Activity-dependent nitric oxide concentration dynamics in the laterodorsal tegmental nucleus in vitro

The behavioral-state related firing of mesopontine cholinergic neurons of the laterodorsal tegmental nucleus appears pivotal for generating both arousal and rapid-eye-movement sleep. Since these neurons express high levels of nitric oxide synthase, we investigated whether their firing increases local extracellular nitric oxide levels. We measured nitric oxide in the laterodorsal tegmental nucleus with a selective electrochemical microprobe (35 microm diam) in brain slices. Local electrical stimulation at 10 or 100 Hz produced electrochemical responses that were attributable to nitric oxide. Stimulus trains (100 Hz; 1 s) produced biphasic increases in nitric oxide that reached a mean peak concentration of 33 +/- 2 (SE) nM at 4.8 +/- 0.4 s after train onset and decayed to a plateau concentration of 8 +/- 1 nM that lasted an average of 157 +/- 23.4 s (n = 14). These responses were inhibited by N(G)-nitro-L-arginine-methyl-ester (1 mM; 92% reduction of peak; n = 3) and depended on extracellular Ca(2+). Chemically reduced hemoglobin attenuated both the electrically evoked responses and those produced by authentic nitric oxide. Application of the precursor, L-arginine (5 mM) augmented the duration of the electrically evoked response, while tetrodotoxin (1 microM) abolished it. Analysis of the stimulus-evoked field potentials indicated that electrically evoked nitric oxide production resulted from a direct, rather than synaptic, activation of laterodorsal tegmental neurons because neither nitric oxide production nor the field potentials were blocked by ionotropic glutamate receptor inhibitors. Nevertheless, application of N-methyl-D-aspartate also increased local nitric oxide concentration by 39 +/- 14 nM (n = 8). Collectively, these data demonstrate that laterodorsal tegmental neuron activity elevates extracellular nitric oxide concentration probably via somatodendritic nitric oxide production. These data support the hypothesis that nitric oxide can function as a local paracrine signal during the states of arousal and rapid-eye-movement sleep when the firing of mesopontine cholinergic neurons are highest.