Steve’s primary interest, however, was to understand the structure and properties of these acetylcholine receptors.

In 1970, Jean-Pierre Changeux and coworkers used biochemical methods to characterize the nAChR as a ligand-gated ion channel formed by five homologous subunit proteins. In order to isolate DNA complementary to the mRNA encoding specific receptor subunit proteins, Steve decided to use a strategy based on recently developed molecular cloning methods. Biochemical methods had previously yielded short stretches DNA Damage inhibitor of amino acid sequences from one of the muscle nAChR proteins; using this information, one could clone the full-length sequence encoding channel subunits from a suitable library of cDNAs. He was on the right path, but just not on time, because in 1982 a research group led by Shosaku Numa cloned the first cDNA encoding a nAChR subunit from the electric organ of the ray Torpedo californica. Having just lost a battle, but not the war, Steve turned his attention to brain ligand-gated ion channels together with Jim Boulter LBH589 and Jim Patrick. The scarcity of the nAChR protein in the brain precluded biochemical derivation of partial peptide sequences, however, and a different tactic was required. Relying on

the assumption that neuronal nAChRs should have some degree of identity with receptor subunits present at the neuromuscular junction, and using low-stringency C-X-C chemokine receptor type 7 (CXCR-7) molecular hybridization techniques, the Heinemann group was able to clone the first neuronal nicotinic receptor subunit, which is now named α3. This was followed by the cloning

of the entire family of neuronal nicotinic receptor α and β subunits, the characterization of channel properties when expressed in heterologous systems, and the differential distribution of receptor subunits in the brain. Finally, in 1994 the α9 nicotinic cholinergic receptor was cloned in the Heinemann laboratory, deciphering the molecular identity of the cholinergic receptor that mediates efferent inhibition of sound amplification within the inner ear. The nicotinic receptor subunit clones were immediately licensed to the pharma industry for the development of drugs to be used in neurological disorders and tobacco addiction. As increasing numbers of nicotinic and the structurally related GABA receptor subunit cDNAs began to yield to maturing techniques in molecular neuroscience, a new set of targets became the object of a feverish cloning race between several laboratories, including that of Steve Heinemann, in the late 1980s and early 1990s. These receptors were, of course, those that mediated excitatory signaling in the CNS, the ionotropic glutamate receptors (iGluRs).