Download Sleep/Neurology-The Orexin System

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Sleep/Neurology-The
Orexin System
Todd J. Swick, M.D. FAAN, FAASM
Assistant Clinical Professor of NeurologyUniversity of Texas Health Sciences CenterHouston
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Speaker Disclosures
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Grants/Research Support


Consultant

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Jazz Pharmaceuticals, UCB, Vanda Pharmaceuticals, XenoPort,
Aerial Pharmaceuticals
Jazz Pharmaceuticals, Vanda Pharmaceuticals, XenoPort, Aerial
Pharmaceuticals, Merck Pharmaceuticals
Honorarium

Jazz Pharmaceuticals, Vanda Pharmaceuticals, XenoPort, Merck
Pharmaceuticals
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Sleep/Neurology-The
Orexin/Hypocretin System
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Objectives
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Historical Overview of Sleep-Wake Signaling
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1998 Discovery of the Orexin/HCRT Neuropeptides
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Neurophysiologic Effects of Orexin/HCRT peptides
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Control of Sleep/Wake mechanisms
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Clinical aspects of orexin/HCRT activity and loss
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Possible role of Orexin/HCRT in the pathogenesis of
MCI/Alzheimer’s
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Possible answer as to “Why we need sleep”
von Economo C. Sleep as a problem of localization. J Nerv Ment Dis. 1930;71(3):249-259.
Original Hypocretin/Orexin Papers
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Sleep/Neurology-The Orexin System
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Two papers were published within 3 weeks of one another in
early 1998
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The San Diego/Stanford groups were looking for genes with
expressed selectivity in the control of appetite, thirst and
other autonomic and arousal functions
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Two peptides derived from a single gene were identified and
named Hypocretins for their hypothalamic location and
sequence homology to secretin
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Sleep/Neurology-The Orexin/HCRT
System
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Team from Dallas led by Masashi Yanagisawa was looking for
ligands for “orphan receptors” with strong homologies to
known G protein receptors but no identified endogenous
ligands
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They identified two novel peptides which were synthesized only
in the hypothalamus
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When injected into the 3rd ventricle they induced feeding
They named the peptides Orexins, after the Greek word for
appetite
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Sleep/Neurology-The Orexin System*
Orexin/HCRT (ORX) neurons originate in the posterior and lateral
hypothalamus as a paired set of nuclei comprising a total of 50,00080,000 neurons
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The ORX system is comprised of neurons producing two ORX
neuropeptides (ORX-A and ORX-B or HCRT-1 and HCRT-2) with
projections throughout the CNS
Preprohypocretin (prepro-orexin) the precursor polypeptide is
composed of 130 residues, undergoes proteolytic cleaving to
produce ORX-A and ORX-B with 33 and 28 amino acids respectively
*Chemelli R, Willie J, Sinton C, et al. Narcolepsy in orexin knockout mice: molecular genetics of sleep regulation.
Cell. 1999;98:437-451.
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Sleep/Neurology-The Orexin System
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ORX-A is highly lipophilic, crosses the BBB via simple
diffusion and is stable in the CSF
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ORX-B is a linear peptide that is not stable in the CSF, has a
short biological half-life due to rapid metabolism and
clearance
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Both peptides bind to two G-protein coupled receptors
(GPCRs)
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The OX1 receptor is a selective receptor with a high affinity
for ORX-A
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The OX2 receptor is a non-selective receptor with equal
affinity for both ORX-A and ORX-B
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Orexin and Orexin Receptors*
(Preprohypocreti
n)
(HCRT-1)
(HCRT-2)
*Inutsuka A, Yamanaka A. The Regulation of Sleep and Wakefulness by the Hypothalamic Neuropeptide
Orexin/Hypocretin. Nagoya J. Med. 2013;75:25-36.
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Knock-Out Mice*
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Yanagisawa’s group used transgenic techniques to construct a null
mutant mouse that did not produce either orexin peptide
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The mice had reduced food intake but not as much as had been
expected and had no effect on weight
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It was observed that the mice would often abruptly cease movement
(cataplectic attack)
*Chemelli R, Willie J, Sinton C, et al. Narcolepsy in orexin knockout mice: molecular genetics of sleep regulation.
Cell. 1999;98:437-451.
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Knock-Out Mice
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They then created mice without the OX2 receptor
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These animals exhibited slowness in movements consistent with
sleep attacks
Mice without the OX1 receptor
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Exhibited disrupted sleep but showed fewer signs of narcolepsy
than the OX2 receptor knockouts
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Orexin/Hypocretin and Genetic
Narcolepsy
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In 1985, Dement et al showed that canine genetic narcolepsy was
caused by an autosomal recessive gene1
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In 1999, Mignot identified the gene responsible for canine narcolepsy
as a mutated, non-functional version of the OX2 receptor gene1
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Human narcolepsy cannot be explained by genetic mutations in that
human narcolepsy is NOT a genetic disease (human narcolepsy is
discordant in identical twins)2
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In 2000, Siegel et al demonstrated that there is massive reduction
(>90%) of postero-lateral hypothalamic orexin neurons in human
narcolepsy/cataplexy pointing to a secondary loss of these cells while
at the same time sparing co-localized neurons (i.e. melanin
concentrating hormone)-raising the possibility of an auto-immune
attack against orexin neurons exclusively2
1Mignot
E. History of narcolepsy at Stanford University. Immunol Res. 2014;58:315-339.
J, Moore R, Thannickal T, al. e. A brief history of hypocretin/orexin and narcolepsy. Neuropsychopharmacology.
2001;25 (Suppl 5): S14-20.
2Siegel
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Sleep/Neurology-The Orexin System
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Orexins are excitatory neurotransmitters
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They act to sustain wakefulness and stabilize sleep
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Facilitatory role in the regulation of muscle tone
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Promote arousal responses to homeostatic challenges
