Calcium permeability of non-N-methyl-D-aspartate receptor channels in immature cerebellar Purkinje cells: studies using fura-2 microfluorometry.

Using fura-2 microfluorometry, I investigated the mechanism by which non-N-methyl-D-aspartate (NMDA) receptor agonists increase the cytosolic free calcium concentration ([Ca]in) in single cerebellar Purkinje cells isolated from 3-10-day-old rats. Kainate and alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionate dose-dependently increased the cytosolic free Na+ concentration, which was measured using sodium-binding benzofuran isophthalate microfluorometry, confirming the Na+ influx through ion channels linked to non-NMDA receptors. The [Ca2+] increases induced by relatively lower concentrations of agonists were entirely dependent on external Ca2+ and were reduced by removal of external Na+ or by addition of a Ca2+ channel blocker, D600. The results indicate that the non-NMDA agonist-induced [Ca]in increase was due mainly to Ca2+ influx through voltage-dependent Ca2+ channels, which were activated by a massive Na+ influx. On the other hand, higher concentrations of agonists dose-dependently increased [Ca]in under conditions in which activation of voltage-dependent Ca2+ channels were blocked by a combination of Na+ removal with D600. These [Ca]in increases were Ca2+ dependent and little affected by adding a competitive NMDA antagonist. Non-NMDA agonists also stimulated influxes of Mn2+ and Co2+, both of which can be monitored by quenching fura-2 fluorescence under the same conditions. These results suggest that ion channels linked to non-NMDA receptors on immature Purkinje cells are permeable to Ca2+, Mn2+, and Co2+.