Cutting Edge: February 2005 Journal of Biological Rhythms

Marta Muñoz, Stuart N. Peirson, Mark W. Hankins, and Russell G. Foster
Long-Term Constant Light Induces Constitutive Elevated Expression of mPER2 Protein in the Murine SCN: A Molecular Basis for Aschoff’s Rule?
J Biol Rhythms 20: 3-14.

Circadian rhythms in behavior, metabolism, and
physiology are based upon transcriptional/translational feedback loops involving
a core set of clock genes that interact to regulate their own expression. In
mammals, the SCN is the site of a master biological clock regulating circadian
locomotor rhythms. The products of the clock genes mPer1, mPer2, mCry1, and
mCry2 form essential components of both negative and positive elements within
the SCN oscillator. The primary aims of this study were to examine clock gene
abundance under longterm LL in an attempt to provide molecular correlates of the
lengthened tau and daily phase delays described by Aschoff’s rule. Wheel-running
behavior was recorded frommice maintained in eitherDDor LLfor 50 days. The
abundance of the clock genes mPer1, mPer2, mCry1, and mCry2 and their protein
products was then examined (every ~4 h) within the SCN using in situ
hybridization and immunocytochemistry. Under LL conditions, mPer1, mPer2, mCry1,
and mCry2 messages remained rhythmic, although the waveform of mCry2 was altered
compared to DD. In LL, mPER1, mCRY1, and mCRY2 protein levels were also rhythmic
and comparable to the patterns observed in DD. However, mPER2 is elevated and
constitutively expressed under LL. Thus, rhythmic expression of these clock
genes is not dependent on the rhythmic production of mPER2, and the acute
up-regulation of mPer1 and mPer2 described for short (nonparametric) light
pulses is not sustained under LL conditions. These findings suggest that mPER2
is important for the generation of phase delays in the molecular clockwork,
providing a possible molecular explanation for Aschoff’s rule: LL lengthens the
circadian period by inhibiting the degeneration of mPER2, and constitutively
elevated levels of mPER2 enhance the phase-delaying limb of the molecular
oscillator.

Hong-Nian Liu, Susumu Ohya, Shinji Furuzono, Jing Wang, Yuji Imaizumi, and Shinsuke Nakayama
Co-contribution of IP3R and Ca²+ Influx Pathways to Pacemaker Ca²+ Activity in Stomach ICC
J Biol Rhythms 20: 15-26.

Intracellular Ca2+ oscillations in interstitial cells
of Cajal (ICCs) are thought to be the primary pacemaker activity in the gut. In
the present study, the authors prepared small tissues of 100-to 300-µm diameter
(cell cluster preparation) from the stomach smooth muscle (including the
myenteric plexus) of mice by enzymatic and mechanical treatments. After 2 to 4
days of culture, the intracellular Ca2+ concentration ([Ca2+]i) was measured. In
the presence of nifedipine, a dihydropyridine Ca2+ channel antagonist,
spontaneous [Ca2+]i oscillations were observed within limited regions showing
positive c-Kitimmunoreactivity, a maker for ICCs. In the majority of cell
cluster preparations with multiple regions of [Ca2+]i oscillations, [Ca2+]i
oscillated synchronously in the same phase. Asmall number of cell clusters (8 of
53) showed multiple regions of [Ca2+]i oscillations synchronized but with a
considerable phase shift. Neither tetrodotoxin (250 nM) nor atropine (10µM)
significantly affected [Ca2+]i oscillations in the presence of nifedipine. Low
concentrations (40µM) of Ni2+ had little effect on the spontaneous [Ca2+]i
oscillation, but SK&F96365 (40µM) and Cd2+ (120µM) terminated it.
Applications of either 2-aminoethoxydiphenyl borate (10µM) or
xestosponginC(10µM) completely and rather rapidly (~2 min) abolished the
spontaneous [Ca2+]i oscillations. The results suggest that pacemaker [Ca2+]i
oscillations in ICCs are produced by close interaction of intracellular Ca2+
release channels, especially inositol 1,4,5-trisphosphate receptor (InsP3R) and
Ca2+ influx pathways, presumably corresponding to store-operated type channels.
Reverse transcription polymerase chain reaction examinations revealed expression
of TRPC2, 4, and 6, as well as InsP3R1 and 2 in ICCs.

