Ca²⁺ regulation of glutamate release from inner hair cells of hearing mice

In our hearing organ, sound is encoded at ribbon synapses formed by inner hair cells (IHCs) and spiral ganglion neurons (SGNs). How the underlying synaptic vesicle (SV) release is controlled by Ca²⁺ in IHCs of hearing animals remained to be investigated. Here, we performed patch-clamp SGN recordings of the initial rate of release evoked by brief IHC Ca²⁺-influx in an ex vivo cochlear preparation from hearing mice. We aimed to closely mimic physiological conditions by perforated-patch recordings from IHCs kept at the physiological resting potential and at body temperature. We found release to relate supralinearly to Ca²⁺-influx (power, m: 4.3) when manipulating the [Ca²⁺] available for SV release by Zn²⁺-flicker-blocking of the single Ca²⁺-channel current. In contrast, a near linear Ca²⁺ dependence (m: 1.2 to 1.5) was observed when varying the number of open Ca²⁺-channels during deactivating Ca²⁺-currents and by dihydropyridine channel-inhibition. Concurrent changes of number and current of open Ca²⁺-channels over the range of physiological depolarizations revealed m: 1.8. These findings indicate that SV release requires ~4 Ca²⁺-ions to bind to their Ca²⁺-sensor of fusion. We interpret the near linear Ca²⁺-dependence of release during manipulations that change the number of open Ca²⁺-channels to reflect control of SV release by the high [Ca²⁺] in the Ca²⁺-nanodomain of one or few nearby Ca²⁺-channels. We propose that a combination of Ca²⁺ nanodomain control and supralinear intrinsic Ca²⁺-dependence of fusion optimally links SV release to the timing and amplitude of the IHC receptor potential and separates it from other IHC Ca²⁺-signals unrelated to afferent synaptic transmission.