Corticotropin-releasing factor (CRF), the primary neuropeptide regulator of responses to stress, is implicated in the
suppression of sleep during stress conditions. We hypothesize that stress-induced disruption of sleep involves
suppressing effect of increased CRF signaling in the brain on the activity of key sleep-regulatory neuronal systems. In
Aim 1, we will determine the effects of pharmacological suppression of CRF signaling, during acute stress, on wakesleep behavior and on sleep-related activity in the preoptic hypothalamus. Rats will be subjected to
intracerebroventricular (ICV) administration of antalarmin (ANT, CRF-Receptor1 antagonist) followed by exposure to
acute species-specific psychological stressor - cage exchange paradigm (see Ref. 21), and recorded for post-stress sleep.
In 5.5 hours after stress exposure (Ref. 21), rats will be subjected to cardiac perfusion. Brain tissue will be processed for
double immunostaining for c-Fos protein and glutamic acid decarboxylase (GAD). In Aim 2, we will determine the
effect of chemogenetic silencing of CRF neurons on stress-induced disturbances of sleep. Separate groups of CRF-irescre mice will be injected AAV-hSyn-DIO-hM4Di-mCherry into the paraventricular nucleus of the hypothalamus
(PVN), central nucleus of the amygdala (CeA) and bed nucleus of the stria terminalis (BNST). In four weeks after the
injection (allowed for gene recombination and expression), mice will be injected clozapine-n-oxide (CNO), placed into
dirty cages from male rats (see Pilot Data) and recorded for sleep. We predict that ANT-treated versus control rats will
exhibit higher amounts and more consolidated sleep, and higher numbers of Fos+GABAerg neurons in the preoptic
hypothalamus. Chemogenetic silencing of CRF neurons will attenuate stress-induced sleep disturbances. Overall, new
knowledge generated after the completion of the proposed studies will (1) contribute to better understanding of
fundamental mechanisms underlying stress-induced disturbances of sleep, and (2) constitute the scientific background
for developing novel strategies to prevent/treat stress-induced abnormalities in sleep and associated health problems.