GROWTH-CONDITIONS DIFFERENTIALLY REGULATE THE EXPRESSION OF PHA-AMINO-3-HYDROXY-5-METHYLISOXAZOLE-4-PROPIONATE (AMPA) RECEPTOR SUBUNITS IN CULTURED NEURONS

We have studied the expression of pha-amino-3-hydroxy-5-methylisoxazol e-4-propionate (AMPA) receptor subunits in cultured cerebellar granule cells [7 days in vitro (DIV)] grown in medium containing different co ncentrations of K+ (10, 25, or 40 mM) with or without 100 muM N-methyl -D-aspartate (NMDA; added once after 2 DIV). All these conditions are known to influence maturation and survival of granule cells, as well a s the functional expression of NMDA receptors during development in cu lture. The expression of both glutamate receptor (GluR) subunit 1 mRNA and receptor protein was low in cultures grown in 10 mM K+ (K10) and increased dramatically in cultures grown in 25 mM K+ (K25), with inter mediate levels found in cultures grown in K10 and chronically exposed to NMDA (K10 + NMDA). In cultures grown in 40 mM K+ (K40), the express ion of GluR1 mRNA and receptor protein was lower than in K25 but still higher than in K10. GluR2 and -3 subunits were differently regulated by growth conditions, with their expression being higher in Kl 0 and p rogressively reduced to the lowest levels in K40 (both mRNA and recept or proteins). GluR4 mRNA levels did not differ between K10 and K25, al though they were reduced by chronic exposure to NMDA. To test how the differential expression of the various subunits affects the functional activity of AMPA receptors, we have measured AMPA-stimulated Ca-45(2) influx and 4beta-[H-3]phorbol 12,13-dibutyrate binding in intact cel ls. Both functional parameters increased along with the K+ concentrati on and were maximal in K40, in coincidence with the lowest expression of the GluR2 subunits. These results indicate that functional diversit y of AMPA receptors can be generated by the degree of chronic depolari zation and/or exposure to NMDA in neurons developing in primary cultur e.