Keisuke Hirai, Muneto Kita, Hiroyuki Ohta, Hisao Nishikawa, Yuu Fujiwara, Shigenori Ohkawa, and Masaomi Miyamoto
Ramelteon (TAK-375) Accelerates Reentrainment of Circadian Rhythm after a Phase Advance of the Light-Dark Cycle in Rats
J Biol Rhythms 20: 27-37.

In vivo pharmacological effects of ramelteon
(TAK-375), a novel, highly MT1/MT2-selective receptor agonist, were studied in
rats to determine ramelteon’s ability to reentrain the circadian rhythm after an
abrupt phase advance. Experiments were also conducted to assess the potential
cognitive side effects of ramelteon and its potential to become a drug of abuse.
After an abrupt 8-h phase shift, ramelteon (0.1 and 1 mg/kg, p.o.) and melatonin
(10 mg/kg, p.o.) accelerated reentrainment of running wheel activity rhythm to
the new lightdark cycle. Ramelteon (3-30 mg/kg, p.o.) and melatonin (10-100
mg/kg, p.o.) did not affect learning or memory in rats tested by the water maze
task and the delayed match to position task, although diazepam and triazolam
impaired both of the tasks. Neither ramelteon (3-30 mg/kg, p.o.) nor melatonin
(10-100mg/kg, p.o.) demonstrated a rewarding property in the conditioned
place-preference test, implying that MT1/MT2 receptor agonists have no abuse
potential. In contrast, benzodiazepines and morphine showed rewarding properties
in this test. The authors’ results suggest that ramelteon may be useful for
treatment of circadian rhythm sleep disorders without adverse effects typically
associated with benzodiazepine use, such as learning and memory impairment, and
drug dependence.

Michael R. Gorman, Magdalena Kendall, and Jeffrey A. Elliott
Scotopic Illumination Enhances Entrainment of Circadian Rhythms to Lengthening Light:Dark Cycles
J Biol Rhythms 20: 38-48.

Endogenously generated circadian rhythms are
synchronized with the environment through phase-resetting actions of light.
Starlight and dim moonlight are of insufficient intensity to reset the phase of
the clock directly, but recent studies have indicated that dim nocturnal
illumination may otherwise substantially alter entrainment to bright lighting
regimes. In this article, the authors demonstrate that, compared to total
darkness, dim illumination at night (<> 26 h. In the presence of dim nocturnal
illumination, however, a majority of hamsters entrained to Ts of 28 h or longer.
The presence or absence of a running wheel had only minor effects on entrainment
to lengthening light cycles. The results further establish the potent effects of
scotopic illumination on circadian entrainment and suggest that naturalistic
ambient lighting at night may enhance the plasticity of the circadian
pacemaker.

Jin Ho Park, Matthew J. Paul, Matthew P. Butler, and Irving Zucker
Binocular Interactions in the Entrainment and Phase Shifting of Locomotor Activity Rhythms in Syrian Hamsters
J Biol Rhythms 20: 49-59.

To assess binocular interactions and possible ocular
dominance in entrainment of circadian rhythms, Syrian hamsters maintained in LL
were subjected for several weeks to schedules of eye occlusion with opaque
contact lenses. In separate groups, the opaque lens was inserted into the left
or right eye for 12 h at the same clock time each day. The left and right eyes
of other groups were alternately occluded for 12 h each day, with initial
occlusion of either the left or right eye for different groups. Amajority of
hamsters entrained their locomotor activity rhythm when 1 eye was occluded for
12 h. The modified visual input imposed by covering 1 eye is sufficient to
induce entrainment. Locomotor rhythms of most animals in which the 2 eyes were
alternately occluded for 12 h each day phasedelayed onset of activity during the
1st few days of the lensing procedure; activity onset then free ran with tau<>

Melanie Rüger, Marijke C. M. Gordijn, Domien G. M. Beersma, Bonnie de Vries, and Serge Daan
Nasal versus Temporal Illumination of the Human Retina: Effects on Core Body Temperature, Melatonin, and Circadian Phase
J Biol Rhythms 20: 60-70.

