1. Sleep and Wakefulness Jonathan W. Bekenstein, M.D., Ph.D.
Associate Professor of Neurology
May 21, 2009

2. OBJECTIVES: Understand the normal human sleep cycle.
Grasp the mechanisms of the circadian clock.
Understand the retinohypothalamic pathway.
Know EEG characteristics of stages of sleep.
Know the neurotransmitter systems and nuclei in the reticular activating system responsible for transitions to different sleep stages.

4. Basic Principles
Humans spend about one-third of their lives sleeping. We do not know the entire function of sleep, but many functions are understood. These include energy conservation and restoration of brain glycogen levels.
Metabolism is diminished at night, and so is body temperature, both of which follow a circadian rhythm and generally follow a pattern of ambient temperature.

5. Developmental Changes
Sleep evolves and changes during life and development. Total sleep requirements change during life. Adolescents and young adults often require more sleep, up to 10 hours per day. Early school schedules often cause adolescents to have daytime sleepiness due to insufficient total sleep time.

6. Sleep at Different Ages
Normal sleep onset is 20 minutes

8. Why do we sleep?
Many experts hypothesize that since humans are highly visual beings, lack of visual stimulation and an inability to fully function in the dark may be part of why we sleep.
Sleep may be a way to shed accumulated body temperature from daytime activities and to reduce the production of body heat due to reduced activity. Growth occurs primarily during slow wave sleep in the early night, when pulsatile release of growth hormone is at its highest level.
Temperature regulation-Growth- Memory
Many experts hypothesize that since humans are highly visual beings, lack of visual stimulation and an inability to fully function in the dark may be part of why we sleep. Sleep may be a way to shed accumulated body temperature from daytime activities and to reduce the production of body heat due to reduced activity. Growth occurs primarily during slow wave sleep in the early night, when pulsatile release of growth hormone is at its highest level.

9. Sleep and Immune System
Interleukin-1 disrupts sleep-wake cycle through receptors in brainstem.
May need sleep to help fight infections.
Maintain fever during sleep and alter thermoregulation during infections.
Sleep deprivation impairs leukocyte function and DNA synthesis for up to 5 days.

10. Interleukins Induced by Serotonin
Serotonergic neurons of the dorsal raphe nucleus, as well as other wake-promoting neurons located in the brainstem and the posterolateral hypothalamus, inhibit sleep-promoting neurons in the anterior hypothalamus, the preoptic area and the adjacent basal forebrain63, 64. In turn, these rostral sleep-promoting neurons inhibit wake-promoting neurons in the brainstem and the posterolateral hypothalamus63, 64. Serotonin (also known as 5-hydroxytryptamine (5-HT)) also induces the synthesis and/or release of sleep-promoting factors, which subsequently inhibit rostral wake-promoting neurons and activate rostral sleep-promoting neurons of the hypothalamus and basal forebrain. Interleukin 1 (IL-1) may be one of the 5-HT-induced sleep factors because serotonergic activation induces IL-1 mRNA expression in the hypothalamus89 and IL-1 inhibits wake-promoting neurons in the hypothalamic preoptic area/basal forebrain82. IL-1 also inhibits wake-promoting serotonergic neurons in the dorsal raphe nucleus74, 75. The schema is not intended to depict all interrelationships between the neuroanatomic regions and neurochemical systems involved in sleep regulation; rather, it is intended to illustrate the potential mechanisms by which 5-HT promotes wakefulness per se and at the same time stimulates the synthesis and/or release of sleep-promoting factors that then drive the sleep that naturally follows wakefulness52. ACh, acetylcholine; DA, dopamine; GABA, -aminobutyric acid; LC, locus coeruleus; LDT–PPT, laterodorsal and pedunculopontine tegmental nuclei; NA, noradrenaline; NREM, non-rapid eye movement; PeF, perifornical region; TMN, tuberomammillary nucleus; VTA, ventral tegmental area; W-REM on, neurons that are active during both wakefulness and rapid eye movement sleep.

