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The circadian system of reptiles: A multioscillatory and multiphotoreceptive system


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Many parameters exhibited by organisms show daily fluctuations that may persist when the organisms are held in constant environmental conditions. Rhythms that persist in constant conditions with a period close to 24 h are called circadian. Although nowadays most research in this field is focused on the molecular and genetic aspects--and therefore mostly on two animal models (Drosophila and mouse)--the study of alternative animal models still represent a useful approach to understanding how the vertebrate circadian system is organized, and how this fascinating time-keeping system has changed throughout the evolution of vertebrates. The present paper summarizes the current knowledge of the circadian organization of Reptiles. The circadian organization of reptiles is multioscillatory in nature. The retinas, the pineal, and the parietal eye (and, possibly, the suprachiasmatic nuclei of the hypothalamus, SCN) contain circadian clocks. Of particular interest is the observation that the role these structures play in the circadian organization varies considerably among species and within the same species in different seasons. Another remarkable feature of this class is the redundancy of circadian photoreceptors: retinas of the lateral eyes, pineal, parietal eye, and the brain all contain photoreceptors.
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The circadian system of reptiles: a multioscillatory and
multiphotoreceptive system
Gianluca Tosini
*, Cristiano Bertolucci
, Augusto Foa
Neuroscience Institute, Morehouse School of Medicine, 720 Westview Drive, SW, Atlanta, GA 30310-1495, USA
Department of Biology, University of Ferrara, Via Borsari 46, Ferrara, Italy
Received 10 May 2000; received in revised form 14 September 2000; accepted 17 October 2000
Many parameters exhibited by organisms show daily fluctuations that may persist when the organisms are held in constant environmental
conditions. Rhythms that persist in constant conditions with a period close to 24 h are called circadian. Although nowadays most research in
this field is focused on the molecular and genetic aspects Ð and therefore mostly on two animal models (Drosophila and mouse) Ð the study
of alternative animal models still represent a useful approach to understanding how the vertebrate circadian system is organized, and how this
fascinating time-keeping system has changed throughout the evolution of vertebrates. The present paper summarizes the current knowledge
of the circadian organization of Reptiles. The circadian organization of reptiles is multioscillatory in nature. The retinas, the pineal, and the
parietal eye (and, possibly, the suprachiasmatic nuclei of the hypothalamus, SCN) contain circadian clocks. Of particular interest is the
observation that the role these structures play in the circadian organization varies considerably among species and within the same species in
different seasons. Another remarkable feature of this class is the redundancy of circadian photoreceptors: retinas of the lateral eyes, pineal,
parietal eye, and the brain all contain photoreceptors. D2001 Elsevier Science Inc. All rights reserved.
Keywords: Pineal; Parietal eye; Melatonin; Retina; Circadian rhythms; Reptiles; Vertebrate; SCN
1. Introduction
Many biochemical, physiological, and behavioral para-
meters exhibited by organisms show daily fluctuations, and
most of these fluctuations persist when the organisms are
maintained in constant environmental conditions, thus
demonstrating that they are driven by an endogenous
oscillator. Rhythms that persist in constant conditions with
a period close to 24-h are called circadian. Circadian
rhythms have been now described from bacteria to humans
and a large amount of information about the physiological,
cellular, and molecular mechanisms responsible for the
generation of circadian rhythmicity is now available. Circa-
dian rhythms are controlled by endogenous clocks that,
now, have been localized to discrete neural anatomical
structures within the nervous system. In vertebrates there
are some structures the removal of which has significant
effects on the behavioral circadian rhythmicity, and there-
fore they can be considered as part of the circadian system.
These structures are the suprachiasmatic nuclei of the
hypothalamus (SCN), the lateral eyes, and the pineal com-
plex. This set of organs constitutes what is now called the
``Vertebrates Circadian Axis'' [37]. Although these struc-
tures are present in all the vertebrates, their contribution to
the circadian system may vary considerably among classes
and even within the same class. The SCN, for example, are
the central circadian pacemaker in mammals, and their
lesion abolish almost all circadian rhythms (the only known
exceptions are some circadian rhythms within the retinas),
while in nonmammalian vertebrates the evidence for SCN
involvement in circadian rhythmicity is far less extensive.
The lateral eyes (retinas) contain self-sustained circadian
oscillators in all vertebrate classes and their removal may
affect physiological and/or behavioral rhythms in amphi-
bians, reptiles, birds, and also in mammals. In addition, in
mammals the eyes are the only structures capable of
perceiving light and thus necessary for circadian entrain-
ment [69].
The pineal gland is a central component in the regulation
of circadian rhythmicity of reptiles and other nonmamma-
* Corresponding author. Tel.: +1-404-756-5214; fax: +1-404-752-
E-mail address: (G. Tosini).
Physiology & Behavior 72 (2001) 461 ± 471
0031-9384/01/$ ± see front matter D2001 Elsevier Science Inc. All rights reserved.
PII: S 0031-9384(00)00423-6\
lian vertebrates [62], whereas the role the gland plays in the
circadian organization of mammals is marginal, since its
removal has little or no effects on overt rhythms [6].
Because of their phylogenetic position and ecology
reptiles have provided Ð and still provide Ð the
circadian field with some of the most interesting models
for understanding circadian organization, its evolution,
and its variability.
2. The pineal complex
The pineal complex (pineal gland and parietal eye) is a
morphologically and functionally related set of organs that
arises as an evagination of the roof of the diencephalon. The
pineal organ is present in almost all vertebrates (alligator
and owl have only a very rudimentary pineal organ) whereas
the parietal eye is present only in some lizards species and in
the tuatara (Sphenodon punctatus).
In Reptiles the pineal gland contains photosensory cells
with secretory activity. The major product of these secretory
cells is the hormone melatonin, and this hormone is believed
to play an important role in the circadian system of reptiles
(see below). Melatonin is synthesized from the amino acid
tryptophan via a well-known biosynthetic pathway. Because
of its capability to respond to changes in illumination and
temperature, the pineal gland is considered to be the photo-
thermoendocrine transducer (via the action of the hormone
melatonin) of changes in photoperiod and environmental
temperature [62].
The parietal eye has a lens, cornea, and retina; the
parietal eye retina is very simple (i.e. is made of photo-
receptors and ganglion cells only) and the photoreceptors
synapse directly onto the ganglion cells, the axons of which
form the parietal nerve. The parietal eye nerve innervates
several areas of the brain (but does not project to the visual
part). The parietal eye seems to be involved in many
physiological functions of lizards (thermoregulation, repro-
duction, and orientation), but, in general, its role seems
marginal or redundant. Almost unknown is the relationships
between the parietal eye and the pineal gland. The parietal
eye synthesizes melatonin [10,52], but in much lower
quantity with respect to the pineal gland. It is likely that
melatonin may simply fulfill a local function within the
parietal eye.
3. The pineal gland as circadian clock: in vitro studies
In most vertebrates melatonin levels (pineal and blood)
show a clear daily rhythmicity, and reptiles are not an
exception to this general pattern. For example, in Testudo
hermanni Ð during the activity season Ð melatonin is high
at night and low during the day [68]. Clear daily rhythms in
melatonin levels are present in the snake Nerodia rhombi-
fera [48], and in S. punctatus [11], in the lizard Anolis
carolinensis [59], Dipsosaurus dorsalis [27], Trachydo-
saurus rugosus [8± 10], Tiliqua rugosa [12], and Iguana
iguana [51,52]. In T. rugosus [8], A. carolinensis [59], D.
dorsalis [27], Podarcis sicula [14], and I. iguana [52] the
melatonin rhythm persisted also when the animals were held
in constant darkness and temperature, demonstrating there-
fore its true circadian nature.
