Muscarinic ACh receptor activation causes transmitter release from isolated frog vestibular hair cells

In the frog, vestibular efferent fibers innervate only type-II vestibular hair cells. Through this direct contact with hair cells, efferent neurons are capable of modifying transmitter release from hair cells onto primary vestibular afferents. The major efferent transmitter, acetylcholine (ACh), is known to produce distinct pharmacological actions involving several ACh receptors. Previous studies have implicated the presence of muscarinic ACh receptors on vestibular hair cells, although, surprisingly, a muscarinic-mediated electrical response has not been demonstrated in solitary vestibular hair cells. This study demonstrates that muscarinic receptors can evoke transmitter release from vestibular hair cells. Detection of this release was obtained through patch-clamp recordings from catfish cone horizontal cells, serving as glutamate detectors after pairing them with isolated frog semicircular canal hair cells in a two-cell preparation. Although horizontal cells alone failed to respond to carbachol, application of 20 microM carbachol to the two-cell preparation resulted in a horizontal cell response that could be mimicked by exogenous application of glutamate. All of the horizontal cells in the two-cell preparation responded to 20 microM CCh. Furthermore, this presumed transmitter release persisted in the presence of d-tubocurarine at concentrations that block all known hair cell nicotinic ACh receptors. The effect on the detector cell, imparted by the carbachol application to the hair cell-horizontal cell preparation, was blocked both by 2-amino-5-phosphonopentanoic acid, a selective N-methyl-D-aspartate antagonist, and the muscarinic antagonist, atropine. Thus vestibular hair cells from the frog semicircular canal can be stimulated to release transmitter by activating their muscarinic receptors.