Narcolepsy is a disorder of unstable wake and sleep states caused by the lack of orexin neurons which degenerate most likely as a consequence of an autoimmune process. The state instability of narcolepsy includes rapid eye movement (REM) sleep intruding into wake in the form of dream-like hallucinations and cataplexy, muscle paralysis (atonia) much like occurs in REM sleep. In mice lacking orexin peptides, cataplexy is also observed with similar presentation as in humans of muscle paralysis during wakefulness which is often triggered by positive emotions. Prior research showed that the activation of the central amygdala is sufficient to promote cataplexy in a mouse model of narcolepsy. The central amygdala (CeA) contains a variety of neuronal types, and we hypothesize that γ-aminobutyric acid (GABA)-ergic neurons expressing the oxytocin receptor (OTR) mediate cataplexy as these neurons project to a known REM sleep atonia-regulating region, the ventrolateral periaqueductal gray (vlPAG)/lateral pontine tegmentum (LPT), and, as oxytocin (OT) sensitive neurons in the amygdala, likely participate in emotional processing and social behavior. In this study, we used fiber photometry to investigate the behavior of these neurons in response to social and rewarding stimuli, during emotion-triggered cataplexy, and across arousal states in an effort to define their potential role in emotion-triggered cataplexy. Initial recordings were conducted at too low an excitation light power to stimulate the green fluorescent calcium indicator, GCaMP6s, but were useful in optimizing MATLAB analysis and behavioral tests later done at higher LED power. The second series of recordings with higher excitation light power and better signal to noise ratio, showed increased activity in response to social interaction and reward, prior to REM transitions, and decreased activity during cataplexy confirming patterns seen in initial recordings. In recordings with higher excitation light, these responses appear to occur before interaction with stimulus mice or reward stimulus. In the future, additional recordings with a higher signal to noise ratio will be needed to confirm these results. In conclusion, responses of CeA-OTR neurons to social and rewarding stimuli, cataplexy, and at REM transitions are in support of a possible role of these neurons in emotion-triggered cataplexy which can be tested using additional methods, such as optogenetics.

The ability of animals to sense, interpret, and respond appropriately to social stimuli in their environment is essential for identifying and distinguishing between members of their own species. In mammals, social interactions both within and across species play a key role in determining if an animal will live to pass on its genes to the next generation or else be removed from the gene pool. The result of this selection pressure can be observed in specialized neural circuits that respond to social stimuli and orchestrate appropriate behavioral responses. This highly conserved network of brain structures is often referred to as the Social Behavior Network (SBN). The medial amygdala (MeA) is a central node in the SBN and has been shown to be involved in transforming information from olfactory sensory systems into social and defensive behavioral responses. Previous research has shown that individual neurons in the MeA of anesthetized mice respond selectively to different chemosensory social cues, a characteristic not observed in its upstream relay, the accessory olfactory bulb (AOB). However, the cause of this stimulus selectivity in the MeA is not yet understood. Here, I hypothesize that a subpopulation of neurons in the MeA that express the enzyme aromatase are involved in the sensory representation of social stimuli in awake, behaving animals. To test this hypothesis, I designed and built a novel behavioral apparatus that allows for discrete presentations of social stimuli in a highly controllable and reproducible environment. I then injected the adeno-associated virus (AAV) AAV-Syn-Flex-GCAMP6s into the MeA of Aromatase:Cre transgenic mice and implanted a fiber optic cannula slightly above the injection site. The combination of this transgenic mouse line and conditional AAV caused GCaMP6s expression to be exclusive to aromatase-expressing neurons. By coupling my novel behavioral apparatus to a fiber photometry system, I successfully recorded the moment-to-moment activity of aromatase neurons in the MeA of awake, behaving animals as they investigated various social stimuli. Aromatase neurons in the MeA of adult male mice respond strongly to conspecific social stimuli, including live adult mice, mouse pups, and mouse urine samples. Sniffing and investigative behaviors correlated strongly with increased GCaMP6s signal in aromatase neurons, reflecting increases in their neural activity. Interestingly, after repeated investigations of the same stimuli the activity of aromatase neurons gradually diminished. Presenting a novel stimulus following repeated investigations of a familiar stimulus reinstated some, but not all of the initial GCaMP6s signal. This points to the potential role that aromatase neurons may play in the habituation to social stimuli that are consistently present in their environment. Investigations of predator stimuli did not evoke significant responses from aromatase neurons, nor did investigations of non-social stimuli. These results demonstrate that aromatase expressing neurons in the MeA of awake, behaving animals encode the sensory representation of conspecific social stimuli, and their responses are highly selective to the type of stimulus presented.