Introduction : A brief
communication in this same journal (Anias et al.
1984) described the existence of circadian
variations in the frequency of spontaneous
yawning in the rat. The observations were
made on a line of Sprague-Dawley rats,
genetically selected to obtain a higher
incidence of yawning (HY rats) (Urba-Holmgren et
al. 1990). In that report an effort was made to
correlate diurnal changes in yawning frequency
with circadian variations in activity (as
reviewed in the literature) of the
neurotransmitter systems that had been
postulated as participating in the central
control and regulation of yawning (Anias et al.
1984). Since yawning frequency was highest
during the last light hour, both under natural
and artificial light-dark (LD) illumination
schedules, it was suggested that the LD
transition might be the "primary synchronizer"
of the circadian rhythm of yawning. But that
suggestion was not supported by a formal
differentiation between circadian rhythms
proper, which are capable of free-running in
absence of the entraining stimuli, and daily or
diurnal rhythms, which "are induced and
dependent upon rhythmic occurrence of
environmental events" (Moore 1980). Nor had the
possibility been explored that yawning behaviour
might be entrainable to "secondary
synchronizing" stimuli, as restricted food or
water availability which had been proved to be
important synchronizers of several behavioural
rhythms (Krieger 1974, Edmonds and Adler 1977,
Krieger et al. 1977, Morimoto et al. 1977,
Takahashi et al. 1977, Krieger and Hauser 1978,
Sulzman et al. 1978, Morimoto et al. 1979,
Phillips and Mikulka 1979, Boulos 1980, Moore
1980, Coleman et al. 1982, Dhume and Cogate
1982, Inouye 1982, Honma et al. 1983,
Mistlberger and Rechtschaffen 1984) (for recent
reviews see Rusak 1981, Takahashi and Katz 1982,
Hiroshige 1984, Turek 1985, Rosenwasser and
Adler 1986). The purpose of the present work is
to study spontaneous yawning behaviour in rats
maintained under continuous illumination, and
examine if food availability, regularly
restricted to 2 or 3 daytime hours can entrain a
rhythmic variation in yawning, both in rats
under constant light, and in animals under
normal 12-12 LD cycles.
Methods : This study is based on the
observation of several groups of male
Sprague-Dawley rats (46 animals in total), from
the 8th, 9th, 13th and 16th generations of the
HY subline, genetically selected at our Animal
House. Their general housing and feeding
conditions and the method we use for the
observation of yawning have already been
described (Anias et al. 1984, Holmgren et al.
1985). At the preliminary stage we observed
yawning for 24 h a day in sessions of
différent durations irregularly
distributed throughout several days. But as this
procedure seemed to introduce unwanted
variability, in each experimental condition
yawning was at the end monitored during an
uninterrupted session of 24-26 h. Finally,
because of the inadequateness of averaging data
from différent animals in a free running
condition, we introduced a balanced experimental
design in which each, animal was its own control
and we monitored-yawning continuously during
56-60 h on two occasions,separated by 22 days.
Different group of animals were studied
exclusively under one of the following
conditions (A-D):
A. Constant light and ad libitum
feeding The observation of yawning
behaviour, under constant illumination (LL), was
done in twelve animals (two rats per cage) from
the 8th and 9th generations of the HY subline.
At the age of 2 months they were transferred
from the Animal House to a laboratory
observation room of 18 m' permanently
illuminated with two 80 watts 2.20 m long
fluorescent tubes from a 4 m high celling. The
room was not exclusive for the experimental
animals: during the day it was randomly used for
observation of other animals and study purposes.
Although silence was recommended, slight noise
and personnel traff ic were inevitable between
08:00 and 19:00. After 3 weeks of adaptation to
the LL regime, with food and water available ad
libitum, yawning occurrence was monitored
continuously during 24 h by trained couples of
observers replacing each other every 2 h.
Finally, two additional groups of four rats
each, from HY F13, after the same period of
adaptation to LL, were subject to yawning
monitoring in uninterrupted observation sessions
of 24 h.
B. Restricted daytime feeding in
animalsunder constant illumination For this
purpose the rats were changerd to clean boxes
every day, at regular intervals before giving
them their daily ration of standard laboratory
rodent pellets (Purina), placed directly on the
layer of wood shavings covering the floor of the
boxes. During the adaptation period to
restricted feeding, which lasted 18 days, the
animals were sometimes weighed before and after
eating, in order to estimate their daily food
intake. Excess of food was rernoved from the
cages at the end of the feeding time. The
animals had free access to water over 24 h. Two
groups of 4 rats each and one of 6 animals were
subject to three slightly différent
feeding and observation schedules: one group had
food avallable during 3 h (14.00 to 17.00) and
the observation sessions took place over 30 days
after the initial period of adaptation to this
regime; the second group had a restricted
feeding time (RFT) of only two hours (12.00 to
14.00),the observations being made during 45
days after the adaptation period; the third
group had food available during two and a half
hours (08:00 to 10:30) and was observed
continuously in a single session of 26 h, which
began with the animals eating their daily ration
of food pellets, and ended with the rats fasting
until 10:00 next day.
