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Synaptic short-term plasticity is a key regulator of neuronal communication and is controlled via various mechanisms. A well established property of mossy fiber to CA3 pyramidal cell synapses is the extensive short-term facilitation during high-frequency bursts. We investigated the mechanisms governing facilitation using a combination of whole-cell electrophysiological recordings, electrical minimal stimulation, and random-access two-photon microscopy in acute mouse hippocampal slices. Two distinct presynaptic mechanisms were involved in short-term facilitation, with their relative contribution dependent on extracellular calcium concentration. The synchronization of multivesicular release was observed during trains of facilitating EPSCs recorded in 1.2 mM external Ca2+ ([Ca2+]e). Indeed, covariance analysis revealed a gradual augmentation in quantal size during trains of EPSCs, and application of the low-affinity glutamate receptor antagonist y-D-glutamylglycine showed an increase in cleft glutamate concentration during paired-pulse stimulation. Whereas synchronization of multivesicular release contributed to the facilitation in 1.2 mM [Ca2+]e, variance-mean analysis showed that recruitment of more release sites (JV) was likely to account for the larger facilitation observed in 2.5 mM [Ca2+]e. Furthermore, this increase in N could be promoted by calcium microdomains of heterogeneous amplitudes observed in single mossy fiber boutons. Our findings suggest that the combination of multivesicular release and the recruitment of additional release sites act together to increase glutamate release during burst activity. This is supported by the compartmentalized spatial profile of calcium elevations in boutons and helps to expand the dynamic range of mossy fibers information transfer. [ABSTRACT FROM AUTHOR]

Morphological studies have provided ample evidence for synaptic connections between cerebellar Purkinje cells (PCs), but the functional properties of these synapses remain elusive. We report on direct recordings of synaptically connected PC5 in mice cerebellar slices. In PCs filled with a fluorescent dye to aid axon visualization and postsynaptic target identification, presynaptic action potentials elicited unitary inhibitory postsynaptic currents in neighboring PCs in 10% of potential connections tested. In 11 pairs. postsynaptic currents had a delay onset of 1.62 ± 0.16 ms with respect to the presynaptic spike, a 10—90% rise time of 2.20 ± 0.33 ms. and a monoexponential decay with a time constant of 13.3 ± 1.7 ms. Average values for peak current and variance-to-mean ratio were 55 ± 14 and 30 ± 3 pA, respectively. In contrast to the depressing nature of the synapse between PC5 and deep cerebellar nuclei neurons, PC-PC synapses exhibited strong facilitation operating within a time window of a few milliseconds; paired-pulse ratios for 3-and 20-ms intervals were 1.79 ± 0.18 and 1.01 ± 0.14, respectively (n = 6). The facilitation is of presynaptic nature because it is accompanied by a decrease in failure rate. Trains of action potentials evoked in presynaptic varicosities volume-averaged calcium transients whose peak increased 1.7-fold as the frequency increased from 50 to 166 Hz. We suggest that PC-PC synapses are tuned for high fidelity of transmission during bursts of PC activity and that their operation in the cerebellar circuit modulates synchronized PC firing. [ABSTRACT FROM AUTHOR]

Activity-dependent elevation of calcium within presynaptic boutons regulates many aspects of synaptic transmission. Here, we examine presynaptic residual calcium (Cares) transients in individual presynaptic boutons of cerebellar granule cells at near-physiological temperatures using two-photon microscopy. Properties of Cares under conditions of zero-added buffer were determined by measuring Cares transients while loading boutons to a steady-state indicator concentration. These experiments revealed that, in the absence of exogenous calcium buffers, a single action potential evokes transients of Cares that vary widely in different boutons both in amplitude (400-900 nM) and time course (25-55 ms). Variation in calcium influx density, endogenous buffer capacity, and calcium extrusion density contribute to differences in Cares among boutons. Heterogeneity in Cares within different boutons suggests that plasticity can be regulated independently at different synapses arising from an individual granule cell. In a given bouton, Cares signals were highly reproducible from trial to trial and failures of calcium influx were not observed. We find that a factor contributing to this reliability is that an action potential opens a large number of calcium channels (20-125) in a bouton. Presynaptic calcium signals were also used to assess the ability of granule cell axons to convey somatically generated action potentials to distant synapses. In response to pairs of action potentials or trains, granule cell boutons showed a remarkable ability to respond reliably at frequencies up to 500 Hz. Thus, individual boutons appear specialized for reliable calcium signaling during bursts of high-frequency activation such as those that are observed in vivo. [ABSTRACT FROM AUTHOR]