These data show that the Ca2+-CaM dependent Munc13-1 mediated replenishment of the rapidly releasable SV pool does not significantly affect SSD levels in young calyx of Held synapses. Because the relative contribution of mechanisms that define the steady-state EPSC amplitudes during train stimulation changes during postnatal maturation of the calyx (Crins et al., 2011; Erazo-Fischer et al., 2007; Sonntag et al., 2011; Taschenberger et al., 2002, 2005; Taschenberger and von Gersdorff, 2000; Wang et al., 2008), we tested whether STD differs between more mature WT and

Munc13-1W464R synapses. We measured SSD levels during trains of 25 APs at frequencies of 2–100 Hz in P14–P17 calyces. SSD levels in calyces of Munc13-1W464R Fulvestrant supplier mice this website were significantly lower than those of WT mice at all frequencies tested (Figures 7A–7C, S3E, and S3F), whereas the initial EPSC amplitudes were unchanged (100 Hz train; WT, 20.55 ± 2.3 nA, n = 16; Munc13-1W464R 24.3 ± 3.02 nA, n = 17; p > 0.05). The stronger SSD in P14–P17 Munc13-1W464R KI calyces was accompanied by significantly smaller PPRs in Munc13-1W464R mutants as compared to WT animals (Figure 7D). Presynaptic Ca2+ current amplitudes (WT, 1.85 ± 0.2 nA, n = 5; Munc13-1W464R, 1.96 ± 0.3 nA, n = 6; p > 0.05),

and facilitation of the Ca2+ current during trains of step depolarizations were similar in Munc13-1W464R and WT calyces (Figures 7E–7G), and therefore cannot account for the differences observed in pr. These data demonstrate that genetic perturbation of Ca2+-CaM signaling to Munc13-1 results in aberrant STD nearly in the calyx of Held after hearing onset, but not at calyces

of juvenile mice. STD during high-frequency AP trains is a feature of many synapses in the mammalian brain, including the calyx of Held (Figures 6 and 7). It primarily reflects a transient and activity dependent decrease in neurotransmitter release, which can be caused by several different processes, including reduced Ca2+ influx into presynaptic terminals (Xu and Wu, 2005), changes in the AP waveform (Geiger and Jonas, 2000), depletion of the RRP of SVs (Rosenmund and Stevens, 1996; Sakaba and Neher, 2001; Wu and Borst, 1999), and delayed clearance of SV release sites (Hosoi et al., 2009). STD is counteracted by the SV priming machinery, which consists of Munc13 and CAPS proteins and determines the rate of RRP refilling and the RRP size after strong stimulation (Augustin et al., 1999b; Jockusch et al., 2007; Junge et al., 2004; Rhee et al., 2002; Rosenmund et al., 2002; Varoqueaux et al., 2002).