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Drive motivated behavior such as seeking food
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Excite neurons of the mesolimbic reward pathways
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ORX antagonists have been shown to reduce the motivation to
seek drugs of abuse
Activated by humoral indicators of hunger such as low glucose or
high levels of ghrelin
+ Integrative Physiologic Roles of
Orexin/Hypocretin Peptides*
5-HT, serotonin; ACh, Acetylcholine; Arc, arcuate nucleus; BST, bed nucleus of the stria
terminalis; DA, dopamine; DR, dorsal raphe nucleus; GABA, gamma-aminobutyric acid;
HA, histamine; LC, locus coeruleus; LDT, laterodorsal tegmental nucleus; NA,
noradrenaline; NPY, neuropeptide Y; POA, preoptic area; POMC, proopiomelanocortin; PPN,
pedunculopontine tegmental nucleus; TMN, tuberomammillary nucleus; VTA, ventral
tegmental area
*Inutsuka A, Yamanaka A. The Regulation of Sleep and Wakefulness by the Hypothalamic Neuropeptide
Orexin/Hypocretin. Nagoya J. Med. 2013;75:25-36.
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Wake-Promoting Neurochemical
Systems
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Acetylcholine (ACh)
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Basal Forebrain (BF) contain ACh neurons that promote
wakefulness and REM sleep
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Also contains GABA which increases cortical activation by
inhibiting cortical interneurons
Pons
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Laterodorsal and pedunculopontine tegmental nuclei
(LDT/PPT) contain ACh neurons which project to subcortical
regions (active during wakefulness and REM sleep)
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Thalamus
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Lateral Hypothalamus
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Basal Forebrain
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Wake-Promoting Neurochemical
Systems
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Norepinephrine (NE)
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Locus coeruleus (LC)
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Fire most rapidly during wakefulness
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Less active during NREM sleep
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Virtually silent during REM sleep
Histamine (HA)
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Tuberomammillary nucleus (TMN)
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Fire most rapidly during wakefulness
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Less active during NREM sleep
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Least active during REM sleep
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Wake-Promoting Neurochemical
Systems
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Serotonin (5-HT) [Binds to at least 15 different receptors with
varied effects]
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Neurons in the dorsal raphe (DR) nucleus and other raphe nuclei
scattered along the midline of the brainstem
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Promote wakefulness and reduces REM sleep
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Fire most rapidly during wakefulness
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Less active during NREM sleep
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Least active during REM sleep
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Wake-Promoting Neurochemical
Systems*
Dopamine (DA)
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Ventral (and ventral-lateral)periaqueductal gray (vPAG/vlPAG) of the pons
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Potent wake-promoting effects
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Sleep promoting effects of DA antagonists
Orexin/Hypocretin
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ORX neurons are located exclusively in the lateral and posterior
hypothalamus
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Project widely and heavily innervate all arousal regions with particular
dense innervation of the LC and TMN
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ORX neurons fire mainly during wakefulness and are silent
during NREM and REM sleep
*Swick T. The Neurology of Sleep 2012. In: Teofilo Lee-Chiong J, MD, ed. Biology of Sleep. Vol 7. Philadelphia, PA: W.B.
Saunders; 2012:399-415.
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WAKE/AROUSAL and OREXIN NUCLEI*
Wake/Arousal Promoting Nuclei
Orexin Projections to Wake Promoting
Nuclei
*Espana R, Scammell T. Sleep neurobiology from a clinical perspective. Sleep. 2011;34:845-858.
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REM-NREM Sleep Switch
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Two populations of mutually inhibitory neurons in the upper
pons form a switch for controlling transitions between REM
and NREM sleep
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GABAergic neurons in the vlPAG and the adjacent LPT fire during
non-REM states to inhibit entry into REM sleep
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During REM sleep the vlPAG and LPT neurons are inhibited by
GABAergic neurons in the sublaterodorsal regions (SLD) that fire
during REM sleep
This mutually inhibitory relationship produces a REM-NREM
flip-flop switch, promoting rapid and complete transitions
between these states
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REM-NREM Sleep Switch
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The core REM switch is modulated by other neurotransmitter
systems
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NA neurons in the LC and 5-HT neurons in the DR inhibit REM
sleep work on both sides of the flip-flop switch (exciting REM-off
and inhibiting REM-on neurons
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During REM sleep these monoaminergic amine neurons are
silent whereas the ACh neurons (LDT/PPT) promote REM sleep by
having opposite actions on the same two neuronal populations
(vlPAG and the LPT)
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REM-NREM Sleep Switch
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Orexin neurons inhibit entry into REM sleep by exciting
neurons in the REM-off population (vlPAG/LPT)
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VLPO neurons promote entry into REM sleep by inhibiting
the vlPAG/LDT
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During REM sleep, glutamatergic neurons in the
sublaterodorsal (SLD) nuclei activate a series of inhibitory
interneurons in the medulla and spinal cord, inhibiting motor
neurons producing the atonia of REM sleep
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REM-NREM Sleep Switch*
GABAergic neurons in the vlPAG and the adjacent
LPT fire during non-REM to inhibit entry into REM
sleep. During REM sleep they are inhibited by a
Glutamatergic neurons in the SLD
population of GABAergic neurons in the
activates a series of inhibitory
sublaterodorsal region (SLD) that fire during REM
interneurons in the medulla and spinal
sleep. NA neurons in the LC and 5-HT neurons in the
cord, inhibiting motor neurons
DR inhibit REM sleep, during REM sleep they are
producing the atonia of REM sleep
silent.
*Saper C, Fuller C, Pedersen N, al. e. Sleep state switching. Neuron. 2010;68:1023-1042.
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State-Specific Firing Rates of Brainstem
and Cortical Neuronal Groups*
*Swick T. The Neurology of Sleep 2012. In: Teofilo Lee-Chiong J, MD, ed. Biology of Sleep. Vol 7. Philadelphia, PA: W.B.
Saunders; 2012:399-415.
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Narcolepsy/Cataplexy
