Sleep is essential for optimal health and performance. Prolonged waking beyond its natural duration leads to a homeostatic sleep response based on the duration of prior wakefulness. Increases in homeostatic sleep response are associated with increased sleepiness and decreased alertness. Thus, a clear understanding of the mechanisms regulating the homeostatic sleep response is important in designing targeted treatments for sleepiness and associated neurobehavioral deficits experienced by military personnel and for the treatment of sleep disorders in veterans. Previous work from our laboratory demonstrated the importance of extracellular adenosine ([AD]ex) in sleep homeostasis within the basal forebrain (BF) wakefulness center. While there is ample evidence demonstrating a sleep deprivation (SD)-induced [AD]ex increase in BF, the mechanisms for this regional and localized increase is yet unknown. The purine nucleotide adenosine triphosphate (ATP) is released into extracellular space as a co-transmitter and via gliotransmission during neuronal activity in wakefulness. The neuromodulatory effects of [ATP]ex is exerted either by its direct action on P2 receptors or after its rapid breakdown by localized ectonucleotidase to [AD]ex, which acts via P1 receptors. Recent evidence suggests a dense presence of a neuronal ectonucleotidase in BF but not in cortex. Using state-of-the-art, multidisciplinary methods we will test the overarching hypothesis that an increase in [ATP]ex in the BF mediates sleepiness via its localized catabolism by ectonucleotidase to AD, which then inhibits wake promoting BF neurons.
In specific aim 1 we will test the hypothesis that during SD [ATP]ex increases in BF and frontal cortex, and produce an elevation of the homeostatic sleep response as determined by an increase in the delta activity (1-4.5 Hz) during recovery NREM sleep in rats. We will examine the time course of SD-induced changes in [ATP]ex and determine the relative effect of two mechanisms of [ATP]ex on the homeostatic sleep response: direct action on P2 receptor versus rapid degradation of [ATP]ex to [AD]ex by selective actions of ectonucleotidase with the prediction that ectonucleotidase inhibitors, but not P2 antagonists, will attenuate homeostatic sleep response.
In specific aim 2, will use a mouse model in which astrocytic release of [ATP]ex is prevented. The transgene in this mice (astrocyte-selective dominant negative SNARE (dnSNARE))is conditionally regulated with dietary doxycycline. , We will test the hypothesis that SD increases gliotransmission of [ATP]ex in BF. We predict allowing dnSNARE expression (-doxycycline) will prevent [ATP]ex release and the increase in [AD]ex during SD, whereas suppressing dnSNARE expression (+doxycycline) will show increases in [AD]ex and a homeostatic response, due to gliotrasmission release of [ATP]ex->[AD]ex.
In specific aim 3, using GAD67-GFP mice, we will test the hypothesis that [ATP]ex will cause an inhibition of cortically projecting BF neurons in vitro, due to its breakdown to AD and activation of A1 receptor. We will also determine the time course of ATP breakdown to [AD]ex in the BF.
In specific aim 4, we will test the hypothesis that ATP-derived AD's action on the A1 receptor mediates increased sleepiness and consequent neurobehavioral performance decrements following SD. We will use two novel behavioral tests (i) Our rodent version of the human multiple sleep latencies test that provides a direct measure of sleepiness, and (ii) our rodent version of the human psychomotor vigilance test to measure sustained attention (vigilance) following 3h and 6h of SD during the light period. Receptor specificity will be tested by reverse microdialysis during SD of antagonists of the P2 and, separately, to the A1 receptor. We predict that A1 receptor antagonists will decrease sleepiness (sleep latencies) and vigilance whereas the P2 antagonists will be much weaker effect, indicating a predominantly adenosinergic mediation of sleepiness. The successful completion of this comprehensive investigation will shed light on purinergic mechanisms involved in homeostatic sleep controls.

Public Health Relevance

Sleep is universal and a highly regulated behavior in mammals. While increasing evidence indicate that adequate sleep is essential for key brain functions, sleep loss is, unfortunately, frequent among military personnel as well as general public. Sleep loss triggers selective changes in the molecular pathways leading to excessive sleepiness, impaired attention and cognition. Studies proposed in this application are directed towards investigating the mechanism excessive sleepiness that follows sleep deprivation. A clear understanding of the molecular and cellular processes triggered by sleep deprivation is important in designing targeted treatments for sleepiness and associated neurobehavioral deficits experienced by military personnel and for the treatment of sleep disorders in veterans.

National Institute of Health (NIH)
Veterans Affairs (VA)
Non-HHS Research Projects (I01)
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Neurobiology C (NURC)
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VA Boston Health Care System
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Yang, Chun; Larin, Andrei; McKenna, James T et al. (2018) Activation of basal forebrain purinergic P2 receptors promotes wakefulness in mice. Sci Rep 8:10730
Yang, Chun; McKenna, James T; Brown, Ritchie E (2017) Intrinsic membrane properties and cholinergic modulation of mouse basal forebrain glutamatergic neurons in vitro. Neuroscience 352:249-261
Dworak, Markus; Kim, Tae; Mccarley, Robert W et al. (2017) Creatine supplementation reduces sleep need and homeostatic sleep pressure in rats. J Sleep Res 26:377-385
Zielinski, Mark R; Gerashchenko, Dmitry; Karpova, Svetlana A et al. (2017) The NLRP3 inflammasome modulates sleep and NREM sleep delta power induced by spontaneous wakefulness, sleep deprivation and lipopolysaccharide. Brain Behav Immun 62:137-150
Gvilia, Irma; Suntsova, Natalia; Kostin, Andrey et al. (2017) The role of adenosine in the maturation of sleep homeostasis in rats. J Neurophysiol 117:327-335
Zant, Janneke C; Kim, Tae; Prokai, Laszlo et al. (2016) Cholinergic Neurons in the Basal Forebrain Promote Wakefulness by Actions on Neighboring Non-Cholinergic Neurons: An Opto-Dialysis Study. J Neurosci 36:2057-67
Shukla, Charu; Basheer, Radhika (2016) Metabolic signals in sleep regulation: recent insights. Nat Sci Sleep 8:9-20
Kalinchuk, Anna V; Porkka-Heiskanen, Tarja; McCarley, Robert W et al. (2015) Cholinergic neurons of the basal forebrain mediate biochemical and electrophysiological mechanisms underlying sleep homeostasis. Eur J Neurosci 41:182-95
Kim, T; Ramesh, V; Dworak, M et al. (2015) Disrupted sleep-wake regulation in type 1 equilibrative nucleoside transporter knockout mice. Neuroscience 303:211-9
Kim, Tae; Thankachan, Stephen; McKenna, James T et al. (2015) Cortically projecting basal forebrain parvalbumin neurons regulate cortical gamma band oscillations. Proc Natl Acad Sci U S A 112:3535-40

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