dizocilpine-maleate and delta-philanthotoxin

Earlier findings had suggested that spontaneous and evoked glutamate release activates non-overlapping populations of NMDA receptors. Here, we evaluated whether AMPA receptor populations activated by spontaneous and evoked release show a similar segregation. To track the receptors involved in spontaneous or evoked neurotransmission, we used a polyamine agent, philanthotoxin, that selectively blocks AMPA receptors lacking GluR2 subunits in a use-dependent manner. In hippocampal neurons obtained from GluR2-deficient mice, philanthotoxin application decreased AMPA-receptor-mediated spontaneous miniature EPSCs (AMPA-mEPSCs) down to 20% of their initial level within 5 min. In contrast, the same philanthotoxin application at rest decreased the subsequent AMPA-receptor-mediated evoked EPSCs (eEPSCs) only down to 80% of their initial value. A 10-min-long perfusion of philanthotoxin further decreased AMPA-eEPSC amplitudes to 60% of their initial magnitude, which remained substantially higher than the level of AMPA-mEPSC block achieved within 5 min. Finally, stimulation after removal of philanthotoxin resulted in reversal of AMPA-eEPSC block, verifying strict use dependence of philanthotoxin. These results support the notion that spontaneous and evoked neurotransmission activate distinct sets of AMPA receptors and bolster the hypothesis that synapses harbor separate microdomains of evoked and spontaneous signaling.

Topics: Anesthetics, Local; Animals; Animals, Newborn; Cells, Cultured; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Excitatory Postsynaptic Potentials; Female; Glutamic Acid; Hippocampus; Lidocaine; Male; Mice; Mice, Knockout; Neurons; Patch-Clamp Techniques; Polyamines; Receptors, AMPA; Tetrodotoxin; Valine

alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor (AMPA-R)-mediated neurotoxicity was studied in relation to subunit expression and the presence of Ca(2+)-permeable receptor channels. AMPA-mediated toxicity had two components: 1) a direct AMPA-R-mediated component, which was not due to Ca(2+) influx through voltage-gated Ca(2+) channels, reversal of the Na(+)/Ca(2+) exchanger or release of calcium from dantrolene-sensitive intracellular Ca(2+) stores, and 2) a minor, indirect component involving activation of NMDA receptor channels, because of glutamate release and removal of the Mg(2+) block of the NMDA receptor on AMPA-R stimulation. The involvement of Ca(2+) influx through AMPA-R was also examined. The number of neurons possessing Ca(2+)-permeable AMPA-R increased during culture development, concurrently with an increasing susceptibility for AMPA-induced toxicity during development. GluR2(R) levels also increased during development, and channel blockers of Ca(2+)-permeable AMPA-R lacking the GluR2(R) subunit (spermine and philanthotoxin) failed to prevent neurotoxicity or increases in [Ca(2+)](i). Thus, the direct AMPA-R-mediated toxicity may be explained by initiation of cell death by Ca(2+) fluxing through AMPA-R containing GluR2(R). The components of direct AMPA-R-mediated toxicity are proposed to be 1) toxicity mediated by GluR2(R)-lacking AMPA-R and 2) toxicity mediated by low-Ca(2+)-permeability AMPA-R containing GluR2(R).

Topics: alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid; Animals; Apoptosis; Benzothiadiazines; Calcium; Calcium Channel Blockers; Calcium Channels; Calcium Signaling; Cells, Cultured; Cerebral Cortex; Dizocilpine Maleate; Excitatory Amino Acid Agonists; Excitatory Amino Acid Antagonists; Flunarizine; Gene Expression Regulation, Developmental; Ion Channel Gating; Lanthanum; Macromolecular Substances; Mice; Nerve Tissue Proteins; Neurons; Neuroprotective Agents; Nifedipine; omega-Conotoxins; Polyamines; Protein Subunits; Receptors, AMPA; Sodium; Sodium Channels; Sodium-Calcium Exchanger; Spermine; Tetrodotoxin