Using [3H]flumazenil as a probe we investigated how benzodiazepine site pharmacology of alpha beta gamma ternary combinations of GABAA receptors can be influenced upon expression of different isoforms of alpha, beta and gamma subunits. The nature of the beta subunit did not alter the pharmacology of this site in that the affinities of alpha 5-containing GABAA receptors for various benzodiazepine modulatory ligands were essentially unchanged upon a comparison of different beta-variant forms (alpha 5 beta 1 gamma 2, alpha 5 beta 2 gamma 2 and alpha 5 beta 3 gamma 2). In contrast, both alpha and gamma variants contributed to notable differences in benzodiazepine site pharmacology. Thus alpha 1 beta 2 gamma 2, alpha 3 beta 2 gamma 2 and alpha 5 beta 2 gamma 2 receptors showed high, intermediate and low affinities for zolpidem, respectively. Exchanging gamma 2 for gamma 3 reduced the affinities of alpha 1 beta 2 gamma and alpha 3 beta 2 gamma receptors for zolpidem by factors of > 150 and > 5.8, respectively. The alpha 1 beta 2 gamma 3, alpha 3 beta 2 gamma 3 and alpha 5 beta 2 gamma 3 receptors exhibited, in contrast, higher affinity for CL218872 than their corresponding gamma 2 receptors. The information on these different recombinant GABAA receptor pharmacological profiles should help in the elucidation of native GABAA receptor subtype diversity.