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Pentad of symptoms [REM sleep characteristics that intrude into
wakefulness/sleep]
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Excessive daytime sleepiness
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Cataplexy
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Hypnogogic hallucinations
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Sleep paralysis
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Disrupted nocturnal sleep
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Narcolepsy/Cataplexy
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Canine narcolepsy caused by an exon-skipping mutation in
the OX2 receptor gene (1999)
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Murine narcolepsy found after the deletion of the gene
coding for both orexin peptides (severe sleepiness and
cataplexy) (1999)
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Humans with narcolepsy/cataplexy had a >90% loss of
orexin producing neurons (2000) with loss of other markers
of the orexin neurons (dynorphin and pentraxin) while
sparing intermingled MCH neurons
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High association with HLA DQB1*06:02 leading to the hypothesis
that orexin loss is secondary to an autoimmune process
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Narcolepsy/Cataplexy
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Impaired orexin signaling causes behavioral states to
become unstable
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Patients with narcolepsy/cataplexy have normal amounts of wake
and sleep but they have many more transitions between states
(accounts for disrupted nocturnal sleep)
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The loss of orexin neurons permits more frequent transitions into
and out of REM sleep throughout the day (pathognomonic of
narcolepsy)
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Can be partial REM sleep states, e.g. cataplexy (abrupt onset of
loss of muscle tone); hypnogogic hallucinations (onset of
dream mentation in the transition between wakefulness and
sleep); sleep paralysis (sleep onset muscle atonia)
+ Insomnia
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Characterized by
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Affects approximately 70 million Americans (one or more symptoms)
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Difficulty falling asleep
Difficulty staying asleep
Early AM awakenings
Poor quality of nocturnal sleep
Daytime consequences (psycho-motor complaints, memory deficits, etc.)
23.5 million (~10% of US adult population) have symptoms consistent with the
diagnosis of insomnia
8 million have had prescriptions written and 10 million use OTC or other sleep
aids (alcohol or herbal products)
Chronic insomnia is associated with Hyperarousal (hypermetabolic)
changes (as seen on PET scans from chronic insomnia patients*) in all
the wake-promoting neural centers of the brain stem, diencephalon and
cortex suggesting orexin overdrive
*Nofzinger E, Buysse D, Germain A, Price J, Miewald J, Kupfer D. Functional Neuroimaging Evidence for
Hyperarousal in Insomnia. Am J Psychiatr. 2004;161:2126-2129.
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Insomnia
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Prescription hypnotics typically work on augmenting sleep
promoting centers or blocking specific wake-promoting
centers
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GABA (enhance GABA signaling via GABAA receptors)
 Benzodiazepines (diazepam, clonazepam, lorazepam, triazolam)
 Non-Benzodiazepine receptor agonists [NBZAs] (zolpidem,
eszoplicone, zaliplon)
Histamine (Blocks HA H1 receptors in TMN)
 Central antihistamines (diphenhydramine, doxylamine,
hydroxyzine)
Block DA receptors (mainly D2)
 Typical antipsychotics (chlorpromazine, haloperidol,
thioridazine)
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Insomnia
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Orexin Receptor Antagonists*
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Evidence from null ORX knock-out mice showed that sleep propensity
was greatest in those animals where there was absence of both ORX
subtypes
If chronic insomnia is due to an overactive wake system then dampening
of the entire wake-promoting system should facilitate sleep by blocking
orexin activity
 This led to the development of dual orexin receptor antagonists
(DORA)
 Suvorexant (Belsomra®) was approved by the FDA in 2015 for the
treatment of insomnia (sleep onset and sleep maintenance) in patients
ages 18 and above
 Almorexant (another DORA) was withdrawn from consideration
before it got to the FDA approval stage
*Ruoff C, Guilleminault C. Hypocretin receptor antagonists for insomnia: rationale and clinical data. Clin
Invest. 2012;2(6):623-637.
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β-Amyloid and Orexin*
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In 2009 Holtzman’s group from Washington University in St. Louis
showed that the amount of interstitial fluid (ISF) levels of Aβ
correlated with wakefulness
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ISF Aβ significantly increased during acute sleep deprivation (in mice)
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ISF Aβ significantly increased during orexin infusion
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ISF Aβ decreased with infusion of a DORA
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Chronic sleep restriction significantly increased Aβ plaque formation in
amyloid precursor protein transgenic mice
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DORA infusion decreased Aβ plaque formation in amyloid precursor
protein transgenic mice
*Kang J, Lim M, Bateman R, et al. Amyloid-β Dynamics Are Regulated by Orexin and the Sleep-Wake Cycle. Science.
2009;326:1005-1007.
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Sleep-Disordered Breathing Advances
Cognitive Decline in the Elderly†
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In 2015, Osorio et. al. from NYU reported on patients from the
Alzheimer’s Disease Neuroimaging Initiative (ADNI) looked
at the following questions
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Is sleep-disordered breathing (SDB) associated with an earlier
age of onset of mild cognitive impairment (MCI) or Alzheimer’s
disease (AD) onset?
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†Osorio
SDB is associated with sleep fragmentation and cyclical
hypoxemia/hypercarbia
Does treatment with CPAP delay the onset of cognitive decline?
R, Gumb T, Pirraglia E, et al. Sleep-disordered breathing advances cognitive decline in the elderly.
Neurology. 2015;84:1-8.
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Sleep-Disordered Breathing Advances
Cognitive Decline in the Elderly
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Results
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SDB patients had a younger age at onset of MCI and AD-dementia
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CPAP treated patients appeared to delay progression of cognitive
impairment
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Survival Curves of Age at MCI or ADDementia Onset*
Excluded patients with
missing data
Excluded patients with
missing data
Excluded pts with ambiguous group
allocation
Excluded pts with ambiguous group
allocation
Documented MCI or AD based on
F/U assessment
Documented MCI or AD based on F/U
assessment
*Osorio, RS, Gumb, T, Pirraglia, E, et al. Sleep disordered breathing advances
cognitive decline in
the elderly. Neurology, 2015;84:1-8.
+ Associations of Brain Lesions at Autopsy
with Polysomnography Features Before
Death

