Fig. 8. Localization of postsynaptic Ca2+ entry through NMDAR channels and VDCCs.a, Infrared video image (left) of an MNTB synapse from which a simultaneous presynaptic and postsynaptic recording was done. The presynaptic terminal was loaded with MagFura-2 (0.4 mm) by the pipette on the right, and the postsynaptic neuron was loaded with OGB-5N (0.4 mm). The right image shows the overlay of the presynaptic and postsynaptic fluorescence images (MagFura-2 pseudocolor code,yellow; OGB-5N pseudocolor code, blue). Scale bar, 10 μm. b, Two presynaptic APs (Vpre
) elicited by afferent stimulation evoked a large NMDAR-mediated postsynaptic current at the synapse shown in a at a holding potential of −80 mV in Mg2+-free extracellular solution. The estimated Ca2+ current through NMDAR channels is shown below, assuming a Pf of 11.6% (ICa,NMDA
). AMPARs were blocked by 10 μm NBQX. The average prestimulus fluorescence image was subtracted to obtain difference images (ΔF,right images), which represent the postsynaptic fluorescence changes of OGB-5N. ΔF images are shown at ∼50, 150, and 350 msec after afferent stimulation, at times when the total accumulated Ca2+ charge was 24, 44, and 55 pC, respectively. White corresponds to the largest fluorescence change. c, In the same MNTB neuron as shown in a and b, voltage steps from −80 to −10 mV (Vpost
) evoked a large inward Ca2+ current (ICa,post
) in the presence of TTX and TEA to block Na+ and K+ currents. On the right, OGB-5N difference images (ΔF) after subtraction of the prestimulus image are shown at ∼50, 150, and 350 msec after stimulation. Total accumulated Ca2+ charge was ∼40 pC.