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Abstract

Voltage-gated Ca²⁺ channels (VGCCs) of the P/Q-type, which are expressed at a majority of mammalian nerve terminals, show two types of Ca²⁺-dependent feedback regulation—inactivation (CDI) and facilitation (CDF). Because of the nonlinear relationship between Ca²⁺ influx and transmitter release, CDI and CDF are powerful regulators of synaptic strength. To what extent VGCCs inactivate or facilitate during spike trains depends on the dynamics of free Ca²⁺ ([Ca²⁺]i) and the Ca²⁺ sensitivity of CDI and CDF, which has not been determined in nerve terminals. In this report, we took advantage of the large size of a rat auditory glutamatergic synapse—the calyx of Held—and combined voltage-clamp recordings of presynaptic Ca²⁺ currents (I_Ca(V)) with UV-light flash-induced Ca²⁺ uncaging and presynaptic Ca²⁺ imaging to study the Ca²⁺ requirements for CDI and CDF.



We find that nearly half of the presynaptic VGCCs inactivate during 100 ms voltage steps and require several seconds to recover. This inactivation is caused neither by depletion of Ca²⁺ ions from the synaptic cleft nor by metabotropic feedback inhibition, because it is resistant to blockade of metabotropic and ionotropic glutamate receptors. Facilitation of I_Ca(V) induced by repetitive depolarizations or preconditioning voltage steps decays within tens of milliseconds. Since Ca²⁺ buffers only weakly affect CDI and CDF, we conclude that the Ca²⁺ sensors are closely associated with the channel. CDI and CDF can be induced by intracellular photo release of Ca²⁺ resulting in [Ca²⁺]i elevations in the low micromolar range, implying a surprisingly high affinity of the Ca²⁺ sensors.