NO reduces evoked release and frequency of spontaneous release in a cGMP-independent manner.

<p>(A) NO suppresses evoked release (eEJC) over a time course of 55 min. Insets show representative single eEJCs at 40 min for both conditions. (B) Mean eEJC amplitudes (left axis) and QC (right axis) of <i>w</i>^<i>1118</i> NMJs are reduced following NO exposure (at 40 min). The sGC inhibitor ODQ (50 μM) did not affect the response to NO. (C) Mean eEJC amplitudes (left axis, black) and QC (right axis, grey) of NOS^C and NOS^Δ15 NMJs. (D) Raw mEJC recordings of <i>w</i>^<i>1118</i> NMJs and mEJC parameters (top to bottom: amplitude, frequency, decay). Top insets show representative mEJC recordings. Bottom insets show single mEJCs (grey) and averaged mEJC (red) with single exponential fit to the decay. (E) Raw mEJC recordings for both NOS^C and NOS^Δ15 genotypes. Below, mEJC quantal parameters: amplitude and frequency, Student <i>t</i> test each relative to <i>w</i>^<i>1118</i> Ctrl, *<i>p</i> = 0.04, ***<i>p</i> = 0.001. (F) cGMP content of larval brains under the conditions indicated (NO: 40 min NO exposure, NO + ODQ: 40 min NO exposure in the presence of 50 μM ODQ, NO + Zap: 40 min NO exposure + PDE inhibitor Zap, 20 μM). (G) FlincG3 fluorescence images of a Ctrl and stimulated NMJ (20 Hz for 10 s, duty cycle: 1 min for total of 10 min). (H) Summary of FlincG3 fluorescence (in a.u.’s). The raw data can be found in <a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.2003611#pbio.2003611.s011" target="_blank">S1 Data</a>. Data denote mean ± SEM in all graphs, ANOVA with post hoc Tukey-Kramer, *<i>p</i> < 0.05, **<i>p</i> < 0.01, ****<i>p</i> < 0.0001. a.u., arbitrary unit; cGMP, cyclic guanosine monophosphate; Ctrl, control; eEJC, evoked EJC; EJC, excitatory junction current; mEJC, miniature EJC; NMJ, neuromuscular junction; NO, nitric oxide; ODQ, 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one; PDE, phosphodiesterase; QC, quantal content; sGC, soluble guanylyl cyclase; Zap, zaprinast.