Figure 1.

Inhibition of NOS prevents NMDA-induced ΔΨ_m dissipation. A, Simultaneous measures over time of NMDA-induced changes in [Ca²⁺]_i (fura-2) and ΔΨ_m (TMRM) in the absence (left) and presence (right) of l-NAME (100 μm for 3 h). Maximal ΔΨ_m dissipation is induced with FCCP (dotted line) at the end of each experiment. To eliminate contributions to changes in mitochondrial TMRM intensity made by plasma membrane depolarization, TMRM intensities from a mitochondria-rich region of each neuron at each point in time was divided by the corresponding intensity from a mitochondria-poor region within the same neuron. Each ratio was normalized to the mean baseline ratio (100%) and the mean ratio obtained after FCCP (0%). NMDA induces a modest ΔΨ_m dissipation that is blocked by l-NAME. B, Population summary of mean peak [Ca²⁺]_i increases during NMDA in the presence and absence of NOS blockade. [Ca²⁺]_i increases are reported with fura-FF (K_d of 5.5 μm). NMDA-induced [Ca²⁺]_i increases do not significantly differ between control and l-NAME. Inset, Representative examples of raw fura-FF responses to NMDA in control and l-NAME-treated neurons. Calibration bar, 1 min. C, NMDA-induced ΔΨ_m dissipation is decreased during NOS blockade (l-NAME, 7-NI), and the blockade is overcome with supplemental l-arginine (Arg). MnTBAP, a superoxide dismutase mimetic, increases NMDA-induced ΔΨ_m dissipation, suggesting that a portion of the NO generated by NMDA is converted to peroxynitrite. Statistical comparisons made between control and each treatment. ^*p < 0.05; ^†p < 0.01. A.U., Arbitrary units.