The Role of Mobile Calcium Buffers in Synaptic Transmission at the Inner Hair Cell Ribbon Synapse

Abstract

Temporally precise sound encoding at the inner hair cell (IHC) ribbon synapse is tightly regulated by calcium. The mobile calcium buffers calbindin, parvalbumin alpha and calretinin might contribute to shaping the presynaptic Ca2+ signals. We investigated the function of these calcium binding proteins in IHC synaptic transmission by examining the auditory phenotype of double and triple buffer knockout mice. Our results show that buffer deficiency does not significantly alter hearing thresholds; however, we observed a slight increase in peak and steady-state sound-driven spike rates of spiral ganglion nerve fibers in knockout mice. The presynaptic function of IHCs was first studied by perforated patch-clamp recordings of Ca2+ currents and exocytic membrane capacitance increments. The absence of mobile calcium buffering proteins augmented sustained exocytosis in IHCs while leaving the amplitude and kinetics of exocytosis of the readily-releasable pool unchanged. Further, Ca2+-dependent inactivation of calcium currents was stronger in IHCs of triple buffer knockout mice. In ruptured patch experiments we then tried to restore the calcium buffer capacity by adding exogeneous buffers. We estimated the concentration of endogenous buffers in IHCs to be equivalent to 0.5-1 mM BAPTA, which agrees well with previous estimates obtained by quantitative immunogold electron microscopy (Hackney et al., 2005). Our results demonstrate that calbindin, parvalbumin alpha and calretinin are involved in the regulation of synaptic transmission at the IHC ribbon synapse; however they do not seem to be essential for hearing, at least not during transient sound stimulation.