Received for publication May 16, 2006.
Revised June 30, 2006.
Accepted for publication July 5, 2006.

Incorporation of the 3 subunit has a dominant negative effect on the function of recombinant central-type neuronal nicotinic receptors

Abstract

The 3 neuronal nicotinic subunit is localised in dopaminergic areas of the central nervous system, where many other neuronal nicotinic subunits are expressed. So far, 3 has only been shown to form functional receptors when expressed together with the 3 and 4 subunits. We have systematically tested in Xenopus oocytes the effects of co-expressing human 3 with every pairwise functional combination of neuronal nicotinic subunits likely to be relevant to the CNS. Expression of 7 homomers or / pairs (2, 3, 4 or 6 together with 2 or 4, produced robust nicotinic currents for all combinations, save 6 2 and 6 4. Co-expression of wild-type 3 led to a nearly complete loss of function (measured as maximum current response to ACh) for 7 and for all functional / pairs except 34. This effect was also seen in hippocampal neurones in culture, which lost their robust 7-like responses when transfected with 3. The level of surface expression of nicotinic binding sites (34, 4 2 and 7) in tsA201 cells was only marginally affected by 3 expression. Furthermore, the dominant-negative effect of 3 was abolished by a VS mutation in the 9' position of the second transmembrane domain of 3, a mutation thought to facilitate channel gating. Our results show that incorporation of 3 into neuronal nicotinic receptors other than 34 has a powerful dominant negative effect, probably due to impairment in gating. This raises the possibility of a novel regulatory role for the 3 subunit on neuronal nicotinic signalling in the central nervous system.

Key words: Nicotinic cholinergic, Func. analysis receptor/ion channel mutants

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