However, the presence of a circadian melatonin rhythm
per se cannot be used as a reliable indicator of the presence
of a self-sustained oscillator, since the rhythmic melatonin
synthesis/release could be driven by circadian oscillators
located outside the pineal, as it occurs in mammals.
An easy way to demonstrate the presence or absence of a
circadian clock in the pineal is that of preparing an in vitro
culture of the gland for a few days (at least three), while
simultaneously measuring melatonin release at fixed inter-
vals of time. This approach was firstly pioneered in chicken
pineal [31] and since then has been applied to the pineal (but
also to the retina) of many other animals. In the lizards A.
carolinensis,Sceloporus occidentalis,D. dorsalis,I. iguana
(Iguanidae), and Christinus marmoratus (Gekkonidae) mel-
atonin synthesis and release persisted in isolated cultured
pineals for several days, and the synthesis/release was
rhythmic under light/dark cycles [27,35,36,42,44,52].
However, only the pineal of A. carolinensis,S. occiden-
talis, and I. iguana showed a persistent rhythm in melatonin
release when cultured in constant darkness and temperature,
thus demonstrating the presence of a circadian oscillator
Fig. 1. In vitro pattern of melatonin release (as percent of the mean) from
the pineal glands of three different species of iguanid lizards. A circadian
rhythm of melatonin release is present in A. carolinensis and I. iguana, but
not in D. dorsalis. The figures are redrawn from the following sources: A.
carolinensis [36], D. dorsalis [27], and I. iguana [52].
G. Tosini et al. / Physiology & Behavior 72 (2001) 461±471462
within the pineal itself (Fig. 1). On the other hand, cultured
D. dorsalis pineals secreted melatonin in large Ð but not
rhythmic Ð quantities (Fig. 1). Exposing the cultured pineal
to bright illumination greatly reduced melatonin synthesis
and/or release and abolished rhythmicity in I. iguana [49]
and in C. marmoratus [42].
In A. carolinensis and I. iguana the circadian rhythm of
melatonin synthesis/release from cultured pineal has been
shown to be temperature compensated [36,53] (Fig. 2).
Recent studies have also shown that the parietal eye may
contain a circadian clock controlling the synthesis/release of
melatonin [49,52].
Fig. 2. In vitro temperature compensation of melatonin circadian rhythm from cultured pineal gland of (A) A. carolinensis and (B) I. iguana. Although the
period of the rhythm is affected by the temperature, the rhythm is temperature compensated, since the Q
(1.1 ± 1.2, respectively) is in the range 0.8 ± 1.3
(redrawn from Menaker and Wisner [36] and Tosini and Menaker, unpublished data).
Table 1
Effects that manipulation of the circadian axis causes on reptiles' circadian rhythms
Species Manipulation Effects Reference(s)
A. carolinensis PINX abolishes, CRL, affects ELR [47,58]
S. olivaceus PARX no effect on CRL [55]
S. olivaceus PINX changes in CRL aand t[55]
S. olivaceus ENU changes in CRL t[65]
S. occidentalis PINX changes in CRL t[57]
S. occidentalis ENU changes in CRL t[57]
D. dorsalis PINX no effects on CRL [27]
D. dorsalis ENU changes in CRL t[27]
D. dorsalis SCNX abolishes CRL [28]
I. iguana PARX changes in tof CRL [52]
I. iguana PINX no effect on CRL, abolishes CRT [52]
I. iguana PINX affects BTS and ELR [40,51]
I. iguana ENU no effect [2]
G. galloti PINX abolishes CRL [41]
P. sicula PARX abolishes BTS for 1 week [24]
P. sicula PINX changes in t[13]
P. sicula PINX the effect changes with the season [25,26]
P. sicula PINX abolishes BTS for 2 ± 3 weeks [24]
P. sicula RETX marked changes in CRL t[13]
P. sicula ONX marked changes in CRL t[38]
P. sicula SCNX abolishes CRL [39]
PINX: pinealectomy; PARX: parietalectomy; RETX: retinalectomy; ENU: bilateral enucleation; ONX: optic nerve lesion; SCNX= electrolytic lesion to the
SCN. CRL = circadian rhythm of locomotor activity; CRT = circadian rhythms of body temperature; BTS = circadian rhythm of behavioral temperature
selection; ELR = ciracadian rhythm in the electroretinogram.
G. Tosini et al. / Physiology & Behavior 72 (2001) 461±471 463
4. The pineal and melatonin in the regulation of
circadian rhythms
The pineal gland is considered the neuroendocrine trans-
ducer of variation in environmental (light and temperature)
conditions, and such action is likely to be mediated via the
hormone melatonin. An easy way to address the role that the
pineal plays in the circadian organization is to remove this
gland and observe the effect that this removal has on the
circadian rhythms (see Table 1).
Circadian rhythms in locomotor activity have been
reported for several species of reptiles (see Ref. [62]).
Removal of the pineal gland abolishes circadian rhythms
of locomotor activity in A. carolinensis and in some
individuals of S. olivaceous [55,58], Gallotia galloti [41],
affects the period of the rhythm in S. olivaceous [55], S.
occidentalis [57], P. sicula [13,26], and has no effect in D.
dorsalis [27] or in I. iguana [52]. Pineal transplantation in
previously pinealectomized P. sicula induced significant
changes in the free-running period of locomotor activity
rhythms [18]. In I. iguana a circadian rhythm in body
temperature has also been demonstrated [50], and the
pineal organ is centrally involved in the generation and
control of this rhythm, since rhythmicity disappears after
its removal [52].
Circadian rhythms in behavioral thermoregulation have
been reported for several reptiles [7,30,46]. Pinealectomy
temporarily abolished the circadian rhythms in behavioral
thermoregulation in P. sicula [24], and reduced the ampli-
tude of these rhythms in I. iguana [51].
Finally, a circadian rhythm in electroretinogram has also
been reported in several lizards (review in Ref. [47]. Pine-
alectomy affected the amplitude of the circadian rhythm of
electroretinographic response in A. carolinensis and I.
iguana, suggesting an involvement of the pineal in the
modulation of this rhythm [40,47]
The behavioral effects of pinealectomy (see Table 1) are
likely to be mediated by melatonin, because of the following
observations: (i) pinealectomy greatly reduces the amount of
circulating melatonin and abolishes its circadian rhythmicity
(review in Ref. [49]); (ii) daily injections of exogenous
melatonin can entrain locomotor rhythms [4,66]; (iii) daily
12-h melatonin infusions that in S. occidentalis closely
mimic the normal, rhythmic pattern of pineal melatonin
secretion in this species entrain locomotor rhythms of both
pineal-intact and pinealectomized lizards [21]; (iv) chronic
administration of melatonin (in silastic capsules) lengthens
the period of circadian rhythms in S. olivaceous and S.
occidentalis [56,57], in D. dorsalis [27], and in P. sicula
[15]; (v) a phase response curve to melatonin has been
described in lizards (S. occidentalis) [61], and (vi) finally,
melatonin administration altered the circadian rhythm of
body temperature selection in I. iguana [51].
