Fig. 2.

Activation of extrasynaptic NMDARs induces large amplitude Ca²⁺ elevations. (A) Single-cell Ca²⁺ transients (fura-2FF) in young neurons triggered by PTX (100 μM) activation of synaptic NMDARs, followed by MK-801 (10 μM) to block open (i.e., synaptic) NMDARs. With this low-affinity Ca²⁺ probe, submicromolar synaptically induced somatic transients are barely visible in the 0- to 3-min portion of the trace. Application of NMDA after washout activates only previously unactivated extrasynaptic NMDARs, leading to large Ca²⁺ elevations. (Inset) Experiment parallel to the first 7 min of the main trace but using high-affinity fura-2 shows that low-amplitude synaptically induced Ca²⁺ elevations (average of 9 cells) are blocked by MK-801; they are also abolished by TTX pretreatment (Fig. S3B). Synaptic Ca²⁺ spikes evoked by bicuculline are comparable to those for PTX (Fig. S3A). (B) Death rate after bath NMDA exposure (100 μM, 30 min; global) was not different from that evoked by activation of only extrasynaptic NMDARs (extrasynaptic). (C) Extrasynaptic Ca²⁺ elevations are mainly eliminated in the presence of Co 101244. Average traces are shown in bold.