CHOLINERGIC AND PURINERGIC SIGNALING IN OUTER HAIR-CELLS OF MAMMALIANCOCHLEA

Outer (OHC) and inner (IHC) hair cells in the organ of Corti of the ma mmalian cochlea process sound. OHC and their efferent synapse are part of a feedback system assumed to control and modulate information carr ied by afferent neurons passing from IHC to the brain. Underlying mech anisms are not well understood. This paper discusses recent progress. In vivo and in vitro information is presented on structure, pharmacolo gy, function and localization of the pre- and postsynaptic acetylcholi ne receptors (AChRs) at the efferent synapse. Recent data are given on a presynaptic M3 AChR subtype, probably an autoreceptor involved in t ransmitter release. Data from our lab on specific binding of [H-3]-qui nuclidinyl benzilate ([H-3]3-QNB) to non-enzymatically isolated guinea pig OHC reveal a K-D several 100 x higher than that for any known mus carinic receptor subtype, including the above-mentioned presynaptic mu scarinic AChR of the OHC efferent synapse. The extremely high concentr ations of [H-3]3-QNB needed for any binding at all to OHC thus rule ou t presynaptic membrane impurities as the cause of such binding, and al so the presence of a typical mAChR subtype on OHC. The number of [H-3] 3-QNB binding sites (similar to 10(6)/OHC) we found on OHC was 1/10th of that we found for binding of nicotinic ligands to OHC, further maki ng it questionable that an ACh-binding site on OHC binds [H-3]3-QNB. O bservations may instead point to the possibility of another binding si te, e.g. an (allosteric) site involved with the as yet not understood 'weak' muscarinic properties of the OHC AChR. Further new data on the OHC AChR confirm reversible sc-bungarotoxin, nicotine and d-tubocurari ne binding. [H-3]alpha-Bungarotoxin and [H-3]-nicotine binding sites a re estimated at similar to 6.10(7) sites/OHC. Strychnine, a glycine re ceptor blocker suggested to interfere with cholinergic sites of the ef ferent OHC synapse, was found to bind to OHC (cold strychnine for unsp ecific binding). This binding, not seen in the presence of high [glyci ne], increased in the presence of. depolarizing [K+], while ACh (100 m u M) had no significant effect. Results suggest strychnine binding to the outside of OHC, but also to sites accessible only after cell depol arization, possibly to the hyperpolarizing Ca2+-dependent K+ channel. Recent molecular cloning of the OHC AChR indicates a novel alpha-subun it. An often observed ACh-activated Ca2+-influx close to zero into OHC leaves an unanswered question. OHC also carry P-2-purinergic receptor s (P(2)Rs), a more rapid ionotropic P(2z)R-like subtype and a quantita tively dominating slow metabotropic P(2y)R subtype coupled to a G prot ein-phospholipase C cascade and not desensitized. Both contribute to i ncreased cytoplasmic [Ca2+], from respectively external and internal s ources, Whether or not such receptors are part of efferent synaptic ac tivity is unknown; their localization on the OHC plasma membrane is so far only indirect and synaptic vesicles of the efferent nerve endings have not yet been analyzed for their ATP content. Localization, funct ion and interaction of [Ca2+] increases triggered by, respectively, AT P and ACh are currently studied in this laboratory.