Parietalectomy (see Table 1) did not affect locomotor
rhythms in A. carolinensis and S. olivaceous [55,58] and in
P. sicula [24], while in I. iguana it produces slight changes
in the circadian rhythms of locomotor activity and body
temperature [52]. In P. sicula parietalectomy temporarily
abolishes (1 week) the circadian rhythm in behavioral
thermoregulation [24].
5. The pineal gland as seasonal clock
Investigations in the lizards A. carolinensis and T.
rugosa demonstrated that 24 h cycles of both light and
temperature can entrain the pineal melatonin rhythm and
that differences in length of daily photoperiod or thermo-
period affect the phase, amplitude, and duration of this
rhythm [12,59,67]. Hence, the current ambient lighting and
Fig. 3. Seasonal differences in the behavioral effects of pinealectomy in the
lacertid lizard P. sicula. (A,B) Means (  S.E.M.) of the absolute changes in
the free-running period of locomotor rhythms |Dt|) and circadian activity
time (|Da|) induced by pinealectomy (PIN-X) in different seasons and by
sham pinealectomy (SHAM). Pinealectomy was effective in altering the
free-running period (t) in all seasons. Changes in twere significantly
greater in summer than in winter, spring, and autumn. Circadian activity
time (a) was found to change significantly in response to pinealectomy
only in spring and summer. Since no seasonal differences in |Dt| and |Da|
were found among the four seasonal groups of SHAM, the data were
pooled. (C) Locomotor activity record of one lizard subjected to
pinealectomy (PIN-X) in summer. Each horizontal line is a record of 1
day's activity, and consecutive days are mounted one below the other.
Pinealectomy markedly lengthens t, shortens a, and abolishes the bimodal
locomotor pattern (redrawn from Refs. [25,26]).
G. Tosini et al. / Physiology & Behavior 72 (2001) 461±471464
temperature conditions (and their seasonal change) are
readily translated into an internal cue in the form of the
pineal melatonin rhythm. This cue can be used to regulate
both the daily and annual physiology of lizards [63]. In the
Fig. 4. Circadian locomotor activity of lacertid lizards P. sicula free-running in constant temperature (29°C) and darkness (DD). Lizards were collected and
subjected to daily melatonin injections in autumn (A), winter (B), spring (C), and summer (D), respectively. (A ± C) Starting and ending dates of melatonin
treatment are shown on the left of each record. The vertical line drawn through each record shows the time of day of melatonin injections during the whole
injection period. (D) On July 31st the time schedule of melatonin injections was advanced (Shift) from 7:00 p.m. to 3:00 p.m. On September 4th melatonin was
replaced with vehicle solution. Although the locomotor rhythms of the lizards tested in summer entrain successfully to the 24-h period of melatonin injections,
the locomotor rhythms of lizards tested in all other seasons do not entrain to the 24-h period of the injections (redrawn from Ref. [4]).
G. Tosini et al. / Physiology & Behavior 72 (2001) 461±471 465
Lacertid lizard P. s i c u l a the pineal was shown to be
involved in the seasonal reorganization of the circadian
system that is typical of this lizard [17,25]. In constant
temperature and constant darkness pinealectomy in P.
sicula actually induces an immediate transition from the
typical circadian locomotor behavior of summer, character-
ized by a marked bimodal pattern, short free-running
period (t) and long circadian activity time (a), to the
typical circadian locomotor behavior of autumn, character-
ized by an unimodal pattern, a long tand short a(Fig.
3C). Again, the behavioral effects of chronic implants of
exogenous melatonin (in silastic capsules) were found to
be the same as those of pinealectomy in summer: the
abolition of the bimodal pattern after application of the
implants was always associated with a lengthening in t
and shortening in a[15]. Robust circadian rhythms of
blood-borne melatonin expressed by intact P. sicula in late
summer become heavily disrupted or abolished in response
to either pinealectomy or melatonin implants [14]. Taken
together, these results strongly support the view that the
transition from a summer locomotor pattern to an autumn ±
winter one in response to both pinealectomy and melatonin
implants is due to the concomitant suppression of circadian
melatonin rhythms in the blood. Accordingly, in contrast to
the situation in summer, in autumn and winter circadian
rhythms of blood-borne melatonin do not seem to be
required for the expression of the locomotor pattern typical
of these seasons, and therefore in autumn± winter the
behavioral effects of pinealectomy are expected to be
substantially reduced with respect to those observed in
summer. For instance, in the tortoise T. hermanni annual
changes in melatonin rhythms have actually been shown to
occur under natural conditions, with maximal amplitude of
these rhythms in summer and their complete disappearance
in winter [68]. The results of an investigation that com-
pared systematically the effects of pinealectomy on circa-
dian locomotor behavior of P. sicula at different times of
the year actually confirmed the existence of marked annual
changes in the role of the pineal in the circadian organiza-
tion of this lizard [26]. Seasonal differences in the beha-
vioral effects of pinealectomy have been reported in one
other nonmammalian species, the burbot Lota lota [32]. In
the burbot, pinealectomy was found to induce a drastic
lengthening in tin winter, and a comparably drastic
shortening in tin summer. Hence, unlike the case of the
lizard P. sicula, the pineal in burbots is centrally involved
in determining circadian organization both in winter and in
summer. Further investigations demonstrated that daily
injections of exogenous melatonin are capable of entrain-
ing circadian locomotor rhythms of P. sicula exclusively
during the summer (Fig. 4) [4]. Altogether, these findings
demonstrate that the pineal Ð via its hormonal product
melatonin Ð is centrally involved in determining the
circadian organization of the Lacertid lizard P. sicula in
summer, while it is only marginally (or not at all) involved
in the other seasons. As mentioned before, the effects of
pinealectomy on circadian locomotor behavior of lizards
may vary consistently depending on the species (arrhyth-
micity, period changes, no effects). On the other hand,
because of the seasonal differences in the behavioral
effects of pinealectomy we have found in P. sicula,it
seems reasonable to doubt that the differences among
lizards are completely interspecific in nature. Instead they
may, at least in part, depend on the particular season in
which the behavioral effects of pinealectomy have been
examined in each different species. The Lacertidae, as well
as many Iguanidae, inhabit temperate zones, i.e. zones in
which seasonal changes in circadian organization are likely
to have evolved in response to the regular seasonal
fluctuations in photoperiod and/or thermoperiod experi-
enced by the lizards throughout the year [17,63]. Hence,
before deciding about interspecific differences, one should
verify whether, for instance, A. carolinensis and G. galloti
were tested in a season when the behavioral effects of
pinealectomy are maximal and whether D. dorsalis was
tested in a season when these effects are minimal.
6. The role of the retinas in the circadian system
The retinas of reptiles can participate in circadian func-
tion not only as photosensory inputs to the clock, but also as
loci of circadian oscillators: in I. iguana the retina isolated
in culture drives circadian rhythms of melatonin synthesis
[52]. Bilateral ocular enucleation under constant bright light
(LL) was found to induce a marked shortening in tin S.
olivaceus,S. occidentalis, and in some case arrhythmicity in
S. olivaceus [57,65]. Enucleation has a modest effect on the
circadian rhythms of body temperature and locomotor
activity in I. iguana, however enucleation plus pinealectomy
abolished both rhythms in 30% of the animals tested [2].
