09 - 1.9 Chronobiology

1.9 Chronobiology

treatment outcomes in major depressive disorder. Psychiatr Clin North Am. 2007;30:105. Jarahi M, Sheibani V, Safakhah HA, Torkmandi H, Rashidy-Pour A. Effects of progesterone on neuropathic pain responses in an experimental animal model for peripheral neuropathy in the rat: A behavioral and electrophysiological study. Neuroscience. 2014;256:403–411. Winterer G, McCarley RW. Electrophysiology of schizophrenia. In: Weinberger DR, Harrison PJ. Schizophrenia. 3rd ed. Hoboken, NJ: Blackwell Publishing Ltd; 2011:311. 1.9 Chronobiology Chronobiology is the study of biological time. The rotation of the Earth about its axis imposes a 24-hour cyclicity on the biosphere. Although it is widely accepted that organisms have evolved to occupy geographical niches that can be defined by the three spatial dimensions, it is less appreciated that organisms have also evolved to occupy temporal niches that are defined by the fourth dimension—time. Much like light represents a small portion of the electromagnetic spectrum, the 24-hour periodicity represents a small time domain within the spectrum of temporal biology. A broad range of frequencies exist throughout biology, ranging from millisecond oscillations in ocular field potentials to the 17-year cycle of emergence seen in the periodic cicada (Magicicada spp.). Although these different periodicities all fall within the realm of chronobiology, circadian (Latin: circa, about; dies, day) rhythms that have a period of about one day are among the most extensively studied and best understood biological rhythms. A defining feature of circadian rhythms is that they persist in the absence of time cues and are not simply driven by the 24-hour environmental cycle. Experimental animals housed for several months under constant darkness, temperature, and humidity continue to exhibit robust circadian rhythms. Maintenance of rhythmicity in a “timeless” environment points to the existence of an internal biological timing system that is responsible for generating these endogenous rhythms. The site of the primary circadian oscillator in mammals, including humans, is the suprachiasmatic nucleus (SCN), located in the anterior hypothalamus. The mean circadian period generated by the human SCN is approximately 24.18 hours. Like a watch that ticks 10 minutes and 48 seconds too slowly per day, an individual with such a period gradually comes out of synchrony with the astronomical day. In slightly more than 3 months, a normally diurnal human would be in antiphase to the day–night cycle and thus would become transiently nocturnal. Therefore, a circadian clock must be reset on a regular basis to be effective at maintaining the proper phase relationships of behavioral and physiological processes within the context of the 24-hour day. Although factors such as temperature and humidity exhibit daily fluctuations, the environmental parameter that most reliably corresponds to the period of Earth’s rotation around its axis is the change in illuminance associated with the day–night cycle. Accordingly, organisms have evolved to use this daily change in light levels as a time cue or zeitgeber (German: zeit, time; geber, giver) to reset the endogenous circadian clock. Regulation of the circadian pacemaker through the detection of changes in

