Cholinergic Modulation of Narcoleptic Attacks in Double Orexin Receptor Knockout Mice

Abstract

To investigate how cholinergic systems regulate aspects of the sleep disorder narcolepsy, we video-monitored mice lacking both orexin (hypocretin) receptors (double knockout; DKO mice) while pharmacologically altering cholinergic transmission. Spontaneous behavioral arrests in DKO mice were highly similar to those reported in orexin-deficient mice and were never observed in wild-type (WT) mice. A survival analysis revealed that arrest lifetimes were exponentially distributed indicating that random, Markovian processes determine arrest lifetime. Low doses (0.01, 0.03 mg/kg, IP), but not a high dose (0.08 mg/kg, IP) of the cholinesterase inhibitor physostigmine increased the number of arrests but did not alter arrest lifetimes. The muscarinic antagonist atropine (0.5 mg/kg, IP) decreased the number of arrests, also without altering arrest lifetimes. To determine if muscarinic transmission in pontine areas linked to REM sleep control also influences behavioral arrests, we microinjected neostigmine (50 nl, 62.5 µM) or neostigmine + atropine (62.5 µM and 111 µM respectively) into the nucleus pontis oralis and caudalis. Neostigmine increased the number of arrests in DKO mice without altering arrest lifetimes but did not provoke arrests in WT mice. Co-injection of atropine abolished this effect. Collectively, our findings establish that behavioral arrests in DKO mice are similar to those in orexin deficient mice and that arrests have exponentially distributed lifetimes. We also show, for the first time in a rodent narcolepsy model, that cholinergic systems can regulate arrest dynamics. Since perturbations of muscarinic transmission altered arrest frequency but not lifetime, our findings suggest cholinergic systems influence arrest initiation without influencing circuits that determine arrest duration.

Pharmacogenetic inhibition of the subcoeruleus region influences rem sleep and cataplexy in narcoleptic mice

“…These findings are supported by a large body of evidence which pointing to acetylcholine as an important neurotransmitter system for cataplexy. In narcoleptic dogs increasing cholinergic drive systemically and focally into pontine regions regulating REM sleep atonia increase cataplexy; these findings have been recently reproduced and built upon in narcoleptic mice [36][37][38]. Our findings compliment these prior studies.…”

“…Pharmacological administration of cholinergic agonists have been shown to increase cataplexy in narcoleptic dogs and mice [36,37]. Histological findings indicate increased cholinergic activity in canine and murine models of narcolepsy.…”

International Union of Basic and Clinical Pharmacology. LXXXVI. Orexin Receptor Function, Nomenclature and Pharmacology

“…Constitutive gene knockouts and Ox/ATX transgenics more closely mimic narcolepsy and varying aspects of the disorder than what might be expected from pharmacological manipulation. For example, behavioral arrests have been observed in Hcrt and OX 2 receptor knockouts (Chemelli et al, 1999;Willie et al, 2003) and in mice in which both orexin receptors have been mutated (Kalogiannis et al, 2011). Although it is currently unclear whether the behavioral episodes observed in the later model constitute cataplexy , cataplectic episodes have not been observed in response to even high doses of dual orexin receptor antagonists across multiple species .…”

Orexin/Hypocretin Antagonists in Insomnia: From Bench to Clinic

“…As orexin knockout mice have relatively normal baseline ventilation (Kalogiannis et al 2011), it seems unlikely that orexin antagonists would significantly reduce respiratory drive. However, they may reduce the response to hypercarbia.…”

“…This has been a b (Sakurai 2007), OX2R KO mice manifest some features of narcolepsy, including an inability to sustain wakefulness (Willie et al 2003). DKO mice display the most profoundly disturbed sleep phenotype of all three models: narcolepsy with cataplexy (transient episodes of behavioral arrest) (Kalogiannis et al 2011). The robust narcoleptic phenotype in DKO mice indicates a synergistic role between OX1R and OX2R in the maintenance of wakefulness.…”