11. Proposed principles by which changes in sleep architecture promote recovery from infection.
Infection-induced alterations in sleep are such that increases in non-rapid eye movement (NREM) sleep provide energy savings, and short NREM-sleep bouts reduce heat loss. The reduction in REM sleep allows the animal to shiver. The combined changes in NREM and REM sleep facilitate the production of fever. Fever imparts survival value because it increases the efficiency of many facets of immune function and alters the host environment to make it less favourable for pathogen reproduction97.

12. Possible Functions of Sleep
Protein synthesis and RNA transcription are most abundant during slow wave sleep, which may have implications for maintaining synaptic function or making memory more persistent. Lastly, mood and cognition, particularly concentration are adversely affected by sleep deprivation or sleep with frequent interruptions.

13. Circadian Rhythms and Sleep
(Circa=about; dia-day) Circadian rhythms are those that occur with about a 24 hour periodicity. Humans have an internal “clock” that runs in the absence of external clues about darkness and light. The internal clock “free-runs” with a period of about 24.2 hours, just slightly longer than a day on Earth. The internal clock can quickly be entrained with light shining into the eyes at a set time each day. Special cells in the retina contain a pigment called melanopsin. Axons from these retinal ganglion cells project to the suprachiasmatic nucleus of the anterior hypothalamus.

14. Paraventricular nucleus
Body of fornix and column of the fornix Midline blue is paraventricular nuc and lower blue is supraoptic nucle, medial and lateral hypothalamic nuclei (brown and red)
Supraoptic nucleus

15. Phototransduction
Phototransduction from the retina to the suprachiasmatic nuclei occurs primarily via the retinohypothalamic tract. Across a normal waking day, clock-dependent (circadian) alertness is usually lowest in the early morning and increases into the late afternoon or evening, thus opposing the growing sleepiness from having been awake all day.

16. Pathway for signaling Circadian light changes
Melanopsin containing retinal ganglion cells project to the suprachiasmatic nucleus of the anterior hypothalamus.
SCN projects to the paraventricular nucleus of the hypothalamus. Those cells send axons to the preganglionic sympathetic neurons in the intermediolateral zone in the thoracic spinal cord.
Intermediolateral cells project to the superior cervical ganglion.

17. Pathways (continued)
Postganglionic axons project to the pineal gland (pinecone shaped).
Melatonin is synthesized from L-tryptophan.
Melatonin levels rise in CSF about two hours before sleep onset and about 7 hours before the core body temperature falls to its nadir. Thus, sleep onset is 5 hours or so before the lowest core body temperature is reached.

19. Pathways

20. What Role Does Light Play?
Light inhibits melatonin production during the day.
Increasing Melatonin is required for sleep.
Serotonin is converted to Melatonin in the Pineal gland.

21. Synchronization of Circadian Timekeeping among SCN Neurons
Pacemaking neurons generate near-24-hr rhythms in gene expression, firing rate, and peptide release through a transcription-translation negative-feedback loop. These neurons are all GABAergic, and a subset in the ventral (core) SCN release VIP. VIP and its receptor VPAC2 are necessary for synchronization of circadian periods among SCN neurons. Daily GABA application can synchronize SCN neurons, and blockade of GABAA receptors interferes with rhythm coordination between the dorsal (shell) and ventral SCN. Gap junctions have also been
implicated in spike-for-spike synchrony between neighboring
SCN neurons.
Have a clock but need to synchronize all cell in the nucleus to be on the same time zone, so to speak.

23. Explanation
Robust rhythm generation in SCN neurons is thought to rely on circadian expression of Period (Per1, 2), Cryptochrome (Cry1, 2), Rev-Erb, Ror, and Bmal1 genes, and constitutive expression of Clock. Period and Cryptochrome proteins (PER/CRY) form a negative-feedback loop, repressing transcriptional activation by CLK/BMAL1 through E-Box sequences on Per and Cry promoters. Casein kinase 1 and (CK1Δ) causes a phosphorylation-dependent delay in PER/CRY feedback. REV-ERB and ROR proteins form a second feedback loop, binding to ROR-element (RORE) promoters of the Bmal1 gene to repress and enhance expression, respectively.