Bilateral retinalectomy induces a marked shortening in tin
P. sicula kept in constant darkness (DD). These data suggest
that the retinae may either (S. olivaceus) be a component of
the primary pacemaker that drives locomotor rhythms, or (S.
occidentalis,P. sicula) at least play an important modulating
role, that is independent of light perception (P. sicula), on
this primary pacemaker [13,34,57]. In P. sicula electrolytic
lesions of both optic nerves at the level of the optic chiasm
in DD produce the same behavioral effects as bilateral
retinalectomy [13,38]. This demonstrates that the influence
of the retinae on the circadian system of P. sicula is neurally
mediated. Accordingly, in the iguanid lizard S. occidentalis
the influence of the retinae on the circadian system appears
to be neurally mediated, since bilateral optic nerve section
induces marked changes in shape of the phase ± response
curve to light [60].
The retinas play also a role in entrainment of circadian
rhythms to LD cycles, since the light threshold for entrain-
ment is lower in sighted than in blinded S. olivaceus [54].
Several investigations made it clear that extraretinal
photoreceptors participate in mediating entrainment of cir-
G. Tosini et al. / Physiology & Behavior 72 (2001) 461±471466
cadian rhythms of reptiles to LD cycles. In nine species of
lizards, representing five different taxonomic families (Igua-
nidae, Gekkonidae, Eublepharidae, Xantusidae, and Lacer-
tidae) the locomotor rhythms can be entrained to LD cycles
after enucleation of the lateral eyes [23,54,60,64,65]. Hence,
intact retinas are not necessary for entraining behavioral
rhythms of lizards to LD cycles.
Studies carried out in the Iguanid lizards S. olivaceus and
the Lacertid lizard P. sicula showed that ablation of all
known photoreceptive structures (lateral eyes, pineal, and
parietal eye) in the same individual animal does not prevent
entrainment of their circadian rhythms of locomotor activity
to light [16,65]. Furthermore, shielding the brains of
blinded± pinealectomized S. olivaceus entrained to LD
cycles causes them to free-run. Taken together, these data
demonstrate the existence of brain photoreceptors mediating
entrainment of locomotor rhythms to LD cycles. Reptile
extraretinal photoreceptors must be quite sensitive, because
blind lizards can be entrained to an LD cycle as dim as 1 lx
[54]. Brain photoreceptors mediating entrainment have been
documented in Alligator missisipiensis [33].
Recent attempts to localize photoreceptors in the deep
brain by using antibodies were successful. In the Iguanid
lizards A. carolinensis and I. iguana anti-opsins antibodies
labeled neurons in the basal region of the lateral ventricles
[19,20]. A brain rhodopsin was recently cloned in P. sicula,
but its location in the brain is unknown [43].
Two different mechanisms, but not mutually exclusive,
have been proposed for entrainment [1,45]: in one, only
the transitions from light to dark and from dark to light
are considered effective for entrainment to 24-h LD
cycles (nonparametric entrainment); in the other, light
and darkness are assumed to exert a more or less
continuous action on the velocity of circadian oscillators
(parametric entrainment). The observation that the velo-
city of oscillators changes by changing the intensity of
LL (Aschoff's rule) supports the model of parametric
entrainment. In diurnal animals, for example, the light
portion of the 24 h LD cycle may increase the velocity of
the oscillators and the dark portion decrease their velocity,
with the net effect of entraining period of the zeitgeber.
Hence, it may be interesting to examine what array of
photoreceptors mediate the response of the circadian
system to changes in LL intensities. Underwood and
Menaker [65] investigated this aspect of circadian orga-
nization in S. olivaceus (Iguanidae) and P. sicula (Lacer-
tidae), by testing the locomotor behavior of these diurnal
lizards exposed to different levels of LL intensities after
bilateral enucleation. When intact, both S. olivaceus and
P. sicula obey Aschoff's rule for diurnal animals [22,65].
After blinding, S. olivaceus continuestoobeyto
Aschoff's rule, while P. sicula cannot discriminate among
different levels of LL and between LL and DD [65]. This
led to the conclusion that in S. olivaceus extraretinal
photoreceptors can mediate the response of the circadian
system to changes in level of LL, while in P. sicula this
function is accomplished only by the retinas of the lateral
eyes. In contrast with this, new investigations in P. sicula
showed clearly the existence in this lizard of extraretinal
photoreceptors that are capable of mediating the effects of
changing in level of LL on circadian locomotor behavior
(Fig. 5) [16]. Such a disagreement may depend on genetic
differences between animals, since the P. sicula used in
the experiments by Underwood and Menaker were col-
Fig. 5. Locomotor activity record of a Lacertid lizard P. sicula subjected to
pinealectomy (PIN-X) and then to retinalectomy (RET-X) under LL 30 lx.
This lizard can discriminate between different levels of LL (Aschoff's rule)
after PIN-X-RET-X: tshortens under LL 600 lx and lengthens under LL 30
lx. The final part of the record shows entrainment of the activity rhythm of
the PIN-X-RET-X lizard to a 24-h light cycle. These data show the
existence of extrapineal ± extraretinal photoreceptors in P. sicula (redrawn
from Ref. [16]).
G. Tosini et al. / Physiology & Behavior 72 (2001) 461±471 467
lected in north-east Italy and Croatia, whereas the P.
sicula used by Foa
Áet al. were collected in central Italy
(about 500± 700 km apart). Even if this interpretation is
correct, it is still unclear why some lizards can use
extraretinal photoreception to discriminate among different
intensities of LL, while others have only the retinas
available to accomplish this function.
7. The SCN and their role in the circadian organization
In mammals, namely rodents, the SCN have been
shown to contain a circadian multioscillator system that
acts as the primary (master) pacemaker for a host of
physiological and behavioral rhythms. During the last
decade the SCN have also been recognized to play a role
in the circadian system of lizards [28,39]. First of all, the
lizard SCN are topographically similar to the SCN of
rodents, and receive a direct retinal projection. As in
rodents, the lizard SCN lies just dorsal to the optic chiasm
and adjacent to the third ventricle, in the region of
transition from the preoptic area to the hypothalamus
(Fig. 6) [5]. Furthermore, as in rodents, the SCN of the
Iguanid lizard D. dorsalis were shown to bind antibodies
raised against neuropeptide Y [29]. Collectively, these
data strongly support the contention that the SCN of
lizards are homologous to the SCN of mammals. Electro-
lytic lesions to 90% or more of the SCN (SCN-X) were
found to abolish circadian rhythms of locomotor activity
both in D. dorsalis and P. sicula (Figs. 6C and 7A)
[28,39]. Except SCN lesions, no experimental treatment or
lesion has so far succeeded in abolishing circadian loco-
motor rhythmicity in both P. sicula and D. dorsalis. This
evidence supports the contention that in both species the
SCN contain the primary circadian pacemaker driving
locomotor rhythms.
Fig. 6. (A) Schematic reconstruction of a transverse brain section at the level of the SCN of the lizard P. sicula. The SCN lies just dorsal to the optic chiasm and
adjacent to the third ventricle, in the region of transition from the preoptic area to the hypothalamus. Dotted lines encompass the area of the section containing
the SCN. Magnifications of this area are presented in the photomicrographs of transverse brain sections reported in the right panel. Sections (B) and (C) are
stained with cresyl violet. Scale bar 40 mm. (B) Brain section of a lizard in which the SCN (indicated by arrows) remained intact. (C) Brain section of a lizard in
which the SCN (indicated by arrows) were completely lesioned. Abbreviations: cpp, posterior pallial commissure; Cxl, cortex lateralis; Cxm, cortex medialis;
DVR, dorsal ventricular ridge; lfb, lateral forebrain bundle; mfb, medial forebrain bundle; OC, optic chiasm; PH, nucleus periventricularis hypothalami; V, III
ventriculus (redrawn from Ref. [39]).
