THE PRODUCTION OF A STABLY TRANSFORMED INSECT-CELL LINE EXPRESSING ANINVERTEBRATE GABA(A) RECEPTOR BETA-SUBUNIT

We have produced a stable insect cell line derived from Spodoptera fru giperda (Sf9) cells expressing a cDNA encoding a beta-subunit of the L ymnaea stagnalis GABA(A) receptor. The cDNA was randomly integrated in to the insect cell genome under the control of a baculovirus immediate early gene (IE-1) promoter. Stable cell lines were established by tra nsformation of Sf9 cells with the expression vector pIEK1.LG beta 1 to gether with a plasmid encoding a selectable marker which confers neomy cin (G418) resistance. Following growth in the presence of G418, neomy cin resistant clones were selected, amplified and analysed for the pre sence of functional GABA-gated chloride channels. Electrophysiological analysis of one cell line showed the presence of a picrotoxin-sensiti ve chloride channel not present in control Sf9 cells. These channels w ere also sensitive to GABA, albeit at relatively high (mM) concentrati ons.