptors. ATPA and to a lesser extent IW appear to be suitable drugs for low-affinity KA receptors. Since toxicity-mediated by low-affinity KA receptors seem likely to contribute to neurodegenerative conditions, our study importantly examines the excitotoxic profile of these novel agonists.
Giardina
S F
SF
Department of Pharmacology, Monash University, PO Box 13E, Clayton, Victoria, 3800, Australia.
Beart
P M
PM
D000818:Animals; D002467:Cell Nucleus / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D002531:Cerebellum / Q000166:cytology / Q000187:drug effects*; D004396:Coloring Agents; D053938:DNA Fragmentation / Q000187:drug effects; D018690:Excitatory Amino Acid Agonists / Q000494:pharmacology*; D007150:Immunohistochemistry; D007202:Indicators and Reagents; D051379:Mice; D011419:Propidium; D018091:Receptors, AMPA / Q000502:physiology; D018092:Kainic Acid Receptors / Q000819:agonists*
D004396:Coloring Agents; D018690:Excitatory Amino Acid Agonists; D007202:Indicators and Reagents; D011419:Propidium; D018091:Receptors, AMPA; D018092:Kainic Acid Receptors