illuminance requires a photoreceptive apparatus that communicates with the central oscillator. This apparatus is known to reside in the eyes, because surgical removal of the eyes renders an animal incapable of resetting its clock in response to light. The circadian clock drives many rhythms, including rhythms in behavior, core body temperature, sleep, feeding, drinking, and hormonal levels. One such circadianregulated hormone is the indoleamine, melatonin. Melatonin synthesis is controlled through a multisynaptic pathway from the SCN to the pineal gland. Serum levels of melatonin become elevated at night and return to baseline during the day. The nocturnal rise in melatonin is a convenient marker of circadian phase. Exposure to light elicits two distinct effects on the daily melatonin profile. First, light acutely suppresses elevated melatonin levels, immediately decreasing them to baseline levels. Second, light shifts the phase of the circadian rhythm of melatonin synthesis. Because melatonin can be assayed easily, it provides a convenient window into the state of the circadian pacemaker. Any perturbation of the clock is reflected in the melatonin profile; thus melatonin offers an output that can be used to study the regulation of the central circadian pacemaker. SLEEP AND CIRCADIAN RHYTHMS Sleep Regulation Restful consolidated sleep is most appreciated when sleep disturbances are experienced. Sleep is the integrated product of two oscillatory processes. The first process, frequently referred to as the sleep homeostat, is an oscillation that stems from the accumulation and dissipation of sleep debt. The biological substrates encoding sleep debt are not known, although adenosine is emerging as a primary candidate neuromodulator of the sleep homeostat. The second oscillatory process is governed by the circadian clock and controls a daily rhythm in sleep propensity or, conversely, arousal. These interacting oscillations can be dissociated by housing subjects in a timeless environment for several weeks. The circadian cycle in arousal (wakefulness) steadily increases throughout the day, reaching a maximum immediately before the circadian increase in plasma melatonin (Fig. 1.9-1). Arousal subsequently decreases to coincide with the circadian trough in core body temperature. Experiments imposing forced sleep schedules throughout the circadian day have shown that an uninterrupted 8-hour bout of sleep can only be obtained if sleep is initiated approximately 6 hours before the temperature nadir. This nadir typically occurs at approximately 5:00 AM to 6:00 AM. In healthy individuals, initiating sleep between 11:00 PM and 12:00 AM affords the highest probability of getting 8 solid hours of sleep.

FIGURE 1.9-1 Relative phase relationship of sleep in young adults to other circadian phase markers. (From Dijk D-J, Lockley SW. Invited review: Integration of human sleep-wake regulation and circadian rhythmicity. J Appl Physiol. 2002;92:852, with permission.) It should be stressed that diurnal preference varies among individuals as a function of age, endogenous circadian periods, and other factors. This variability is paralleled by physiology. Clinically, diurnal preference can be quantified using the Horne–Östberg (HO) Morningness-Eveningness Questionnaire (MEQ). In qualitative terms, morning people or morning larks tend to awaken earlier and experience the core body temperature minimum at an earlier clock time relative to night people or night owls. Sleep deprivation studies have shown that the homeostatic component of sleep is remarkably similar among individuals of similar age. (It should be noted that there is a well-established agedependent decline in sleep need.) Therefore, diurnal preference is dictated almost exclusively by the circadian component of sleep regulation. Circadian Sleep Disorders Advanced sleep phase syndrome (ASPS) is a pathological extreme of the morning lark

phenotype. An autosomal-dominant familial form of ASPS (FASPS) recently has been genetically characterized. Afflicted family members exhibit a striking 4-hour advance of the daily sleep–wake rhythm. They typically fall asleep at approximately 7:30 PM and spontaneously awaken at approximately 4:30 AM. Affected individuals have a single nucleotide polymorphism in the gene encoding hPER2, the human homolog of the mouse Per2 clock gene. This adenine-to-guanine nucleotide polymorphism results in serine-toglycine amino acid substitution that causes the mutant protein to be inefficiently phosphorylated by casein kinase Iε, an established component of the circadian molecular clockwork. Similarly, delayed sleep phase syndrome (DSPS) has been shown to be influenced by genetics. A length polymorphism in a repeat region of the hPER3 gene appears to be associated with diurnal preference in patients with DSPS, the shorter allele being associated with evening preference. The advent of the light bulb has extended the human day into the natural night. This encroachment on the night, although increasing productivity, has affected human sleep patterns (Fig. 1.9-2). Typical use of artificial lights results in a single, consolidated bout of sleep lasting approximately 8 hours. This pattern of sleep is uncommon among most other mammals, which typically experience more fractured sleep. Human sleep under more natural photoperiods, where the duration of the night is longer, becomes decompressed. Specifically, a bimodal distribution of sleep is observed; bouts of sleep occur in early and late night. Periods of quiet wakefulness are interspersed between the two primary bouts of sleep. This natural sleep pattern is more similar to the sleep patterns of other mammals.