G. Tosini et al. / Physiology & Behavior 72 (2001) 461±471468
Other experiments confirmed the role of the SCN as
primary pacemaker in the P. sicula circadian system. Daily
injections of exogenous melatonin entrain locomotor
rhythms of intact, pinealectomized, and unilaterally SCN-
lesioned P. sicula, but are incapable of restoring rhythmicity
in subjects previously rendered arrhythmic by SCN-X (Fig.
7B) [4] Furthermore, the fact that the circadian rhythm of
behavioral temperature selection of P. sicula is not definitely
abolished after both parietalectomy and pinealectomy sug-
gests that the SCN or neighboring hypothalamic areas may
also be involved in driving this rhythm [24]. Interestingly, in
D. dorsalis the daily bout of voluntary hypothermia dis-
appears after lesion to the periventricular preoptic area of
the hypothalamus [3].
The SCN may be important to the circadian organization
of lizard species besides P. sicula and D. dorsalis. In both S.
occidentalis and I. iguana the removal of all known circa-
dian components (retinas, pineal, and parietal eye) does not
abolish circadian rhythms of locomotor activity [52,57],
demonstrating the existence of oscillators elsewhere in the
system. In A. carolinensis pinealectomy abolishes locomo-
tor rhythms in constant conditions, but the entrained phase
of these rhythms in pinealectomized individuals shows the
existence of oscillators elsewhere [58]. The data in P. sicula
and D. dorsalis suggest that such oscillators in S. occiden-
talis,I. iguana, and A. carolinensis may be contained within
the SCN.
8. Conclusions and perspectives
Reptiles because of their phylogenetic position and
ecology still provide the circadian field with some of the
most interesting model to understand circadian organiza-
tion, its evolution, and its variability. In lizards the pineal
complex, the retinas of the lateral eyes, and more recently
the SCN were all shown to participate in the control of
circadian rhythms. Noteworthy, the role these structures
play in circadian organization may vary interspecifically, as
it is the case of the pineal. However, the profound seasonal
differences in the behavioral effects of pinealectomy dis-
covered in P. sicula suggest that some of the interspecific
differences reported so far among lizards may, at least in
part depend on the particular season in which the beha-
vioral effects of pinealectomy have been examined in each
different species. Systematic studies of pinealectomized
lizards in different seasons across several species will
certainly solve the problem. Due to the central role of
the SCN in the circadian organization of two lizards (D.
dorsalis and P. sicula), similar studies should be extended
to further species of reptiles and to other aspects of
circadian behavior, such as, for instance, photic entrain-
ment. This will improve and extend phylogenetic compar-
isons among vertebrates concerning the circadian role of
the SCN. Again, several functional aspects of the redun-
dancy of circadian photoreceptors in reptiles (lateral eyes,
pineal, parietal eye, and deep brain photoreceptors) await
clarification. As regards deep brain photoreceptors, future
investigations in reptiles should be aimed at establishing
the precise location(s) of those brain photoreceptors that
effectively mediate photic entrainment, their characteriza-
Fig. 7. Effects of complete electrolytic lesions to the SCN (SCN-X) on
circadian locomotor rhythms in the Lacertid lizard P. sicula. Locomotor
activity records were double-plotted to aid in interpretation. (A) Record of a
lizard free-running in constant darkness (DD), which became arrhythmic in
response to SCN-X. (B) While intact, a lizard tested during the summer
entrained to the 24-h period of melatonin injections. After SCN-X this
lizard became behaviorally arrhythmic. Melatonin injections continued after
surgery, and their schedule was shifted on September 7th from 11:00 a.m. to
03:00 p.m. The SCN-X lizard remained arrhythmic during the whole
injection period (redrawn from Refs. [4,39]).
G. Tosini et al. / Physiology & Behavior 72 (2001) 461±471 469
tion at the molecular level, and the pathway(s) from them
to the rest of the circadian system.
Work in the A. Foa
Áand C. Bertolucci laboratory was
supported by grants from the Ministero dell'Universita' e
della Ricerca Scientifica. Work in the G. Tosini laboratory
was supported by grants from the National Institute of
Neurological Disorders and Stroke.
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... D'autres processus physiologiques présentant une rythmicité circadienne peuvent être étudiés, comme la température corporelle et le rythme de synthèse et sécrétion de la mélatonine, sécrétée la nuit et considérée comme un signal d'obscurité (Bhadra et al., 2017;Binkley, 1977;Tosini et al., 2001). Cette hormone est synthétisée et sécrétée par la glande pinéale de façon rythmique. ...
... Il est cependant à noter que la plupart des souris de laboratoires ne synthétisent pas de mélatonine (Kennaway, 2019). Chez les oiseaux (Chaurasia et al., 2005), certains reptiles (Tosini et al., 2001) et le poisson zèbre (Vatine et al., 2011), le rythme de synthèse de mélatonine persiste quand on place la glande pinéale en culture en condition d'obscurité constante, et au moins chez le poulet et le poisson zèbre elle est directement entrainable par des signaux extérieurs comme la lumière. Chez le poisson zèbre, la transcription du gène aanat2, l'enzyme limitante de la voie de synthèse de la mélatonine, est aussi utilisée comme reflet de la synthèse de la mélatonine chez la larve et l'adulte. ...
... Cependant dans ces conditions il est difficile de distinguer un rôle dans le photoentrainement d'un rôle dans le masking (effet direct de la lumière sur l'activité locomotrice). Les lézards S.olivaceus à la fois énuclés, pinéalectomisés et dont la lumière ne peut pas pénétrer dans le cerveau ont ainsi une activité locomotrice qui « free-run » en LD (Tosini et al., 2001). De même les lézards P.sicula à la fois pinéalectomisés, énucléés et qui ont reçu une injection d'un antisens dirigé contre l'opsine ps-RH2 exprimée dans l'hypothalamus antérieur ont une activité locomotrice qui « free-run » en LD pendant 6 à 7 jours après l'injection de l'antisens, alors que l'ablation de toutes les structures connues pour être photosensibles (oeil, glande pinéale, oeil pariétal) n'a pas d'effet sur le rythme de l'activité locomotrice en LD (Pasqualetti et al., 2003). ...