Honolulu-Asia Aging Study (prospective cohort study of
Japanese American men in Honolulu)
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Neuropathologic analysis
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Braak stage
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Neurofibrillary tangle and neuritic plaque counts
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Microinfarcts
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Generalized brain atrophy
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Lacunar infarcts
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Lewy body (LBs)
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Neuronal loss and gliosis in the locus coeruleus
+ Associations of Brain Lesions at Autopsy
with Polysomnography Features Before
Death
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167 patients (all males) were included in the analysis who
underwent polysomnography in 1999-2000 (mean age=84
years) and died through 2010 (mean 6.4 years to death)
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Results
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SpO2<95% was associated with higher numbers of microinfarcts
(OR=3.88; CI=1.10-13.76)

Greater SWS (Stage N3) duration was associated with less
generalized brain atrophy (OR=0.32; CI=0.10-1.03) and slower
cognitive score reductions
+ Associations of Brain Lesions at Autopsy
with Polysomnography Features Before
Death‡
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Discussion
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Men with lower SpO2 during sleep were more likely to have higher
levels of microinfarcts
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Men with less SWS (slow wave sleep) a presumed marker of poor sleep
quality and strongly associated with fragmented sleep had more brain
atrophy at autopsy
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Men with greater hypoxemia during REM sleep exhibited more gliosis
and neuronal loss in the LC (locus coeruleus)
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AHI (a measure used in standard definitions of OSA and by Medicare to
determine CPAP eligibility) and # of arousals were not associated with
any of the lesions whereas O2 saturation level was associated with
microinfarcts, the major lesion of vascular dementia
‡Gelber
R, Redline S, Ross G, et al. Associations of brain lesions at autopsy with polysomnography features before
death. Neurology. 2015;84(10.1212/WNL.001163):296-303.
+