FIGURE 1.9-2 Change in sleep structure in response to artificial lighting. Total sleep time is reduced, and periods of quiet wakefulness are abolished by extending daytime into nighttime through artificial lighting. (From Wehr TA, Moul DE, Barbato G, et al. Conservation of photoperiod-responsive mechanisms in humans. Am J Physiol. 1993;265:R846, with permission.) SEASONALITY The 24-hour period of the Earth’s rotation around its axis is unchanging. However, the Earth’s axis is tilted 23.45 degrees from the plane of its own orbit around the sun (the

ecliptic). As a result, the relative proportion of daytime to nighttime within the 24-hour astronomical day varies as the Earth proceeds through its orbit of the sun. Many organisms are capable of synchronizing physiology to the seasonal cycle to maximize survival. For example, precise seasonal cycles in reproduction are seen throughout the plant and animal kingdoms. Large mammals that typically have long gestation periods, such as sheep, conceive in the fall when the nights are long and the days are short, so birth occurs during the relatively mild season of spring. These animals are referred to as short-day breeders. Conversely, mammals with gestation periods of only a few weeks, such as hamsters, conceive and give birth during spring and summer, when the days are long and the nights are short. Hence, these animals are referred to as long-day breeders. Like circadian rhythms, many of these yearly (circannual) rhythms tend to persist in the absence of seasonal cues with endogenous periods of approximately 1 year. Melatonin and Seasonality The most reliable environmental parameter providing a faithful representation of the solar day is the day–night cycle. Similarly, the most reliable environmental parameter reflecting the progression through the seasons is the change in day length, the fraction of the 24-hour day between sunrise and sunset. In seasonally breeding animals, day length is physiologically encoded through the melatonin profile. As described previously, melatonin levels are elevated during the night. A long night, such as that experienced during the short day lengths of winter, results in an elevated melatonin profile of a relatively long duration. A short summer night, by contrast, results in a short duration of elevated melatonin. This seasonal signal is interpreted by the reproductive axis, resulting in an appropriate reproductive response. Melatonin’s role in transducing day length was elucidated by pinealectomizing seasonally breeding animals, thereby removing the primary endogenous source of melatonin. Melatonin was then infused in profiles mimicking long days or short days. The duration of elevated melatonin was the primary determinant of seasonal reproductive status, even when the infused profile was administered under a conflicting day length. Variations in other parameters, such as the amplitude of the melatonin profile, the amount of total melatonin synthesized, or the phase relationship of the profile to the light–dark cycle, are of limited importance in producing a humoral signal that transduces day length. Reproductive responses to changing day length can be dramatic. A male Siberian hamster (Phodopus sungorus) maintained in long days is reproductively competent and typically has a testicular weight of approximately 250 mg per testis. Under short days, however, the testes regress to approximately 15 mg per testis, representing a 94 percent decrease in testicular mass. The same degree of regression is observed in response to melatonin infusions that mimic short days. Communication of the hormonally transduced day length to the reproductive axis is likely to be mediated, at least partially, through melatonin receptors in the pars tuberalis of the pituitary gland. The exact mechanism remains unknown, but activation of these receptors is hypothesized to indirectly regulate an unidentified factor putatively named tuberalin. Tuberalin, in turn,