La lumière exerce une influence majeure sur la physiologie et le comportement de la plupart des organismes. En particulier, elle synchronise les rythmes circadiens par un processus appelé photoentrainement. Chez les mammifères, le photoentrainement dépend de la rétine, et plus particulièrement d'une classe de cellules ganglionnaires de la rétine (RGCs) exprimant le photopigment mélanopsine (opn4). Ces RGCs sont intrinsèquement sensibles à la lumière bleue et sont appelés ipRGCs pour "intrinsically photosensitives RGCs". Les ipRGCs intègrent l'information lumineuse médiée par la mélanopsine avec celle provenant des photorécepteurs classiques pour contrôler le photoentrainement mais aussi le masking, un effet direct de suppression ou d'élévation de l'activité locomotrice par la lumière. Bien que les ipRGCs soient les médiateurs des effets circadiens et directs de la lumière sur le comportement des mammifères, leurs rôles chez les vertébrés non mammifères ne sont pas élucidés. Grâce à son développement externe rapide, à sa transparence et à sa capacité de manipulation génétique facile et d'analyse comportementale à haut débit, le poisson zèbre (PZ) est apparu comme un puissant modèle de vertébré diurne non mammifère pour la chronobiologie. Contrairement aux mammifères, la rétine n'est pas la seule structure photosensible chez le PZ. En particulier chez cette espèce, la glande pinéale contient des photorécepteurs et des neurones de projection (PNs, équivalent des RGCs) et peut donc recevoir une information lumineuse et la transmettre au cerveau. De plus, tous les organes adultes du PZ testés, y compris la glande pinéale, sont directement photoentraînables lorsqu'ils sont placés en culture. Il reste donc à déterminer quels sont les rôles respectifs de la photodétection périphérique (au niveau de tous les organes) et de la photodétection centrale (au niveau de structures spécialisées telles que la rétine et la glande pinéale) dans le photoentraînement. Nous nous sommes intéressé.e.s aux rôles des gènes de mélanopsine et des cellules les exprimant chez le PZ. Parmi les 5 gènes de mélanopsine présents chez cette espèce, opn4xa et opn4b sont exprimés dans les RGCs larvaires. De plus, nous avons montré qu'opn4xa est exprimé dans une sous-population de PNs. Ainsi, opn4xa définit une nouvelle population de PNs présentant une photosensibilité à la lumière bleue et verte dépendante d'opn4xa et qui fonctionne en mode LIGHT ON (Publications 1,2). Dans un second temps, nous avons cherché à comprendre la fonction des RGCs (opn4xa+ et opn4xa- - mutant lakritz -) et de la photosensibilité intrinsèque des RGCs et PN opn4xa+ (mutant opn4xa) dans l'influence directe et circadienne de la lumière sur l'activité locomotrice chez la larve de PZ. Nous avons trouvé que les RGCs sont impliqués dans le masking et la réponse locomotrice aux transitions lumineuses, mais indépendamment de la photosensibilité pilotée par opn4xa. Enfin, les mutants opn4xa-/-, lakritz-/- et opn4xa -/- ; lakritz -/- n'ont montré aucun défaut de photoentraînement à un pulse de lumière blanche administré au début de la nuit suggérant que la photodétection de la rétine (y compris celle des ipRGCs) et la photodétection pilotée par la mélanopsine opn4xa ne sont pas nécessaires au photoentraînement chez la larve de PZ (Publication 2). Ces résultats soulèvent des différences majeures dans le photoentrainement circadien chez les mammifères et le poisson zèbre.
... Reptiles have discrete times of the day and year when activity is concentrated (Tosini, Bertolucci and Foà, 2001;Dayananda, Jeffree and Webb, 2020), yet movement and activity are not always predictable. Movement, basking, nesting and brumation behaviours may be influenced by biotic (e.g., prey availability, reproductive condition, experience of individuals) and physical factors (e.g., climate, the attributes of the landscape, topography (Meeske, 2000;Meeske and Mühlenberg, 2004;Ficetola, Thuiller and Padoa-Schioppa, 2009;Vilardell-Bartino et al., 2015;Ottonello et al., 2017a;Canessa et al., 2016)). ...
Behavioural and spatial distribution analyses were quantified during a phase of activity and lethargy in a wild population of the European pond turtle inhabiting a protected internal wetland area of the Venice lagoon. The marked individuals (13 males and 16 females) provided informative radiotracking data to study differential patterns of activity, dispersion and habitat use between the two study periods ("October-November both 2019 and 2020" and "June-July 2020"). The differences in the movements behaviours and habitat selection were affected by period. Movements were higher in the period of activity than lethargy, but they were not influenced by sex and size. The presence of the European pond turtle in the transitional woodland/shrubs and brackish water valley habitats was significantly higher in the period of activity than lethargy. During the latter one, pond turtles were observed to brumate gregariously in a small area for brumation, usually in shallow water. In contrast, all individuals have changed water bodies during the activity period. Part of those movements has occurred towards aquatic habitat with higher salinities 1-17 (mean: 10.64). These findings provide a set of information to better understand the behavioural ecology of Emys orbicularis in the lagoon area. This is of relevance for management actions and for the conservation of this threatened species.
... Some circadian clocks in the other vertebrate classes such as birds, reptiles, fish, or amphibians have direct co-localization between light-sensing proteins and the clock (e.g., chicken pinealocytes (Deguchi 1981)). Other circadian clocks in nonmammalian vertebrates are not directly photosensitive, but receive light input from non-retinal (deep brain) photoreceptors (e.g., lizard SCN (Underwood 1973, Tosini et al. 2001). Mammals do not have evidence of deep brain photoreceptors (Underwood and Groos 1982); direct exposure of the brain to light in enucleated rodents does not evoke circadian responses to light (Groos and van der Kooy 1981). ...
Intrinsically photosensitive retinal ganglion cells (ipRGGs) are the most recently discovered photoreceptor class in the human retina. This Element integrates new knowledge and perspectives from visual neuroscience, psychology, sleep science and architecture to discuss how melanopsin-mediated ipRGC functions can be measured and their circuits manipulated. It reveals contemporary and emerging lighting technologies as powerful tools to set mind, brain and behaviour.
... Rhythmic activity has been also reported in reptiles, along with per2, cry1, and clock expression in both central and peripheral oscillators (Bertolucci et al. 2017). However, it is noteworthy that 24 h periods of both light (photoperiod) and temperature (thermoperiod) can impact per2 and pineal melatonin rhythms in many species of reptiles, resulting in seasonal variation of rhythmicity, suggesting differences in clock organization between reptiles and mammals (Tosini et al. 2001;Bertolucci et al. 2017). ...
Over the past decades the molecular mechanisms responsible for circadian phenotypes of animals have been studied in increasing detail in mammals, some insects, and other invertebrates. Particular circadian proteins and their interactions are shared across evolutionary distant animals resulting in a hypothesis for the canonical circadian clock of animals. As the number of species for which the circadian clockwork has been described increases, the circadian clock in animals driving cyclical phenotypes becomes less similar. Our focus in this review is to develop and synthesize the current literature to better understand the antiquity and evolution of the animal circadian clockwork. Here, we provide an updated understanding of circadian clock evolution in animals, largely through the lens of conserved genes characterized in the circadian clock identified in bilaterian species. These comparisons reveal extensive variation within the likely composition of the core clock mechanism, including losses of many genes, and that the ancestral clock of animals does not equate to the bilaterian clock. Despite the loss of these core genes, these species retain circadian behaviors and physiology suggesting novel clocks have evolved repeatedly. Additionally, we highlight highly conserved cellular processes (e.g., cell division, nutrition) that intersect with the circadian clock. The conservation of these processes throughout the animal tree remains essentially unknown, but understanding their role in the evolution and maintenance of the circadian clock will provide important areas for future study.
... 35 The exposure of shorter blue wavelengths on the retinal pigment epithelium can danger photoreceptors via photochemical damage. 36,37 On the other side, emission wavelengths greater than 475 nm sacrifice color saturation and result in poor image quality. As a result, the lighting industry is moving toward the spectral window of 460−475 nm, and in this study, we employed blue LEDs with a wavelength of 460 nm. ...
