Abstract

Anxiety disorders, such as posttraumatic stress disorder (PTSD), are characterized by a high prevalence and debilitating symptoms. However, the current first-line treatment for these conditions, which consists of selective serotonin reuptake inhibitors (SSRIs) and cognitive behavioral therapy, alongside symptomatic treatment with benzodiazepines, does not represent by far a functional solution for all affected patients. Therefore, identifying and characterizing novel candidates for alternative anxiolytic therapies are a crucial focus of psychiatric and neurobiological research. This study focuses on Neuropeptide S (NPS), a newly identified endogenous neuropeptide that has been shown to exert strong anxiolytic effects upon intracerebral injection in rodents. In an approach that combines basic research with incipient clinically relevant application, novel mechanisms and brain targets of NPS-mediated anxiolytic effects were identified, and a noninvasive application procedure also applicable in patients, namely the intranasal administration, was established for the first time for NPS in mouse models. In a first step, the feasibility of intranasal NPS delivery was established in mice using fluorophore-coupled NPS to allow intracerebral tracking. This method permitted for the first time tracking of intranasally applied substances within the brain at a single-cell resolution. These results not only proved the applicability of intranasal NPS administration in the mouse, but also allowed identification and characterization of hitherto undescribed cerebral NPS target cells, which were shown to be most likely exclusively neurons. Moreover, specific uptake of fluorescently labeled NPS in the hippocampus provided the first direct evidence linking this brain region, a well-known major player in the regulation of fear expression, to the NPS circuitry. Further investigation into the functional role of the hippocampus in NPS-elicited anxiolytic effects revealed that local microinjections of NPS into the ventral CA1 (vCA1) region are sufficient to elicit anxiolysis in C57BL6/N mice on the elevated plus maze (EPM). In a second step, behavioral and molecular effects of intranasal NPS treatment were characterized in C57BL/6N mice. Intranasal application of NPS was shown here to produce anxiolytic effects similar to those described by others after intracerebral injection. This finding represents the basis for the implementation of a future NPS-based therapy via nasal sprays in patients suffering from anxiety disorders. Furthermore, the molecular effects of NPS treatment on cerebral protein expression were examined here for the first time. This research led to identification of novel downstream targets of NPS-mediated regulation in the hippocampus and the prefrontal cortex. These new targets include proteins involved in the glutamatergic system and in synaptic plasticity, both of which are known to be dysregulated in anxiety disorders. Finally, the effects of intranasal NPS treatment, hitherto described only in non-pathological animal models, were examined for the first time in mouse models of anxiety disorders, namely the high anxiety behavior (HAB) mice and a mouse model of PTSD. In HAB mice, NPS treatment elicited anxiolytic effects similar to those observed in C57BL/6N mice. In the mouse model of PTSD, NPS counteracted disease-related changes in expression levels of hippocampal synaptic proteins. To sum up, this work expands the current state-of-knowledge concerning the molecular and mechanistic background of NPS-mediated anxiolysis by characterizing the role of the hippocampus in the NPS circuitry and by identifying novel downstream targets of NPS. The data on anxiolytic effects of intranasal NPS treatment especially in mouse models of anxiety disorders furthermore establishes the therapeutic potential of NPS as a novel anxiolytic treatment.