controls gene expression and prolactin release from lactotrophs in the adenohypophysis of the pituitary. Seasonality in Humans Whether humans are truly seasonal is still a point of considerable debate. Several lines of evidence exist that suggest the presence of a residual tendency toward seasonality. A peak in the rate of suicide occurs in the summer; this peak is cross-cultural. Birth rates also tend to show a seasonal variation; a small but distinguishable peak in the rate of births occurs in spring and summer. This pattern, however, is itself variable and is heavily influenced by unknown cultural and geographic factors. Of interest, the amplitude of the spring–summer birth rate peak has diminished as societies have become industrialized. The decompressed bimodal structure of human sleep during long nights indicates that the length of natural sleep is related to the length of the night. Potentially, a twooscillator system could function to maintain proper sleep patterns during changing photoperiods. Such a proposed system would consist of an evening oscillator that tracks the transition from day to night (dusk) and a morning oscillator that tracks the transition from night to day (dawn). The relative phase differences between these oscillators may encode the changing day lengths associated with the passing of the seasons. Biological evidence for a two-oscillator system exists in rodents and humans. The melatonin profile of many vertebrates, including some humans, is bimodal, with evening and morning peaks. In rodents, metabolic and electrophysiological studies of the SCN typically have been done in brain slices cut in the coronal plane. Results of electrophysiological studies conducted in brain slices cut in the horizontal plane have provided new insights. The action potential frequency in SCN neurons from horizontally cut preparations is bimodal, with peaks in the early and late subjective day. Furthermore, the interpeak interval varies as a function of the photoperiod in which the animal was housed. These studies lend credence to long-standing suspicions that the SCN of seasonally breeding mammals and, perhaps, nonseasonal mammals harbor a morning and evening oscillator that interact to convey day-length information. Effect of Aging In general, as humans age, the circadian period shortens, the circadian phase advances resulting in earlier waking times and bedtimes, the amplitudes of most circadian rhythms decrease, and dramatic phase shifts such as those caused by jet-lag are less tolerated. Again, a mouse model has provided interesting insight into the interaction of the aging process and the circadian clock. The effect of chronic jet-lag on aged mice has dramatic consequences on mortality. About half of aged mice forced to phase advance 6 hours once per week survive this treatment compared with an 83 percent survival rate in unshifted age-matched mice. Aged mice subjected to weekly 6-hour phase delays show an intermediate survival of 68 percent. These profound effects of phase shifting are not observed in younger mice. The pathogenesis of chronic jet-lag remains to be determined.

Of interest, these mice did not have an increased rate of tumorigenesis. It is likely that in humans, as in mice, the internal desynchrony of oscillators that result from a rotating light schedule may have dire consequences that may be exacerbated by aging. CIRCADIAN RHYTHMS AND PHARMACOTHERAPY Circadian rhythmicity can be affected by drugs, and conversely, the circadian clock can modulate the efficacy of drugs throughout the course of the day. A better understanding of these interactions will lead to more effective pharmacotherapies. Some of the beststudied interactions between medications and the circadian clock have included the circadian effects of antidepressants. Elevated nocturnal body temperature is a common feature among depressed patients. This effect may be due to a reduced amplitude of the master circadian oscillator in the hypothalamus that drives body temperature. Tricyclic antidepressants (TCAs) and selective serotonin reuptake inhibitors (SSRIs) reduce elevated nocturnal body temperature while simultaneously enhancing circadian amplitude. Similarly, many depressed patients exhibit a dampened amplitude in daily activity rhythms. Like body temperature, the amplitude in daily activity cycles of depressed individuals may be augmented by TCA or SSRI therapy. The use of lithium to treat bipolar disorder has been long established. However, lithium also affects the circadian system, resulting in a lengthening of circadian period. The molecular mechanism by which this occurs remains unknown. Glycogen synthase kinase 3β (GSK3β) has been implicated in participating within the molecular clock mechanism. Of interest, GSK3β is inhibited by lithium. In cell culture, GSK3β has been shown to stabilize the negative clockwork regulator REV-ERBα via phosphorylation. REV-ERBα typically represses the transcription of the BMAL1 gene. In the presence of lithium, however, GSK3β is inhibited, thereby preventing the phosphorylation and stabilization of REV-ERBα, which as a consequence is targeted for proteasomal degradation. The degradation of REV-ERBα results in the de-repression of BMAL1 transcription. Whether lithium’s influence on circadian behavior is attributable to its inhibitory effect on GSK3β-mediated stabilization of REV-ERBα remains to be determined. Short-acting benzodiazepines (e.g., triazolam [Halcion] and brotizolam [Lendormin]) also exert chronobiological effects. In hamsters, triazolam or brotizolam administered during the middle of the subjective day induces circadian phase advances in activity. Brotizolam has been shown to reduce the light-induced expression of clock genes Per1 and Per2 in the SCN. Although benzodiazepines are allosteric modulators of γaminobutyric acid A receptors (GABAA), several lines of evidence indicate that the circadian effects of short-acting benzodiazepines require an intact serotonergic system. When the 5-HT1A/7 receptor agonist 8-hydroxy-2-(di-n-propylamino)-tetralin (8-OHDPAT) is injected into hamsters at subjective midday, phase advances in locomotor behavior and SCN neuronal activity are observed in addition to a reduction in Per1 and Per2 gene expression in the SCN. Recreational drugs of abuse also affect the circadian system. 3,4-Methylenedioxymethamphetamine (MDMA), or “ecstasy,” can act as a

