No ancillary proteins are essential for the functional heterologous expression of the homomeric α6 nicotinic acetylcholine receptor from the brown castor tick Ixodes ricinus.

Nicotinic acetylcholine receptors (nAChRs) are ligand-gated ion channels that facilitate rapid neural signaling. nAChRs respond to acetylcholine, their natural agonist, as well as to nicotine and neonicotinoids, the latter used to control ectoparasites and agricultural pests. The nAChRs are well studied in mammals and some arthropods; however, the nAChR subunits in the ectoparasitic tick Ixodes ricinus remain less understood. This study examined the α6 nAChR subunit in I. ricinus at both the molecular and functional levels. RT-PCR identified two alternative splice variants of the α6 subunit: Iricα6A, which includes two variants of exon 3 (3a and 3b) and Iricα6B, which contains only 3b. Protein structure predictions identified a loop of 15 amino acids within the extracellular domain of Iricα6A, originating from exon 3a, that is missing in Iricα6B; however, the role of this loop in receptor non-functionality remains undetermined. In functional studies using Xenopus laevis oocytes, two-electrode voltage-clamping revealed that only Iricα6B forms a functional homomeric receptor and does not require ancillary proteins. The Iricα6B EC50 for ACh was estimated to be 2 μM (n = 7 different recordings from two different batches of oocytes). The neonicotinoids dinotefuran and imidacloprid exhibited varying potencies and efficacies, with EC50 values of 0.42 nM (n = 6 different recordings from two different batches of oocytes) and 11.6 nM (n = 6 different recordings from two different batches of oocytes), respectively, and maximum amplitudes of 112% and 81% relative to ACh, respectively. Nicotine and two additional neonicotinoids also elicited responses in the Iricα6B receptor.