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In the next decade, we will witness the replacement of a majority of conventional light sources with light-emitting diodes (LEDs). Efficient LEDs other than phosphors can enhance their functionality and meet different lighting needs. Quantum dots (QDs) have high potential for future LED technology due to their sensitive band-gap tuning via the quantum confinement effect and compositional control, high photoluminescence quantum yield (PLQY), and mass-production capacity. Herein, we demonstrate white LEDs using QDs that reach over 150 lumens per electrical Watt. For that we synthesized green- and red-emitting ZnCdSe/ZnSe core/shell QDs by low-temperature nucleation, high-temperature shell formation, and postsynthetic trap-state removal. Their cadmium concentration is lower than 100 ppm, satisfying the current EU RoHS regulations, and their PLQY reaches a high level of 94%. The PLQY of QDs is maintained within the device on blue LED via liquid injection, and their integration at optimized optical densities leads to 129.6 and 170.4 lm/W for red-green-blue (RGB)- and green-blue (GB)-based white LEDs, respectively. Our simulations further showed that an efficiency level of over 230 lm/W is achievable using ultraefficient blue LED pumps. The simple fabrication and high performance of white LEDs using QD liquids show high promise for next-generation lighting devices.
Captive animal welfare has benefited from various new technologies and a new generation of welfare-minded and better-informed individuals adopting more welfare-oriented practices. However, for captive reptiles, there remain many aspects that are grounded in and reflect a long history of arbitrary or folklore husbandry and advice, and reptile-keeping continues to be compromised by practices that benefit the keeper rather than the animal that is kept. This second edition of Health and Welfare of Captive Reptiles, like the first volume, contains a diversity of primary classical subjects, each hopefully constituting an advancement in our understanding of reptilian biology and meeting the associated needs of these animals in captivity. Some subjects, comprise miscellaneous considerations that, directly or indirectly, will have a significant bearing on reptile health and welfare. It is these factors that form the basis of this chapter. It is hoped that, at the very least, their inclusion may create or stimulate an awareness of other potential issues that may affect the well-being of captive reptiles.KeywordsAnimal welfareReptile husbandryStressPainSensitivityEnvironmentEuthanasiaKillingEthics
Sleep, accounting for roughly one-third of a person’s life, plays an important role in human health. Despite the close association between sleep patterns and medical diseases proven by several studies, it has been neglected in recent years. Presently, all societies are facing the most challenging health-threatening disease, cancer. Among all cancer types, gastrointestinal (GI) cancers, especially colorectal type, seem to be one of the most relevant to an individual’s lifestyle; thus, they can be prevented by modifying behaviors most of the time. Previous studies have shown that disruption of the 24-h sleep–wake cycle increases the chance of colorectal cancer, which can be due to exposure to artificial light at night and some complex genetic and hormone-mediated mechanisms. There has also been some evidence showing the possible associations between other aspects of sleep such as sleep duration or some sleep disorders and GI cancer risk. This review brings some information together and presents a detailed discussion of the possible role of sleep patterns in GI malignancy initiation.
Subfertility and recurrent miscarriage are present within 6% and 2% of the female population respectively1. These pathologies can bring psychological harm to individuals and families2, however the biological mechanisms that underpin these issues are not fully understood, with up to 50% of the cases lacking an etiology3 Understanding the biochemical environment inside the uterus allows clinicians greater insight into pathologies arising from dysregulation at a cellular level. Current techniques for monitoring these biochemical changes focus on point measurements, such as phlebotomy or invasive draining procedures, which can lack the information that can be garnered by measuring the changes over the course of minutes or hours. Microdialysis has proven to be a technique that can monitor changes with high spatial resolution by bringing a semipermeable membrane in to close contact with the area of interest. Despite its success within the measurement of rapidly changing biomarkers, such as neurotransmitters, the application of microdialysis to long term monitoring is hampered by technical limitations brought about by the inability to preserve temporal resolution whilst allowing patients relative ease of movement. Here, a microdialysis probe with embedded sample storage is developed to overcome these limitations. The use of microfluidics, specifically that of plug flow and capillary droplet traps, allowed for16nL samples to be stored within the body of the probe in addressable arrays, paving the way for further analysis. The integration of semipermeable membranes to the body of these microfabricated probes resulted in novel bonding techniques that allowed for the bonding of Polyethersulfone, a thermally stable and solvent resistant, semipermeable membrane to SU-8, a common epoxy-based photoresist. The resulting microdialysis probes were validated against commercially available probes and showed viability in recovering biomarkers. Finally, a proof-of-concept experiment was undertaken that demonstrated the possibility of carrying out assays on the chip.
Compared to other domestic species, relatively little is known about the reptile eye and its role in health and disease. Until the 1980s, ocular disease was considered rare in reptiles (Millichamp et al. 1983; Ensley et al. 1978). However, with an increase in routine ophthalmic examination and research, we have come to realize that the reptile eye, similar to other domestic species, can be affected primarily or secondary to systemic disease or poor husbandry. When considering a body size range of 3 inches to 10 feet as well as variations in morphology, physiology, and behavioral adaptations, lizards are arguably the most diverse group of reptiles (Barten and Simpson 2019a; O’Malley 2005). Over half the known reptile species are lizards and can be found on every continent but Antarctica. Many lizard species can be kept in captivity, raising the need for knowledge of disease and its management.
Iguanas are the most endangered family of reptiles, with 77% categorised as threatened or near threatened. Further, Cyclura is the most endangered reptile genus, with all 12 species considered threatened. Therefore, it is vital that we develop assisted reproductive technologies for Cyclura spp. to enhance their conservation efforts. The goals of this study were to collect semen, and to measure testicle size and testosterone concentrations in Grand Cayman rock iguana hybrids (Cyclura lewisi×nubila (CLN)) and rhinoceros rock iguanas (Cyclura cornuta (CC)). A prospective longitudinal study was performed in 9.0 CLN and 9.0 CC during their reproductive season in southern Florida (February-July). Serial testicle ultrasound measurements and plasma testosterone concentrations were collected monthly. Testicle measurements (length (L), width (W), height (H)) were collected and testicle volume (V) was estimated using the equation V=0.52(LW2). There were significant differences in testicle L, W, H and V for both species. Testicle size peaked for CLN and CC in April and May respectively. Plasma testosterone concentrations increased from baseline during February, March and April in CLN and in March, April and May in CC. Ultrasound testicle measurements could be used to predict when to collect semen in these seasonally monocyclic iguanas.
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In order to contribute to a comparative view on lacertids, the effect of pinealectomy on the freerunning activity displayed under constant darkness and temperature (27.5°C ± 0.5) has been studied in the lizard Gallotia galloti eisentrauti. Animals showed an entrained motor activity rhythm under an initial light-dark (12:12 hours) routine and freerunning circadian periods ranging between 24.1 and 25.5 h during constant darkness (periodograms obtained by Sokolove & Bushell's method). After pinealectomy, most animals showed no significant circadian rhythm, their locomotor activity becoming diffuse throughout the whole 24 h period. Thus, the pineal gland seems to play an important role as a main pacemaker regulating the endogenous activity rhythm under constant conditions. This result contrasts with that found in Podarcis sicula where after pinealectomy only changes in length of the freerunning period were found.