24. More than half of all SCN neurons require daily VIP-VPAC2 signaling to maintain robust rhythms. This signaling pathway may impinge on the intracellular molecular clockwork through activation of adenyl cyclase (AC), cAMP, protein kinase A (PKA), and CREB-dependent transcription of the Period genes (Travnickova-Bendova et al., 2002 and Itri and Colwell, 2003). This regulation of clock gene transcription may underlie the synchronization and amplification of neuronal rhythms by daily VIP/VPAC2 signaling. (Aton and Herzog, Neuron, 2005: )

26. Stages of Sleep and Some Basic EEG
Defined by EEG criteria, as behavioral distinction is difficult.
a. Alpha frequency: 8-13 Hz (described first)
b. Beta: > 13 Hz (described second)
c. Theta: 4-7 Hz
d. Delta < 4 Hz (for Death, Disease, Disability)

27. There is a progression through stages of sleep.
Drowsiness characterized by diminished eye blinks, diminution and cessation of the posterior dominant alpha rhythm, and some diffuse slowing of the background frequencies. Alpha is the predominant awake rhythm. Beta is seen in the frontal head regions during wakefulness. Theta is a normal rhythm of limbic structures and hippocampus in wakefulness.

28. Nightly Pattern of Sleep Stages

30. Fever Disrupts Normal Sleep Cycle
The relationship between fever and changes in sleep architecture is apparent when interleukin 1 (IL-1) is administered to rats. a | A representative hypnogram depicting sleep–wake cycles of a male Sprague–Dawley rat demonstrates arousal state-dependent changes in brain temperature. Shown is a record of sleep–wake states (green line) and brain temperature (red line) for consecutive 12 s epochs recorded for 12 h after intracerebroventricular injection of vehicle. The injection was given at the beginning of the dark portion of the light–dark cycle. The expanded inset shows the arousal state-dependent changes in brain temperature that occur under normal conditions in a freely behaving rat. Brain temperature declines before entry into and during non-rapid eye movement (NREM) sleep, whereas it increases at the onset of and during REM sleep. Increases in brain temperature that are associated with wakefulness (wake) are of greater magnitude than those that occur during REM sleep. b | After intracerebroventricular injection of 5.0 ng IL-1 into the same rat as in part a, NREM sleep is fragmented, REM sleep is abolished (green line) and fever ensues (red line). The expanded inset shows the extent to which NREM sleep is fragmented during fever. This inset depicts the effects of IL-1 on sleep during the third post-injection hour, the same period presented in the expanded inset of part a. In this animal, the effects of IL-1 on NREM sleep and REM sleep are apparent for almost 6 h. Once NREM–REM sleep cycles reappear, arousal state-dependent changes become apparent and brain temperature subsides to control values.

31. More Definitions
Other than in disease states and in damaged brain, Delta waves are associated with non-REM (rapid eye movement) sleep. Sleep spindles are waves of waxing and waning activity that have a fronto-central predominance at a frequency between 7-16 Hz. They are generated by reticular thalamic cells that are GABAergic.

32. Non-REM Sleep:
Stage I: more theta activity, loss of alpha, and vertex sharp waves
Stage II: theta and delta waves, vertex sharp waves, K complexes (a vertex sharp wave with a superimposed slow and fast component), and sleep spindles.

33. Stage II

34. Sleep Stages
Stage III: higher voltage delta waves that are 20-50% of the background over time.
Stage IV: more than 50% delta slow waves.
During non-REM sleep muscle tone is maintained at a moderate level and breathing is regular.

35. Stage III % Delta

36. Stage IV >50% Delta

37. REM Sleep Return of beta activity to the EEG
Loss of muscle tone except in diaphragmatic muscles
Rapid eye movements
Dreaming
Lasts about 10 minutes per epoch, on average.
Total REM time diminishes from 8 hours at birth to 2 hours at age 20 to 45 minutes at age 70.

38. REM
Note the absence of activity in the chin and leg EMG channels and the LOC and ROC (eye channels) showing eye movements.

40. PGO Waves in REM Sleep
Pontine-geniculo-occipital waves (PGO) waves originate in pontine reticular formation and propagate through lateral geniculate nucleus of thalamus to the occipital cortex in association with the rapid eye movements which may be related to visual hallucinations thought to be the major component of dreams.

