Presynaptic NMDARs in Cerebellar Interneurons Figure 5. Axonal Ca2+ signals elicited by local activation of pre-NMDARs. A: Representative experiment. Top: Fluorescence image (left) and reconstruction (right) of a MLI filled with Alexa 488 (20 mM) and OGB-1 (50 mM). Bottom: Local Ca2+ transients and associated somatic currents obtained in response to axonal (left) and dendritic (right) glutamate release (1 ms laser pulses; 0.9 mM MNI-glutamate; 5 mM NBQX; 0.5 mM TTX). B: Summary data. Left: In responsive axonal spots, the amplitude of Ca2+ transients are correlated to the corresponding somatic current (correlation coefficient R = 0.74). Right: Ca2+ transients elicited in TTX by local glutamate uncaging (Uncaging) had peak amplitudes similar to those obtained in the same axonal spots before TTX application using 4 propagated action potentials (Action potentials). Experiments were carried out in normal extracellular [Mg2+] (1 mM). doi:10.1371/journal.pone.0039983.g005 receptors [35]. In the presence of 300 nM Zn2+, NMDA (30 mM) did not significantly increase the frequency of mIPSC (frequency ratio over control of 2.1262.00, n = 4; Fig. 6D). Additionnally ifenprodil, an NR2B selective antagonist, was used to examine the possible involvement of this subunit in the effect of NMDA application. When slices were treated with 10 mM ifenprodil, application of NMDA (30 mM) still increased the frequency of mIPSCs (frequency ratio over control of 3.5361.57; Fig. 6 C and D), albeit to a smaller extent than in control conditions. These results indicate that both NR2A and NR2B participate in the effects elicited by NMDA on mIPSCs. Because of the high input resistance of MLIs and the rather small volume of their presynaptic terminals, even a small glutamate-evoked current might be sufficient to alleviate Mg2+ block. It is important to PLoS ONE | www.plosone.org 8 June 2012 | Volume 7 | Issue 6 | e39983