Assembly of human neuronal nicotinic receptor alpha 5 subunits with alpha 3, beta 2, and beta 4 subunits

Nicotinic acetylcholine receptors formed from combinations of alpha 3, beta 2, beta 4, and alpha 5 subunits are found in chicken ciliary ganglion neurons and some human neuroblastoma cell lines, We studied the co-expression of various combinations of cloned human alpha 3, beta 2, beta 4, and alpha 5 subunits in Xenopus oocytes, Expression on the surface membrane was found only for combinations of alpha 3 beta 2, alpha 3 beta 4, alpha 3 beta 2 alpha 5, and alpha 3 beta 4 alpha 5 subunits but not for other combinations of one, two, or three of these subunits, alpha 5 subunits assembled inside the oocyte with beta 2 but not with alpha 3 subunits or other alpha 5 subunits, alpha 5 subunits coassembled very efficiently with alpha 3 beta 2 or alpha 3 beta 4 combinations, The presence of alpha 5 subunits had very little effect on the binding affinities for epibatidine of receptors containing also alpha 3 and beta 2 or alpha 3 and beta 4 subunits, The presence of alpha 5 subunits increased the rate of desensitization of both receptors containing also alpha 3 and beta 2 or alpha 3 and beta 4 subunits, In the case of receptors containing alpha 3 and beta 4 subunits, the addition of alpha 5 subunits had little effect on the responses to acetylcholine or nicotine, However, in the case of receptors containing alpha 3 and beta 2 subunits, the addition of alpha 5 subunits reduced the EC(50) for acetylcholine from 28 to 0.5 mu M and the EC,, for nicotine from 6.8 to 1.9 mu M, while increasing the efficacy of nicotine from 50% on alpha 3 beta 2 receptors to 100% on alpha 3 beta 2 alpha 5 receptors, Both alpha 3 beta 2 and alpha 3 beta 2 alpha 5 receptors expressed in oocytes sedimented at the same 11 S value as native alpha 3-containing receptors from the human neuroblastoma cell line SH-SY5Y, In the receptors from the neuroblastoma alpha 3, beta 2, and alpha 5 subunits were co-assembled, and 56% of the receptor subtypes containing alpha 3 subunits also contained beta 2 subunits, The beta 2 subunit-containing receptors from SH-SY5Y cells exhibited the high affinity for epibatidine characteristic of receptors formed from alpha 3 and beta 2 or alpha 3, beta 2, and alpha 5 subunits rather than the low affinity exhibited by receptors formed from alpha 3 and beta 4 or alpha 3, beta 4, and alpha 5 subunits, Nicotine, like the structurally similar toxin epibatidine, also distinguishes by binding affinity two subtypes of receptors containing alpha 3 subunits in SH-SY5Y cells, The affinities of alpha 3 beta 2 receptors expressed in oocytes were similar to the affinities of native alpha 3 containing receptors from SH-SY5Y cells for acetylcholine, cytisine, and 1,1-dimethyl-4-phenylpip erazinium.