41. Note Posterior Alpha
Alpha Activity

42. Vertex Sharp Wave: Stage I

43. Delta Waves: Stage IV Sleep

44. Sleep Spindles

45. EEG During Medical School Lecture
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46. Dolphins and Sleep Sleep with one eye open and one eye closed.
Cetaceans have the slow waves in one half of their brain at a time, while the other half of the brain has low voltage activity.
Each half of the brain exhibits approximately 4 h of SWS per day in the bottlenose dolphin.
No REM sleep in cetaceans.
Maintaining visual contact during sleep is of great importance to cetacean mothers and their newborn calves.

47. No REM in Dolphins
Allows them to generate heat in a “thermically challenging environment.”
Predator avoidance.
Activity of neurons of the locus coeruleus facilitate the maintenance of basal muscle tone. Basal muscle tone accounts for approximately 30% of the heat production required by the mammalian body.

48. Interhemispheric Synchronization
Much smaller corpus callosum in dolphins.
The reduced size of the corpus callosum may also be related to the hemispheric independence in the sleeping cetacean, a smaller connection resulting in a lowering of coherent activity between the two hemispheres.
Mice with abnormal corpus callosum don’t have synchronous EEGs.
Newborn humans don’t have synchronous EEGs. E

49. EEG recorded from two cortical hemispheres and two thalamus during waking (A), unihemispheric slow wave sleep in the right (B) and left (C) hemispheres in a bottlenose dolphin. Recording: 1—right cortex, 2—left cortex, 3—right thalamus and 4—left thalamus. Calibration 1 s and 200 μm (modified from Supin and Mukhametov, 1986.)
Most dolphins have a counterclockwise swim bias during sleep

51. Circuits in Sleep
Circuits involve pontine reticular formation, thalamus, and cortex
In the awake state and in REM sleep, cholinergic nuclei at the pontine-midbrain junction are active. These cells desynchronize the EEG and higher frequency, lower amplitude waves are seen.

52. Other Neurotransmitter Systems
Wakefulness also involves the locus coeruleus (norepinephrine), the raphe nuclei (serotonin), and the tuberomammillary nucleus (orexin/hypocretin). Orexin is a peptide neurotransmitter. Loss of orexin is thought to be cause of narcolepsy, analogous to loss of dopamine in Parkinson’s disease. May be an autoimmune problem (narcolepsy) since there is a relation to HLA type. May do CSF orexin levels as a diagnostic tool in the future. Orexin promotes waking and alertness.

53. REM and Muscle Paralysis

54. Feedback Loop from SCN
Activation of the ventrolateral preoptic nucleus of the hypothalamus induces sleep. Orexin containing neurons are inhibited. Cholinergic neurons in the pons-midbrain are also inhibited.
Melatonin probably helps to activate the VLPO to induce sleep.

55. Completing the Feedback Loop

56. Neurotransmitter and Anatomical Summary

57. Neurotransmitters Histamine linked to wakefulness and arousal.
Norepinephrine and serotonin affect muscle tone and arousal but are not tightly linked to maintaining arousal.
Anterior hypothalamus is the sleep center.
Posterior hypothalamus (histamine, orexin) and basal forebrain are the wake center (glutamate and acetylcholine).

59. Sleep and Memory
Post-learning sleep stabilizes learning, particularly of language and associations, such that there is resistance to interference from distractions during recall.
Sleep also improves recall of motor tasks.

60. Memory
Sleep improves and supports consolidation of both declarative and procedural memories. Declarative memory refers to retention of semantic facts and episodic events and has dependence on the hippocampal formation. Procedural memory refers to memory for motor skills and presumably relies on cortico-striatal and cortico-cerebellar circuits.

61. Which Brain Regions Involved?
Slow wave sleep supports consolidation of hippocampal dependent declarative memories.
REM sleep seems to preferentially support non hippocampal mediated memories. Memory may also be dependent upon the regular cycling between REM and non-REM sleep.

62. Memory Storage Areas
Medial temporal lobe is probably important in the retention of recent memories and neocortex probably important for storing remote memories.
Sleep may be a way to gradually incorporate newly acquired memories into neocortical networks for storage and retrieval.
Thus, recent studies suggest that consolidation of memory takes place during slow wave sleep as memories “redistribute” to neocortical memory systems.