C. Restricted feeding ame in rats under a
LD schedule A group of six two month-old HY
rats, from the 9th generation, housed in two
cages with three animals per cage, were adapted
to a LD cycle (lights on from 7:00 to 19:00)
with food available between 9:00 and 11:00. Once
adapted to this regime for 3 weeks, the rats
were continuously observed and yawning monitored
during 26 h, starting from 9:00 one day to 11:00
next day.
D. Balanced experimental design with
animais being their own controls In these
experiments, we used six two month-old HY-males
from generation F16, randomly distributed in two
groups of three animals each and treated under
continuous illumination according to the
following feeding schedule.
E. Fasting experiments After
completing the observation of yawning along 24 h
of theday, some of the animal groups fasted
during 48 to 96 h, with yawning monitored
several times during that period in order to
study the evolution of spontaneous and drug
induced yawning under fasting conditions. These
experiments will be described in full
separately.
[...]
Discussion
The initial purpose of this work was to
establish whether the daily rhythm in yawning
behaviour described in the rat, with its peak in
frequency preceding the LD transition (Amas et
al. 1984) could bc properly considered as
circadian, i.e. as an endogenous rhythm governed
by a circadian pacemaker, entrainable to the LD
transition and capable of free-rurming in its
absence. In our experiments with rats adapted to
live under constant light, and fed ad libitum,
yawning activity exhibits several peaks during
the 24 h, as if under aperiodic environmental or
internal underlying influences, rather than
under the regular control of a circadian
pacemaker. The vectors representing maximal
yawning hour intervals of the différent
animals over 24 h seem to be randomly
distributed.
The loss in the diurnal rhythm of yawning
behaviour in rats exposed to constant
illumination while fed ad libitum does not
necessarily mean that this rhythms is not
endogenous. Several well documented circadian
rhythm, as those of plasma corticosteroid
concentration, body temperature levels and food
consurription, which in rats are entrainable to
the LD cycle, also tend to disappear under
constant dim light (Takahashi et al. 1977, Honma
and Hiroshige 1978a, 1978b, Morimoto et al.
1979) prolonged for more than one month. The
importance of the time of feeding in modifying
behavioural rhythms has been known for a long
time. Honma et al. (1983) recalled that already
in 1922 Richter described an increase in
locomotor activity in rats before the time of
feeding. Restricted periodical feeding has more
recently been used by many authors and in
different animals to entrain other biological
rhythms: locomotion, running activity, body
temperature, drinking and urinary excretion,
adrenocortical activity, etc..(Krieger 1974,
Edmonds and Adler 1977, Krieger et al. 1977,
Morimoto et al. 1977, Takahashi et al. 1977,
Phillips and Mikulka 1979, Boulos and Terman
1980, Moore 1980, Coleman et al. 1982, Inouye
1982, Honma et al. 1983, Hiroshige 1984,
Mistlberger and Rechtschaffen 1984, Shiraishi et
al. 1984). Our expeYiments with HY rats under
constant light and only one regular daily meal,
show that after 3 weeks' exposure to this
regime, a significant peak in yawning activity
anticipates the time of food availability.
In captive African lions and mandrils in a
Zoo, subjected to regular feeding times,
Baenninger (1987) has recently described a clear
peak in yawning during the last hour before the
animals recelved their food. But this does not
necqsarily mean that yawning might be a sign of
hunger, as suggested by Barbizet (1958). If
fasting was prolonged for more than 24 h
spontaneous yawning diminished, and practically
disappeared if the animals fasted 3 or 4 days.
Under these last conditions even
apomorphine-induced yawning was completely
blocked (Anias and Holmgren, unpublished
results). As to the relative potencies of the
L-D transition and the restricted meal time as
entrainers of the yawning daily rhythm, the
experiment illustrated in Fig. 2C seems quite
convincing: the peak in yawning just before dark
is absent. These results confirin the opinions
of other authors that the time of food
presentation (Edmonds and Adler 1977, Krieger
and Hauser 1978, Sulzman et al. 1978) or of
water availability (Dhume and Cogate 1982) may
be more potent synchronizers than the L-D cycle,
in relation to several rhythmic physiological or
behavioural variables. Since rats under a LD and
free feeding schedule eat mostly during the
early dark hours, this eating habit might be
permanently reinforcing the tendency for a
pre-dark peak in yawning to appear, making the
L-D transition look like a stronger synchronizer
than what it really is.
Yawning has commonly been considered to be
related to drowsiness preceding or following sleep
(Barbizet 1958). But increased yawning behaviour,
anticipatory to the restricted feeding time in
rats, seems to coincide with a state of alertness
of the animals, with increased locomotor activity
(Phillips and Mikulka 1979) or wheel running
(Edmonds and Adler 1977, Coleman et al. 1982, Honma
et al. 1983) and higher plasma levels of
corticosteroids (Krieger 1974, Morimoto et al.
1977). Food "expectancy" might, in a certain way,
be considered as a sort of psycho] ogical
stress or situational anxiety, condition which
recently, in a very brief editorial comment (Shader
and Greenblatt 1985), has been mentioned to be
positively correlated with yawning in humans.
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