Many sensory systems, such as the vestibular (Arenz et al., 2008; Bagnall et al., 2008
), proprioceptive (van Kan et al., 1993
), somatosensory (Jörntell and Ekerot, 2006
), auditory (Lorteije et al., 2009
), and visual (Azouz et al., 1997
) systems, exploit a broad bandwidth of action potential frequencies to represent information as sustained rate codes. Synapses in sensory organs typically employ large, vesicle-tethering, electron-dense cytomatrix structures at their active zones (AZs), the sites where vesicles dock and fuse to release their neurotransmitter content into the synaptic cleft (Südhof, 2004
). These electron-dense structures are decorated with vesicles and vary in size and shape in a species- and cell type-specific manner (Zhai and Bellen, 2004
). Some extend vertically into the cytoplasm and are referred to as ribbons (Lenzi and von Gersdorff, 2001
). These cytomatrix structures are thought to be critical for rapid and sustained vesicle supply at these specialized synapses, which transmit graded signals (Khimich et al., 2005; von Gersdorff et al., 1998
). In contrast, central rate-coded synapses have less prominent cytomatrix structures, but some can nevertheless maintain signaling over a wide bandwidth of action potential frequencies with a relatively small number of conventional release sites (Saviane and Silver, 2006
). This is achieved by a large pool of vesicles and rapid vesicle reloading to the AZ (Saviane and Silver, 2006
), but the molecular mechanisms underlying this rapid reloading are unknown.
To date, at least five protein families have been characterized whose members are highly enriched at the cytomatrix of the AZs: Munc13s, RIMs, ELKS/CAST proteins, Piccolo and Bassoon, and the liprins-α (Kaeser et al., 2009; Schoch and Gundelfinger, 2006
). Bassoon is a very large coiled-coil protein of ~4000 amino acids (~400 kDa) and is one of the core components of the cytomatrix at the AZ of both excitatory and inhibitory synapses (tom Dieck et al., 1998; Wang et al., 2009
). Interestingly, whereas other AZ proteins (e.g., RIMs) are present in both vertebrates and invertebrates (e.g., C. elegans
, homologs of Bassoon and Piccolo (also named Aczonin; Wang et al., 2009
) appear to be de novo developments of vertebrates (Altrock et al., 2003
). At ribbon-type synapses, deletion of Exons 4 and 5 of the Bassoon gene leads to disrupted assembly of the cytomatrix at the AZ (Dick et al., 2003
) as well as impaired auditory signaling (Buran et al., 2010; Khimich et al., 2005
). At conventional synapses Bassoon is involved in trafficking and synaptic delivery of AZ material (Fejtova et al., 2009
) and in partially silencing synapses (Altrock et al., 2003
). However, the function of Bassoon in synaptic transmission remains unclear.
We investigated the role of Bassoon by comparing the properties of transmission at cerebellar mossy fiber to granule cell (MF-GC) synaptic connections in control and Bassoon null mutant (Bsn−/−
) mice. These glutamatergic synapses appear ideally suited to investigate the mechanisms of vesicle reloading because they show rapid vesicle reloading at a limited number of release sites (Saviane and Silver, 2006
). In addition, MF-GC synapses are characterized by highly synchronized vesicular release (Sargent et al., 2005
), a large pool of releasable vesicles (Saviane and Silver, 2006
), and firing frequencies of more than 700 Hz in vivo (Rancz et al., 2007
). The excellent voltage clamp afforded by the postsynaptic granule cell leads to excitatory postsynaptic currents (EPSCs) with rise and decay kinetics in the submillisecond range with only modest desensitization (DiGregorio et al., 2007
), facilitating the analysis of high-frequency signaling.
Here, we show that spontaneous EPSCs and EPSCs evoked at low frequencies are normal at MF-GC synapses in Bsn−/− mice compared to those in control mice. However, the lack of Bassoon caused a pronounced depression during high-frequency transmission that occurred within milliseconds and a delayed recovery from depression. Analysis of the presynaptic and postsynaptic mechanisms of short-term plasticity revealed that the rate of vesicle reloading at AZs of MF-GC terminals was almost halved in Bsn−/− mutants compared with controls. Thus, our data demonstrate that the cytomatrix protein Bassoon speeds high-rate vesicle reloading at AZs of a central excitatory synapse, significantly increasing the achievable rate of transmission.