The mammalian retina contains both visual and
circadian photoreceptors. In humans, nocturnal stimulation of the latter
receptors leads to melatonin suppression, which might cause reduced nighttime
sleepiness. Melatonin suppression is maximal when the nasal part of the retina
is illuminated. Whether circadian phase shifting in humans is due to the same
photoreceptors is not known. The authors explore whether phase shifts and
melatonin suppression depend on the same retinal area. Twelve healthy subjects
participated in a within-subjects design and received all of 3 light
conditions—1) 10 lux of dim light on the whole retina, 2) 100 lux of ocular
light on the nasal part of the retina, and 3) 100 lux of ocular light on the
temporal part of the retina—on separate nights in random order. In all 3
conditions, pupils were dilated before and during light exposure. The protocol
consisted of an adaptation night followed by a 23-h period of sustained
wakefulness, during which a 4-h light pulse was presented at a time when maximal
phase delays were expected. Nasal illumination resulted in an immediate
suppression of melatonin but had no effect on subjective sleepiness or core body
temperature (CBT). Nasal illumination delayed the subsequent melatonin rhythm by
78 min, which is significantly (p= 0.016) more than the delay drift in the
dim-light condition (38 min), but had no detectable phase-shifting effect on the
CBT rhythm. Temporal illumination suppressed melatonin less than the nasal
illumination and had no effect on subjective sleepiness and CBT. Temporal
illumination delayed neither the melatonin rhythm nor the CBT rhythm. The data
show that the suppression of melatonin does not necessarily result in a
reduction of subjective sleepiness and an elevation ofCBT. In addition, 100 lux
of bright white light is strong enough to affect the photoreceptors responsible
for the suppression of melatonin but not strong enough to have a significant
effect on sleepiness and CBT. This may be due to the larger variability of the
latter variables.

Stacy A. Clemes, and Peter A. Howarth
The Menstrual Cycle and Susceptibility to Virtual Simulation Sickness
J Biol Rhythms 20: 71-82.

Virtual simulation sickness (VSS) is a form of
visually induced motion sickness that can result fromimmersion in a virtual
environment (VE). As in their susceptibility to the sickness induced by real
motion, womenhave been reported to be more susceptible than men to VSS, yet the
reason for this difference is not known. The aim of the current study was to
investigate the influence of themenstrual cycle on susceptibility to VSS in 16
naturally cycling women and to compare the responses of this group with control
groups consisting of 1) 16 premenopausal women taking a combined monophasic oral
contraceptive and 2) 16 men. All female participants were immersed in a
nauseogenic VE on days 5, 12, 19, and 26 of their menstrual/pill cycle. These
days were chosen because they fall in line with peaks and troughs of ovarian
hormone levels. Menstrual cycle phase was confirmed by salivary estradiol and
progesterone levels. A 4-week "pseudo-cycle" was assigned to the male
participants. Hormone analysis revealed that 9 participants in the experimental
group had been tested at the desired phases of their cycle. These participants
exhibited a significant increase in susceptibility to VSS on day 12 of their
cycle. The hormone analysis also showed that the cycles of the 7 remaining
members of the experimental group had not precisely followed the expected
pattern, and so these people had been tested on days that did not coincide with
peaks and troughs of ovarian hormone levels. No consistent variation in
susceptibility was observed over the cycle in these volunteers. In addition, no
change in susceptibility was observed over the pill cycle of the oral
contraceptive group nor over the pseudo-cycle applied to the male control group.
The authors conclude that susceptibility to VSS varies over the menstrual cycle
as a consequence of hormonal variation.

Florian Geier, Sabine Becker-Weimann, Achim Kramer, and Hanspeter Herzel
Entrainment in a Model of the Mammalian Circadian Oscillator
J Biol Rhythms 20: 83-93.

To adapt the timing of processes regulated by the
circadian clock to seasonally varying photoperiods, the phase relation between
the circadian clock and dusk or dawn ("phase of entrainment") must be tightly
adjusted. The authors use a mathematical model of the molecular mammalian
circadian oscillator to investigate the influence of the free-running period (tau)
and the shape of the PRC on the phase of entrainment. They find that a
phase-dependent sensitivity ("gating") of light-induced period gene
transcription enables a constant phase relation to dusk or dawn under different
photoperiods. Depending on the freerunning period tau and on the shaping of the
PRC by gating, the model circadian oscillator tracks either light onset or light
offset under different photoperiods. The study indicates that the phase of
entrainment of oscillating cells can be systematically controlled by regulating
both gating and the free-running period (tau).

Category: Cutting Edge Research

Tags: circadian