AVERMECTIN B-1A BINDS TO HIGH-AFFINITY AND LOW-AFFINITY SITES WITH DUAL EFFECTS ON THE GAMMA-AMINOBUTYRIC ACID-GATED CHLORIDE CHANNEL OF CULTURED CEREBELLAR GRANULE NEURONS

Avermectin B-1a (AVM B-1a), a widely used insecticide and acaricide, i s reported to both activate and inhibit gamma-aminobutyric acid, (GABA (A)) receptor function in mammalian brain. This study attempts to reso lve these seemingly contradictory results by examining the binding pro perties of AVM B-1a and its effects on the GABA-gated chloride channel with primary cultures of rat cerebellar granule neurons as a model sy stem. Specific binding of [H-3]AVM B-1a in intact neuron cultures is t ime- and concentration-dependent and is displaceable by AVM analogs. S catchard analysis of [H-3]AVM B-1a binding reveals high- and low-affin ity sites with K-D values of 5 and 815 nM, respectively. AVM B-1a alte rs the binding of [H-3]ethynylbicycloorthobenzoate at the noncompetiti ve blocker site in a biphasic manner; activation is evident with 10 to 300 nM AVM B-1a after 5 to 10 min incubation and inhibition with an I C50 of 866 nM after 60 min incubation. Consistent with this observatio n, Cl-36(-) influx is stimulated by AVM B-1a at 3 to 100 nM and inhibi ted at 1 to 3 mu M. GABA-stimulated Cl-36(-) influx is completely bloc ked by both [H-3]ethynylbicycloorthobenzoate and 12-ketoendrin (two GA BA-gated chloride channel blockers) and AVM B-1a at 1 to 1.5 mu M. Als o, Cl-36(-) influx induced by AVM B-1a at 10 nM is suppressed by the t wo channel blockers. Thus, AVM B-1a binds to two different sites in th e GABA-gated chloride channel with dual effects, i.e., activating the channel on binding to the high-affinity site and blocking it on furthe r binding to the low-affinity site.