Synaptic vesicle fusion at hair cell ribbon active zones is triggered by Ca2+ entry through L-type (CaV1.3) Ca2+ channels (Platzer et al 2000, Cell 102:89) in response to sound-induced graded receptor potentials. Although ribbon synapses become more Ca2+ efficient with maturation (Johnson et al. 2005 J Physiol 563:177; Wong et al 2014, EMBO J 33:247), how Ca2+ is able to regulate exocytosis at mature ribbon synapses is still mostly undetermined. Calcium nanodomain coupling between a few Ca2+ channels and the readily releasable synaptic vesicles has been proposed to control exocytosis in vertebrate hair cells (Graydon et al 2011, J Neurosci 31:16637; Wong et al 2014). This tight-coupling has the advantage of providing accurate temporal encoding for phase-locking. However, another hypothesis is that the exocytotic coupling is controlled by many channels cooperatively (Ca2+ microdomain) and it is the molecules intrinsic to the synaptic machinery (Ca2+ sensors) that generate the highly Ca2+ efficient exocytosis at mature ribbon synapses (Johnson et al 2010, Nat Neurosci 13:45). These two release mechanisms may in fact co-exist along the same auditory organ, thus emphasizing the different frequency components of the cell’s in vivo receptor potential, respectively. Whole-cell patch-clamp recordings were used to investigate exocytosis in hair cells at specific characteristic frequencies (CF) of the mature gerbil, mouse and bullfrog auditory organs. The physiological coupling between Ca2+ influx and the synaptic machinery was investigated using different intracellular concentrations of EGTA, which buffers increases in intracellular Ca2+ only relatively far away from its source (Neher 1998 Neuron 20:389). Experiments were performed at body temperature and using 1.3 mM extracellular Ca2+ and following UK and USA animal regulations. We show that the coupling between Ca2+ channels and release sensors change as a function of the cell’s frequency position. While low-frequency hair cells (<2 kHz), which are phase-locked to sound stimuli, exhibit a tight, nanodomain, coupling between Ca2+ channels and synaptic vesicles, high-frequency cells have a much more loose coupling, which becomes progressively more microdomain along the gerbil cochlea. We also showed that the level of intracellular Ca2+ buffer affects the speed of recovery from paired-pulse depression. Our findings show that both the nanodomain and microdomain coupling are present in mature auditory hair cells, the function of which is to preserve the precise temporal coding of sound in phase-locked low-frequency hair cells and stimulus intensity in high-frequency cells, respectively.