Involvement of the secretory pathway for AMPA receptors in NMDA-induced potentiation in hippocampus.
A chemical form of synaptic potentiation was produced with a brief bath application of NMDA to rat hippocampal slices. Two methods were used to assess changes in membrane-bound AMPA receptors. Traditional subcellular fractionation was used to isolate synaptic membranes; alternatively, membrane receptors were cross-linked with the membrane-impermeable reagent bis(sulfosuccinimidyl) suberate, and levels of nonmembrane receptors were determined. In both cases, Western blots were used to determine the content of receptor subunits in various subcellular fractions. NMDA-induced potentiation was associated with increased levels of glutamate receptor 1 (GluR1) and GluR2/3 subunits of AMPA receptors in synaptic membrane preparations, whereas no change was observed in whole homogenates. Both KN-62, an inhibitor of calcium/calmodulin kinase, and calpain inhibitor III, a calpain inhibitor, inhibited NMDA-induced potentiation and changes in GluR1 and GluR2/3 subunits of AMPA receptors. Brefeldin A (BFA) inhibits protein trafficking between the Golgi apparatus and cell membranes. Pretreatment of hippocampal slices with BFA significantly decreased NMDA-induced potentiation and completely prevented an NMDA-induced increase in GluR1 levels in membrane fractions. Thus, the levels of GluR1 and GluR2/3 subunits of AMPA receptors are rapidly upregulated in synaptic membranes under conditions associated with potentiation of synaptic responses, and this upregulation requires a functional secretory pathway.
Broutman
G
G
Neuroscience Program, University of Southern California, Los Angeles, California 90089-2520, USA.
Baudry
M
M
D000818:Animals; D015153:Blotting, Western; D020126:Brefeldin A / Q000494:pharmacology; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000037:antagonists & inhibitors; D002154:Calpain / Q000037:antagonists & inhibitors; D003432:Cross-Linking Reagents / Q000494:pharmacology; D004558:Electric Stimulation; D006624:Hippocampus / Q000737:chemistry / Q000166:cytology / Q000378:metabolism*; D066298:In Vitro Techniques; D017774:Long-Term Potentiation / Q000187:drug effects / Q000502:physiology*; D016202:N-Methylaspartate / Q000378:metabolism* / Q000494:pharmacology; D021381:Protein Transport / Q000187:drug effects; D051381:Rats; D017207:Rats, Sprague-Dawley; D018091:Receptors, AMPA / Q000187:drug effects / Q000378:metabolism*; D013347:Subcellular Fractions / Q000378:metabolism; D013570:Synaptic Membranes / Q000378:metabolism; D009435:Synaptic Transmission / Q000187:drug effects; D015854:Up-Regulation / Q000187:drug effects
D020126:Brefeldin A; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D002154:Calpain; D003432:Cross-Linking Reagents; D016202:N-Methylaspartate; D018091:Receptors, AMPA; C104723:glutamate receptor ionotropic, AMPA 3; C104722:glutamate receptor ionotropic, AMPA 2