Alzheimer’s disease: sleep, orexin and
cognitive decline‡‡
Liguori’s group from U. of Rome looked at 48 consecutive untreated AD
patients and 29 healthy controls (CSF ORX levels and PSG findings)
 AD patients were divided into two groups
 Mild AD (MMSE>21; 21 subjects)
 Moderate-severe AD (MMSE<21; 27 patients)
 Results

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Controls and mild AD patients had no significant difference in CSF Orexin
levels
Moderate to severe AD patients showed significantly increased orexin levels
compared to normal controls and mild AD patients
Moderate-severe AD patients exhibited more impaired nocturnal sleep
compared to normal controls and mild AD patients
The global AD group had orexin levels that were positively correlated with
total tau proteins and strictly related to sleep impairment
Cognitive impairment (as measured by MMSE) was correlated with sleep
structure deterioration (reduction in SWS and increased amounts of WASO)
‡‡Liguori C, Romigi A, Nuccetelli M, et al. Orexinergic System Dysregulation, Sleep Impairment, and Cognitive
Decline in Alzheimer Disease. JAMA Neuro. 2014;71(12):1498-1505.
+ Correlations between CSF tau and MMSE (WASO,
and SE) in Patient’s with Moderate to Severe
Alzheimer’s Disease
(Total tau protein)
(Phosphorylated tau protein)
(WASO)
(SE)
+ Correlations between CSF Orexin Levels
and PSG Data in Patient’s with AD
(WASO)
(SE)
(SOL)
+ Sleep Disruption Leads to Aβ Deposition in
Mice¶

