An investigation into neuronal nicotinic receptors with complex compositions
Neuronal nicotinic acetylcholine receptors (nAChRs) are ligand-gated ion channels of the nicotinic superfamily. They are found widely in the autonomic nervous system and in selected regions of the central nervous system and are thought to be of importance in the progression and/or treatment of a number of disorders. Despite all this the exact role of cholinergic neurotransmission remains unclear and current cholinergic drug treatments leave a lot to be desired. As such nAChRs remain areas of immense interest and potential. One of the problems with nAChR research has been that the simple receptors obtained in heterologous expression systems probably bear little resemblance to the more complex stoichiometrics seen in vivo. Therefore my PhD dealt with the characterisation of some more complex nAChR compositions and the development of research tools to aid in the research of not just nAChRs but other ligand-gated and voltage-gated ion channels with complex compositions. In this I primarily used two-electrode voltage clamp methods in Xenopus laevis oocytes. My initial research investigated the role of P3 subunit incorporation into a range of nAChRs surprisingly this subunit, which produced only subtle effects when incorporated into a3p4 receptors, completely abolished the currents produced by all other pair and homomer receptors tested. Similar findings were obtained when hippocampal neurones in primary culture (which have a7-like responses) were transfected with the P3 subunit. This suggests a possible major role for P3 in nAChR regulation. One widely used method of constraining receptor stoichiometry is the use of "tandem" subunits. My research however uncovered serious flaws in this method which may render it worse than useless. Subsequent work showed that expression of "pentamer" constructs has greater potential, as it allows much greater control over receptor composition yet avoids the problems seen with the tandem approach.