Rasmussen_2014_Brain.Res_1542_1

Reference

Title : Biophysical and pharmacological characterization of alpha6-containing nicotinic acetylcholine receptors expressed in HEK293 cells - Rasmussen_2014_Brain.Res_1542_1
Author(s) : Rasmussen AH , Strobaek D , Dyhring T , Jensen ML , Peters D , Grunnet M , Timmermann DB , Ahring PK
Ref : Brain Research , 1542 :1 , 2014
Abstract :

Nicotinic acetylcholine receptors (nAChR's) containing the alpha6 subunit (alpha6) are putative drug targets of relevance to Parkinson's disease and nicotine addiction. However, heterologous expression of alpha6 receptors has proven challenging which has stifled drug discovery efforts. Here, we investigate potential new avenues for achieving functional alpha6 receptor expression. Combinations of chimeric and mutated alpha6, beta2 and beta3 subunits were co-expressed in the human HEK293 cell line and receptor expression was assessed using Ca(2+)-imaging (FLIPR) and whole-cell patch-clamp electrophysiology. Transient transfections of a chimeric alpha6/alpha3 subunit construct in combination with beta2 and beta3(V9'S) gave rise to significant acetylcholine-evoked whole-cell currents. Increasing the beta3(V9'S):beta2:alpha6/alpha3 cDNA ratio, resulted in a significantly higher fraction of cells with robust current levels. Using an excess of wild-type beta3, significant functional expression of alpha6/alpha3beta2beta3 was also demonstrated. Comparing the acetylcholine concentration-response relationship of alpha6/alpha3beta2beta3(V9'S) to that of alpha6/alpha3beta2beta3 revealed the beta3 point mutation to result in decreased current decay rate and increased ACh agonist potency. Ca(2+)-imaging experiments showed preservation of basic alpha6 receptor pharmacology. Our results establish that alpha6/alpha3beta2beta3(V9'S) replicate several basic features of native alpha6 receptors but also highlight several caveats associated with using this construct and may therefore provide guidance for future drug hunting efforts.

PubMedSearch : Rasmussen_2014_Brain.Res_1542_1
PubMedID: 24157862

Related information

Citations formats

Rasmussen AH, Strobaek D, Dyhring T, Jensen ML, Peters D, Grunnet M, Timmermann DB, Ahring PK (2014)
Biophysical and pharmacological characterization of alpha6-containing nicotinic acetylcholine receptors expressed in HEK293 cells
Brain Research 1542 :1

Rasmussen AH, Strobaek D, Dyhring T, Jensen ML, Peters D, Grunnet M, Timmermann DB, Ahring PK (2014)
Brain Research 1542 :1