Hypocretin and GABA Interact in the Pontine Reticular Formation to Increase Wakefulness

Abstract

underlies the pathophysiology of narcolepsy, 4,5 and disruption of hypocretin signaling in mouse, 6 rat, 7,8 and dog 9 leads to narcolepsy-cataplexy. Hypocretinergic neurons project to multiple areas of the brain, including those important for regulating sleep and wakefulness. 1 One such area is the pontine reticular nucleus, oral part (PnO). 1,10 The PnO is the rostral portion of the rodent pontine reticular formation 11 and contributes to the generation of wakefulness and rapid eye movement (REM) sleep. 12 Microinjection of hypocretin-1 into rat PnO causes an increase in wakefulness, 13 and microinjection of hypocretin-1 into cat pontine reticular formation increases the cortically activated states of REM sleep 14 or wakefulness. 15,16 Administering hypocretin-1 into the PnO may increase wakefulness by modulating the release of arousal-promoting neurotransmitters within the PnO. Direct administration of hypocretin-1 to the PnO of isoflurane-anesthetized rat causes a concentration-dependent increase in both acetylcholine (ACh) release 17 and GABA levels 13 within the PnO. Extracellular recording studies of PnO neurons in urethane-anesthetized rat show that iontophoretic application of hypocretin-1 causes a hyperpolarization that is blocked by prior application of bicuculline. 10 This finding indicates that the hypocretin-1-induced inhibition of PnO neurons is mediated by GABA A receptors. Identified GABAergic neurons in brainstem slices of mouse PnO have been shown to be excited by hypocretin-1, 18 and intracellular recording studies in halothane-anesthetized cat show that hypocretin-1 can also cause direct depolarization of PnO neurons and an increase in PnO neuronal firing rate. 14 Numerous studies have demonstrated that GABAergic transmission in the PnO increases wakefulness and inhibits REM sleep. 13,[19][20][21][22][23][24][25] The present study provides the first test of the hypothesis that the wakefulness-promoting effects of delivering hypocretin-1 into the PnO are mediated by GABA A receptors as well as by hypocretin receptors. This hypothesis was evaluated by determining whether (1) microinjection of hypocretin-1 into the PnO causes a concentration-dependent increase in wakefulness, (2) this increase in wakefulness is blocked by coadministration of the hypocretin receptor-1 (hcrt-r1) antagonist SB-334867, and (3) coadministration of the GABA A receptor antagonist bicuculline also blocks the wakefulness response to hypocretin-1. Portions of these data have been presented as abstracts. 26,27 MATERIALS AND METHODS Chemicals and Drug SolutionsHuman hypocretin-1 was purchased from California Peptide Research, Inc. (Napa, CA). Bicuculline methiodide was pur- Study Objectives: Hypocretin-1/orexin A administered directly into the oral part of rat pontine reticular formation (PnO) causes an increase in wakefulness and extracellular g-aminobutyric acid (GABA) levels. The receptors in the PnO that mediate these effects have not been identified. Therefore, this study tested the hypothesis that the increase in wa...

Endogenous GABA Levels in the Pontine Reticular Formation Are Greater during Wakefulness than during Rapid Eye Movement Sleep

“…In contrast, wakefulness is decreased and REM sleep is increased by pontine microinjection of GABA receptor antagonists or drugs that decrease GABA levels (Camacho-Arroyo et al, 1991; Xi et al, 1999, 2001; Sanford et al, 2003; Marks et al, 2008; Watson et al, 2008; Flint et al, 2010). Additionally, the increase in wakefulness caused by microinjecting hypocretin-1 into rat pontine reticular formation is blocked by coadministering the GABA A receptor antagonist bicuculline (Brevig et al, 2010). This finding supports the conclusion that in the pontine reticular formation, hypocretin-1 causes an increase in wakefulness by increasing GABAergic transmission (Watson et al, 2008).…”

Hypocretin and Its Emerging Role as a Target for Treatment of Sleep Disorders

“…In vivo slice electrophysiology studies have shown that hypocretin increases firing rates of monoaminergic neurons in the LC, serotonergic neurons in the DR, histaminergic neurons in the TMN, and cholinergic neurons in the BF and LDT [5••], but have no effect on the GABAergic neurons in the VLPO. First reported by Brevig et al [24], injection of hypocretin-1 in a rat pontine reticular nucleus, a region involved in wakefulness and REM sleep, increases wakefulness that is mediated by GABAergic transmission of GABA A receptors. These findings indicate that activities of wake-promoting regions are without a doubt regulated by hypocretin.…”

Modulation of neurotransmitter release in orexin/hypocretin‐2 receptor knockout mice: A microdialysis study

“…With the assumption that OX2 receptors are coupled exclusively to G q proteins, a disinhibitory action through their presynaptic regulation of inhibitory GABAergic neurons might be a probable mechanism (Kukkonen et al, 2002). Studies have also suggested an excitatory role of OX2 receptors in modulating activity of GABAergic neurons in the medial septum/diagonal band of Broca (Wu et al, 2002) and arcuate nucleus (Burdakov et al, 2003), indicating that, at least in other brain areas, activation of OX2 receptors could result in an increase of GABA release and a subsequent inhibitory action on other neurotransmitters in the area via GABA A receptors (Nuñez et al, 2006;Brevig et al, 2010). An alternative mechanism could involve a direct action through OX2 receptors coupled to G i/o proteins and localized on cholinergic, histaminergic, and norepinephrinergic nerve terminals.…”