Co-release of GABA and ACh from Medial Olivocochlear

During development, inner hair cells (IHCs) in the mammalian cochlea are unresponsive to acoustic stimuli but instead exhibit spontaneous activity. During this same period, neurons originating from the medial olivocochlear (MOC) complex transiently innervateIHCs, regulating their firing pattern which is crucial for the correct development of the auditory pathway. Although the MOC-IHC isa cholinergic synapse, previous evidence indicates the widespread presence of gamma-aminobutyric acid (GABA) signaling markers,including presynaptic GABAB receptors (GABABR). In this study, we explore the source of GABA by optogenetically activating eithercholinergic or GABAergic fibers. The optogenetic stimulation of MOC terminals from GAD;ChR2-eYFP and ChAT;ChR2-eYFP mice(of either sex) evoked synaptic currents in IHCs, which were blocked by α-bungarotoxin. This suggests that GABAergic fibers releaseacetylcholine (ACh) and activate α9α10 nicotinic acetylcholine receptors (nAChRs). Additionally, MOC cholinergic fibers release notonly ACh but also GABA, as the effect of GABA on ACh response amplitude was prevented by applying a GABABR blocker. Usingoptical neurotransmitter detection and calcium imaging techniques, we examined the extent of GABAergic modulation at the singlesynapse level. Our findings suggest heterogeneity in GABA modulation, as only 15 out of 31 recorded synaptic sites were modulatedby applying the GABABR specific antagonist, CGP 35348 (100−200 μM). In conclusion, we provide evidence indicating that GABAand ACh are co-released from at least a subset of MOC terminals. In this circuit, GABA functions as a negative feedback mechanism,locally regulating the extent of cholinergic inhibition at certain efferent→IHC synapses during an immature stage.