LATERAL SEPTAL CHOLINERGIC NEURONS REPRESENT AVERSIVE STIMULI AND MEDIATE AVOIDANCE BEHAVIOR

Cholinergic neurons are key modulators of diverse cognitive processes. While major cholinergic populations in the basal forebrain and brainstem have been extensively characterized, surprisingly little is known about a significant population of cholinergic neurons located in the lateral septum (LS). The aim of this study was to characterize the anatomical hallmarks and cognitive role of LS cholinergic neurons (LSCNs). A Cre-dependent viral tracer was injected into the LS of ChAT-Cre mice and their long-range projections were analyzed. The average number of LSCNs was quantified by stereological methods. To assess their causal role in behavior, channelrhodopsin-2 (ChR2) was expressed in LSCNs and optogenetic stimulation was paired with one chamber in a place preference task. We examined neuronal activity using fiber photometry and electrophysiology during a probabilistic Pavlovian conditioning task, in which one auditory cue predicted reward and another cue predicted punishment. We counted approximately 15,000 cholinergic neurons in the LS, characterized by small somata and densely spined dendrites. These neurons primarily projects to the anterior hypothalamic nucleus, ventromedial hypothalamus and anterodorsal preoptic nucleus, where they release acetylcholine. ChR2-expressing mice avoided the stimulation-coupled chamber to a significantly greater extent than control mice. During Pavlovian conditioning, LSCNs showed minimal or no cue-evoked responses and did not discriminate between reward- and punishment-predicting cues. However, these neurons exhibited robust activation during punishment outcomes. Moreover, their activity encoded punishment prediction error signals. Our findings show that LSCNs are projection neurons that innervate the hypothalamus and the amygdala and may play an important role in aversive conditioning.