serotonin neurotoxin. Hamsters treated with MDMA showed reduced triazolam-induced phase shifts in circadian locomotor activity and a diminished ability to reentrain rhythms posttreatment. MDMA-treated animals exhibited a reduction of serotonergic axonal terminals in the SCN, again emphasizing the importance of an intact serotonergic system in the regulation of the circadian axis. Recreational use of methamphetamine has increased dramatically. Chronic administration of methamphetamine disorganizes rodent activity rhythms. However, administration of methamphetamine to rodents rendered arrhythmic through ablation of the SCN results in a reemergence of rhythmicity. The mechanism involved in the rescue of rhythmicity or site of action remains unknown. The efficacy and toxicity of many pharmacotherapeutics vary as a function of circadian phase. Daily variations in fixed-dose lethal toxicity have been appreciated in rodents for years. Many anticancer drugs, ranging in mechanism from antimetabolites to deoxyribonucleic acid (DNA) intercalators to mitotic inhibitors, have been shown to have 2- to 10-fold changes in tolerability in rodents over the course of the day. Much of this difference is attributed to circadian variations in the body’s ability to absorb, distribute, metabolize, and eliminate toxic compounds. These four processes are affected by underlying circadian rhythms in physiological processes such as daily variations in gastric pH, gastrointestinal mobility, glomerular filtration rate, and membrane viscosity. The rhythmic intake of food during traditionally timed meals also influences how therapeutic drugs are handled by the body. It is becoming clear that to maximize efficacy and minimize toxicity of drugs, the circadian phase of administration must be considered. Appropriate circadian timing of the administration of multiple drugs can be a daunting challenge to infirmed individuals or their caretakers. The development of small implanted programmable pumps that can be directed to administer anticancer drugs or other therapeutics at particular times of day may provide a limited solution to this challenge. The emergence of the field of chronotherapy is a reflection of our increased understanding of the impact of the circadian system on the effectiveness of pharmacological treatments. REFERENCES Delezie J, Challet E. Interactions between metabolism and circadian clocks: Reciprocal disturbances. Ann N Y Acad Sci. 2011;1243:30. Dridi D, Zouiten A, Mansour HB. Depression: chronophysiology and chronotherapy. Biol Rhyth Res. 2014;45:77–91. Eckel-Mahan K, Sassone-Corsi P. Metabolism and the circadian clock converge. Physiol Rev. 2013;93(1):107. Glickman G, Webb IC, Elliott JA, Baltazar RM, Reale ME, Lehman MN, Gorman MR. Photic sensitivity for circadian response to light varies with photoperiod. J Biol Rhythms. 2012;27(4):308. Gonnissen HK, Rutters F, Mazuy C, Martens EA, Adam TC, Westerterp-Plantenga MS. Effect of a phase advance and phase delay of the 24-h cycle on energy metabolism, appetite, and related hormones. Am J Clin Nutr. 2012;96:689. Lanzani MF, de Zavalía N, Fontana H, Sarmiento MI, Golombek D, Rosenstein RE. Alterations of locomotor activity rhythm and sleep parameters in patients with advanced glaucoma. Chronobiol Int. 2012;29(7):911. Loddenkemper T, Lockley SW, Kaleyias J, Kothare SV. Chronobiology of epilepsy: Diagnostic and therapeutic implications of chrono-epileptology. J Clin Neurophysiol. 2011;28:146.

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