Rodent habenulo-interpeduncular pathway expresses a large variety of uncommon nAChR subtypes, but only the alpha3beta4* and alpha3beta3beta4* subtypes mediate acetylcholine release.

Recent studies suggest that the neuronal nicotinic receptors (nAChRs) present in the habenulo-interpeduncular (Hb-IPn) system can modulate the reinforcing effect of addictive drugs and the anxiolytic effect of nicotine. Hb and IPn neurons express mRNAs for most nAChR subunits, thus making it difficult to establish the subunit composition of functional receptors. We used immunoprecipitation and immunopurification studies performed in rat and wild-type (+/+) and beta 2 knock-out (-/-) mice to establish that the Hb and IPn contain significant beta 2* and beta 4* populations of nAChR receptors (each of which is heterogeneous). The beta 4* nAChR are more highly expressed in the IPn. We also identified novel native subtypes (alpha 2 beta 2*, alpha 4 beta 3 beta 2*,alpha 3 beta 3 beta 4*, alpha 6 beta 3 beta 4*). Our studies on IPn synaptosomes obtained from +/+ and alpha 2, alpha 4, alpha 5, alpha 6, alpha 7, beta 2, beta 3, and beta 4(-/-) mice show that only the alpha 3 beta 4 and alpha 3 beta 3 beta 4 subtypes facilitate acetylcholine (ACh) release. Ligand binding, immunoprecipitation, and Western blotting studies in beta 3(-/-) mice showed that, in the IPn of these mice, there is a concomitant reduction of ACh release and alpha 3 beta 4* receptors, whereas the receptor number remains the same in the Hb. We suggest that, in habenular cholinergic neurons, the beta 3 subunit may be important for transporting the alpha 3 beta 4* subtype from the medial habenula to the IPn. Overall, these studies highlight the presence of a wealth of uncommon nAChR subtypes in the Hb-IPn system and identify alpha 3 beta 4 and alpha 3 beta 3 beta 4, transported from the Hb and highly enriched in the IPn, as the subtypes modulating ACh release in the IPn