Parabrachial-insular stimulation does not wake mice.

The parabrachial nucleus (PB) in the upper brainstem receives interoceptive information and sends a massive output projection directly to the cerebral cortex. Its glutamatergic axons primarily target the midinsular cortex, and we have proposed that this PB-insular projection promotes arousal. Here, we test whether stimulating this projection causes wakefulness. We combined optogenetics and video-electroencephalography (vEEG) in mice to test this hypothesis by stimulating PB axons in the insular cortex. Stimulating this projection did not alter the cortical EEG or awaken mice. Also, despite a tendency toward aversion, PB-insular stimulation did not significantly alter real-time place preference (RTPP). These results are not consistent with the hypothesis that the direct PB-insular projection is part of the ascending arousal system. A brainstem region critical for wakefulness overlaps the medial parabrachial nucleus (PB) and has functional and direct axonal connectivity with the insular cortex. In this study, we hypothesized that this direct projection from the PB to the insular cortex promotes arousal. However, photostimulating PB axons in the insular cortex did not alter the cortical EEG or awaken mice. This information constrains the possible circuit connections through which brainstem neurons may sustain arousal.