L-Glutamate binding sites of parasitic nematodes: an association with ivermectin resistance?

Nematode membrane preparations contain high amounts of low-affinity specifi c L-glutamate binding sites. The numbers of these sites were increased in 2 isolates, one field-derived and the other laboratory-derived, of ivermecti n-resistant Haemonchus contortus and a field isolate of ivermectin-resistan t Telodorsagia circumcincta, when compared to control, drug-sensitive isola tes. Specific [H-3]ivermectin binding to these membrane preparations showed no differences between ivermectin-sensitive and resistant isolates and the number of ivermectin binding sites was approximately 100-fold less than th e number of L-glutamate binding sites. Kinetic analysis of L-glutamate bind ing suggested the presence of at least 2 classes of binding site. L-Glutama te binding was blocked by ibotenic acid, kynurenic acid and beta-hydroxyasp artate, but not by ivermectin, argiopine, kainate, quisqualate or NMDA. Com petition assays with ibotenic acid suggested that there were 2 distinct pop ulations of glutamate binding sites and that the site with the lower affini ty for ibotenate was upregulated in the ivermectin-resistant nematodes. In the field isolate of resistant H. contortus we found no coding changes in t he cDNAs encoding glutamate-gated chloride channel subunits HG2, HG3 and HG 4, nor were any changes in channel expression detected using subunit-specif ic antibodies. The low-affinity binding site is unlikely to be associated w ith the ivermectin receptor in these nematodes.