In 2014, a team from Washington U published results of their
work on orexin knock-out mice looking at Aβ deposition
 Results showed that increase in sleep time slowed the
production of Aβ and sleep deprivation increased the amount
of Aβ pathology in the brain
¶Roh
J, Jiang H, Finn B, et al. Potential role of orexin and sleep modulation in the pathogenesis of Alzheimer’s disease. J Exp
Med. 2014;211(13):2487-2496.
+
Why Do We Need Sleep?
 Why
is sleep restorative?
 Why
does lack of sleep impair brain function?
 Sleep




deprivation
Reduces learning
Impairs performance in cognitive and motor tests
Prolongs reaction time
Common cause of seizures
+
Why Do We Need Sleep?

Proteins linked to neurodegenerative diseases (β-amyloid, αsynuclein and tau) are present in the interstitial space
surrounding cells of the brain

The brain lacks a conventional lymphatic system to remove
excess interstitial proteins into the general circulation for
degradation in the liver

CSF recirculates through the brain interchanging with
interstitial fluid (ISF) to remove interstitial proteins including
β-amyloid
+
Why Do We Need Sleep?

The convective exchange of CSF and ISF is organized around
the cerebral vasculature



CSF influx occurs around arteries
ISF efflux occurs along the veins
These pathways were named “glymphatic system” on the
basis of their dependence on astrocytic aquaporin-4 (AQP4)
water channels with results equivalent to the peripheral
lymphatic removal of interstitial metabolic byproducts and
toxins


Removal of AQP4 channels reduce clearance of Aβ by 65%
The interstitial concentration of Aβ is higher in the awake state
compared to the sleep states in rodents and humans
+

Why Do We Need Sleep?
Nedergaard and associates* from U of Rochester in 2013
hypothesized that Aβ clearance increased with sleep and
the sleep-wake cycle regulates glymphatic clearance of
metabolic “neurotoxins”
that
 Using
real-time assessments of tetramethylammonium
(TMA) diffusion and two-photon imaging in live mice it
was demonstrated
 Natural sleep or anesthesia was associated with a 60%
increase in interstitial space

Increasing the convective fluxes of ISF which in turn
increased the rate of β-amyloid clearance during sleep
*Xie L, Kang H, Xu Q, et al. Sleep Drives Metabolite Clearance from the Adult Brain. Science.
2013;342:373-377.
+Sleep Drives Metabolite Clearance from
the Brain
*P<0.05 compared with
awake
A) Time-disappearance curves of I-Aβ after its injection into the frontal cortex in awake
(orange triangles), sleeping (green diamonds) and anesthetized (red squares)
B) Rate constants derived from the clearance curves for I-Aβ infusion
C) Time-disappearance curves of C-inulin after its injection into the frontal cortex of
awake (orange triangles), sleeping (green diamonds) and anesthetized (red
squares)
D) Rate constants derived from the clearance curves for inulin infusion
+ Summary

The orexin system acts as the master conductor of the wake-promoting
system of the sleep/wake and REM/NREM states

The absence of orexin creates an unstable sleep/wake condition and
clinically is cause of narcolepsy/cataplexy

Sleep fragmentation, a common component of such varied conditions
as sleep apnea, insomnia, RLS, PLMDs, and AD is thought to be
secondary to, or associated with, orexin over activity

With an increase in orexin activity, the normal physiologic function of
the glymphatic system is perturbed, potentially leading to a decrease
in Aβ and Tau protein elimination and the development of MCI/AD

Treatment of sleep disordered breathing (and potentially other sleep
and neurologic conditions) with resultant fragmented sleep might
decrease the onset of vascular dementia as well as the development of
other degenerative neurologic disorders