Orexin Receptor Blockade-Induced Sleep Preserves the Ability to Wake in the Presence of Threat in Mice

Abstract

Retention of the ability to wake from sleep in response to dangerous situations is an ideal characteristic of safe hypnotics. We studied the effects of a dual orexin receptor antagonist-22 (DORA-22) and the GABA-A receptor modulator, triazolam, on the ability to wake in response to aversive stimuli. We examined four modalities of sensory inputs, namely, auditory (ultrasonic sound), vestibular (trembling), olfactory (predator odor), and autonomic (hypoxia) stimuli. When the mice fell asleep, one of the four stimuli was applied for 30 s. In the case of auditory stimulation, latency to arousal following vehicle, DORA-22, and triazolam administration was 3.0 (2.0-3.8), 3.5 (2.0-6.5), and 161 (117-267) s (median and 25-75 percentile in the parentheses, n = 8), respectively. Latency to return to sleep after arousal was 148 (95-183), 70 (43-98), and 60 (52-69) s, respectively. Similar results were obtained for vestibular and olfactory stimulation. During the hypoxic stimulation, latencies for arousal and returning to sleep were not significantly different among the groups. The findings of this study are consistent with the distinct mechanisms of these sleep promoting therapies; GABA-A receptor activation by triazolam is thought to induce widespread central nervous system (CNS) suppression while DORA-22 more specifically targets sleep/wake pathways through orexin receptor antagonism. These data support the notion that DORA-22 preserves the ability to wake in response to aversive and consciousness-inducing sensory stimuli, regardless of modality, while remaining effective in the absence of threat. This study provides a unique and important safety evaluation of the potential for certain hypnotics.

Keywords: aversive stimuli; dual orexin receptor antagonist; hypnotics; hypocretin; orexin; triazolam.

Figure 1

Effects of dual orexin receptor antagonist-22 (DORA-22), triazolam, and eszopiclone on sleep time and sleep architecture. Time slept per every 30 min is shown for (A) DORA-22, (B) triazolam, and (C) eszopiclone for 6 h after p.o. administration of the drugs. The same values for the vehicle are shown repeatedly for comparison purposes. Each animal received DORA, triazolam, eszopiclone, and vehicle in a randomized order on spaced days. Data are shown as Mean ± SEM. (D) Comparison among the four treatments (F_(3,12) = 5.101, p = 0.017) revealed that DORA-22 and triazolam, but not eszopiclone, significantly increased total sleep time during a 3-h period starting at 1 h after administration. These drugs did not affect sleep episode duration (F_(3,12) = 0.906, p = 0.467; E) but did decrease awake episode duration (F_(3,12) = 33.0, p < 0.001; F). In (D–F), data from the same animal are connected with lines to show possible interactions between drugs in individual mice. Horizontal lines indicate mean value for each treatment. Statistical results using repeated measure ANOVA followed by Tukey’s multiple comparisons test are indicated in the graph.

Figure 2

Latency to arousal and latency to return to sleep induced by four modalities of sensory stimulation. (A) Auditory stimulus by ultrasonic sound (n = 8). (B) Vestibular stimulus by cage shaking (n = 8). (C) Olfactory stimulus by a predator odor (n = 10). (D) Autonomic stimulus by hypoxia (n = 8). Each animal received vehicle (V), DORA-22 (D), and triazolam (T) in a randomized order on spaced days and were tested for one of the stimuli 2–3 times. The value from those tests were averaged and are represented as a single dot. Data from the same animal are connected with lines to show possible drug interactions within an individual animal. Horizontal lines indicate, from top to bottom, 75 percentile, median, and 25 percentile, respectively. Statistical results using the Friedman test, a repeated measure nonparametric multiple comparisons test, are indicated in the graph. Note that latency to arousal in DORA-22 treated mice is not different from those in vehicle treated mice and significantly shorter than those in triazolam treated mice, with the exception of hypoxia. Also note that latency to return to sleep in DORA-22 treated mice is significantly shorter than those in vehicle treated mice and not different from those in triazolam treated mice at least for auditory and vestibular stimuli.