Glutamate-gated chloride channels (GluCls) are members of the cys-loop ligand-gated ion channel superfamily whose presence has been reported in a variety of invertebrate tissues. In the honeybee, a single gene, amel_glucl, encoding a GluClα subunit, was found in the genome but both the pattern of expression of this gene in the bee brain and its functional role remained unknown. Here we localised the expression sites of the honeybee GluClα subunit at the mRNA and protein levels. To characterise the functional role of GluCls in the honeybee brain, we studied their implication in olfactory learning and memory by means of RNA interference (RNAi) against the GluClα subunit. We found that the GluClα subunit is expressed in the muscles, the antennae and the brain of honeybees. Expression of the GluClα protein was necessary for the retrieval of olfactory memories; more specifically, injection of dsRNA or siRNA resulted in a decrease in retention performances ∼24 h after injection. Knockdown of GluClα subunits impaired neither olfaction nor sucrose sensitivity, and did not affect the capacity to associate odor and sucrose. Our data provide the first evidence for the involvement of glutamate-gated chloride channels in olfactory memory in an invertebrate.