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The pineal complex of reptiles is a morphologically and functionally con-nected set of organs that originates as an evagination of the roof of the dien-cephalon. It is formed by two structures: the pineal organ and the parietal eye. The pineal gland is present in Chelonia, Squamata and Rhynchocephalia, but not in Crocodilia. The parietal eye is present in some species of lizards (Squama-ta) and in the tuatara (Rhynchocephalia). Both the pineal gland and the parietal eye are photosensitive. In particular, the parietal eye is an highly organized pho-toreceptive structure, with a well-defined lens, cornea and retina. The most important (and studied) secretory product of this complex is the hormone melat-onin which is synthesized by both organs (pineal and parietal eye). The pineal organ is believed to be the neuroendocrine transducer of changes in photoperiod and environmental temperature and it has been demonstrated to have a func-tional role in many aspects of reptilian biology. Melatonin has an influence on the mechanisms controlling thermoregulation (behavioral and physiological), because its manipulation or removal may produce significant alterations of behavioral and physiological thermoregulatory parameters. The reptilian pineal complex may also possess self-sustained circadian oscillators which are involved in the circadian organization of these animals and in their reproduction. It is believed that many of the roles played by the pineal complex are mediated by the hormone melatonin, since exogenous administration of melatonin may affect the animal's physiology and/or behavior. The present paper will review the cur-rent knowledge about the neuroendocrinology and functional roles of the reptil-ian pineal complex.
The circadian rhythmicity of eukaroytic organisms is dictated by an innate program that specifies the time course through the day of many aspects of metabolism and behavior. The programmed sequence of events in each cycle of the rhythm has been evolved to parallel the sequence of predictable change (physical and biological) in the course of the day-outside: it constitutes an appropriate day-within. It is a characteristic, almost defining, feature of these circadian programs that their time course is stabilized with almost clocklike precision to parallel the stable time course of the environmental day. There is equally clear functional significance to the program's being driven by a self-sustaining oscillator; thus, the program is subject to entrainment by one or more of the external cycles whose period it closely approximates. It is this entrain ability that provides for proper phasing of the program to the sequence of external changes that it has been evolved to cope with and exploit.
It is only within the last 15 years that a few favorable experimental situations have been identified that have made it possible to study the physiology of circadian systems in some vertebrates (Aschoff 1981). Given the complexity of the systems under study, the long time constants of experimentation, and the relatively small number of scientists engaged in the work, it is not surprising that I must address the search for principles rather than the principles themselves. Insofar as documented principles exist, they are disturbingly vague. However, in discussing them it becomes clear that at least we know where we should be looking further. A few principles, proto-principles and pseudoprinciples are discussed below.
1. 1.|The temperature preference (ST) of Chrysemys scripta elegans was investigated under normal and inverted light regimes (NLR and ILR, 28 experiments) and during continuous light (CLR, 5 experiments). Each experiment lasted 72 h. 2. 2.|In 26 experiments with NLR and ILR, single cosinor analysis yields a significant diel rhythm. The amplitude obtained by the mean cosinor procedure is always significant. 3. 3.|The diel rhythm of ST is based on an endogenous circadian rhythm. 4. 4.|The main 'zeitgeber' for the diel rhythm is protoperiod.
1.1.|Electrolytic lesions were placed stereotaxically in the medial preoptic area, hypothalamus, and in the telencephalon to determine their functions in behavioural thermoregulation in the desert iguana, Dipsosaurus dorsalis.2.2.|Lesioned and sham-operated control lizards were placed on the hot side of the thermal shuttle box, as described in the preceding paper.3.3.|Six characteristics of behavioural thermoregulation were analyzed for each lesioned and sham-operated control lizard in the present study and for each unoperated control lizard from the previous paper. These characteristics are the mean high body temperature at shuttling (HBTS), the mean low body temperature at shuttling (LBTS), the standard deviations of the HBTS and LBTS, the frequency of shuttling, and the body temperatures of lizards taking up special thermal postures. The level of significant difference between a characteristic of a lesioned lizard and the unlesioned groups was accepted at p ≤ 0·05.4.4.|Medial preoptic area lesions dramatically alter thermoregulation. The LBTS was significantly decreased in all seven lizards. The HBTS was also significantly increased in two lizards. The standard deviations of the HBTS and LBTS in some lizards were also significantly large, while the frequency of shuttling is significantly reduced.5.5.|Lesions of the nucleus of the anterior hypothalamus and nucleus suprachiasmaticus, but not the anterior hypothalamic area, resulted in a significant reduction in both the frequency of shuttling and the LBTS, and the standard deviation of the LBTS was significantly reduced in one of these lizards.6.6.|Lesions in the neural structures of the rostral-dorsal hypothalamus and the nucleus of the ventromedial hypothalamus had no effects on thermoregulatory performance. However, a bilateral lesion of the nucleus paraventricularis resulted in a very low standard deviation of the LBTS.7.7.|Large lesions extending from the posterior part of the ventromedial telencephalon to the medial forebrain bundle and large lesions of the medial wall of the telencephalon (medial cortex, septal area) resulted in significantly low HBTS's and LBTS's. The standard deviations of these two temperatures were exceedingly large.8.8.|The medial preoptic area in ectothermic lizards plays an essential role in the neural integration of behavioural thermoregulation, as it does in both physiological and behavioural control of temperatures in the endothermic birds and mammals. The high and low temperature set-point mechanisms, responsible for shuttling behaviour, act independently of each other. Other rostral hypothalamic nuclei and telencephalic structures contribute to the central neural network involved in thermoregulation.
SUMMARY 1. At a constant temperature of 24 °C there was a diel fluctuation in plasma melatonin concentration; highest levels occurring in the scotophase of a reversed daily light-cycle. Parietalectomy did not appear to affect melatonin titres under these conditions. 2. When lizards were subjected to a photoperiod together with a thermo- period (31 °C in the photophase, 24 °C in the scotophase), nocturnal plasma melatonin levels were almost twice as high as those in animals subjected to a photoperiod at constant temperature. 3. Capping the lateral eyes of T. rugosus under these conditions did not alter the phase or amplitude of the rhythm in plasma melatonin content. However, removal of the parietal eye abolished the rhythm, owing mainly to reduced levels during the mid-scotophase and elevated levels during the mid- photophase. 4. It is concluded that plasma melatonin levels are regulated extraretinally, and that the parietal eye may help to mediate environmental input to centres secreting melatonin. It is suggested that the parietal eye may mediate thermal as well as photic information.
Body temperatures of individually housed Sceloporus occidentalis were monitored continuously while the animals moved freely over a thermal gradient during several daily light-dark cycles and a subsequent period of constant darkness. These self-selected body temperatures display a distinct daily pattern, a pattern which is maintained (with an altered cycle length) during constant darkness.
Pinealectomy of the iguanid lizardSceloporus occidentalis freerunning in either continuous illumination or continuous darkness typically causes changes in the period of the activity rhythm as well as changes in the amount of daily activity (). Blinding also alters the period of the freerunning activity rhythm. Continuous long term administration of melatonin via subcutaneous capsules causes a significant lengthening of the period of the activity rhythm (as well as a decrease in ) of pinealectomized and/or blinded lizards showing that melatonin exerts its action at extrapineal and extraocular sites. However, the amount of lengthening induced by melatonin is significantly greater in pinealectomized lizards than in intact lizards. The results indicate that the pineal (and possibly the eyes) act as coupling devices or as the loci of circadian pacemakers within a multioscillator system. Melatonin may function as a chemical messenger between the pineal (or eyes) and the rest of the circadian system.