Locomotor Activity Rhythms in Cave Catfishes, Genus Taunayia, from Eastern Brazil (Teleostei: Siluriformes: Heptapterinae)

We studied the locomotor rhythmicity in heptapterine catfishes, genus Taunayia, under free-running conditions (DD) and LD cycles (12:12). Taunayia sp., anophthalmic and depigmented undescribed species from a cave in northeastern Brazil, is the fourth Brazilian troglobitic catfish studied with focus on circadian rhythms. Weak free-running rhythmicity, with absence of significant circadian components, was observed for this species when compared to the epigean, eyed relatives. On the other hand, the studied troglobitic catfishes in general presented significant circadian rhythms under LD cycles, with activity peaks in the night phase probably corresponding to nocturnal activity pattern inherited from their epigean ancestors. However, no residual oscillations were observed after transition from LD to DD. This indicates masking of activity by light-dark cycles. Regression of circadian rhythmicity in the stable, permanently dark subterranean habitat was also observed for other cave fishes. Such regression corroborates the notion that circadian rhythmicity is mainly selected in the epigean environment by ecological factors, namely daily cycles of light and/or temperature.     

Locomotor Activity Rhythms in Cave Catfishes,Genus Taunayia,from Eastern Brazil (Teleostei: Siluriformes: Heptapterinae)

We studied the locomotor rhythmicity in heptapterine catfishes, genus Taunayia, under free-running conditions (DD) and LD cycles (12:12). Taunayia sp., anophthalmic and depigmented undescribed species from a cave in northeastern Brazil, is the fourth Brazilian troglobitic catfish studied with focus on circadian rhythms. Weak free-running rhythmicity, with absence of significant circadian components, was observed for this species when compared to the epigean, eyed relatives. On the other hand, the studied troglobitic catfishes in general presented significant circadian rhythms under LD cycles, with activity peaks in the night phase probably corresponding to nocturnal activity pattern inherited from their epigean ancestors. However, no residual oscillations were observed after transition from LD to DD. This indicates masking of activity by light-dark cycles. Regression of circadian rhythmicity in the stable, permanently dark subterranean habitat was also observed for other cave fishes. Such regression corroborates the notion that circadian rhythmicity is mainly selected in the epigean environment by ecological factors, namely daily cycles of light and/or temperature.     

Stocking density affects circadian rhythms of locomotor activity in African catfish, Clarias gariepinus

The effect of stocking density on the locomotor activity of African catfish C. gariepinus under different light regimes was investigated. C. gariepinus were stocked under different densities (1, 5, or 10 fish/tank), and their locomotor activity recorded under light-dark (LD), constant light (LL), constant darkness (DD), and LD-reversed (DL) regimens. Under the LD cycle, catfish showed a crepuscular activity pattern, irrespective of stocking density, with most of the daily activity concentrated around the light-onset and light-offset times. When fish were subjected to DD, all 4 tanks with medium (5 fish) and high (10 fish) stocking densities showed circadian rhythmicity, with an average period (?) of 23.3???0.5 and 24.6???0.5?h, respectively. In contrast, only 2 low (1 fish) density tanks showed free-running rhythms. Under LL, activity levels decreased significantly in comparison with levels observed under LD and DD. Moreover, fish of 1, 2, and 3 out of the 4 tanks with low, medium, and high densities, respectively, showed free-running rhythms under these conditions. When the photocycle was reversed (DL), fish of 3, 2, and 4 out of the 4 tanks with low, medium, and high stocking densities, respectively, showed gradual resynchronization to the new phase, and transient cycles of activity were observed. These results suggest that stocking density of fish affected the display of circadian rhythmicity and the intensity of activity levels. Thus, fish kept in higher densities showed more robust rhythmicity and higher levels of daily activity, indicating that social interactions may have an influence on behavioral patterns in the African catfish.     

Circadian rhythms of heart rate in freely moving and restrained American lobsters, Homarus americanus

While circadian rhythms of locomotion have been reported in the American lobster, Homarus americanus, it is unclear whether heart rate is also modulated on a circadian basis. To address this issue, both heart rate and locomotor activity were continuously monitored in light-dark (LD) cycles and constant darkness (DD). Lobsters in running wheels exhibited significant nocturnal increases in locomotor activity and heart rates during LD, and these measures were significantly correlated. In DD, most lobsters exhibited persistent circadian rhythms of both locomotion and heart rate. When heart rate was monitored in restrained lobsters in LD and DD, most animals also demonstrated clear daily and circadian rhythms in heart rate. Overall, this is the first demonstration of circadian rhythms of heart rate in H. americanus, the expression of which does not appear to be dependent on the expression of locomotor activity.     

Circadian Rhythms of Locomotor Activity in the Nile Tilapia Oreochromis niloticus

Behavioral rhythms of the Nile tilapia were investigated to better characterize its circadian system. To do so, the locomotor activity patterns of both male and female tilapia reared under a 12:12 h light-dark (LD) cycle were studied, as well as in males the existence of endogenous rhythmicity under free-running conditions (DD and 45 min LD pulses). When exposed to an LD cycle, the daily pattern of activity differed between individuals: some fish were diurnal, some nocturnal, and a few displayed an arrhythmic pattern. This variability would be typical of the plastic circadian system of fish. Moreover, reproductive events clearly affected the behavioral rhythms of female tilapia, a mouth-brooder teleost species. Under DD, 50% (6 of 12) of male fish showed circadian rhythms with an average period (τ) of 24.1±0.2 h, whereas under the 45 min LD pulses, 58% (7 of 12) of the fish exhibited free-running activity rhythms with an average τ of 23.9±0.5 h. However, interestingly in this case, activity was always confined to the dark phase. Furthermore, when the LD cycle was reversed, a third of the fish showed gradual resynchronization to the new phase, taking 7–10 days to be completely re-entrained. Taken together, these results suggest the existence of an endogenous circadian oscillator that controls the expression of locomotor activity rhythms in the Nile tilapia, although its anatomical localization remains unknown.     

Interruption of the rat circadian clock by short light-dark cycles

Ninety male Sprague-Dawley rats were exposed to 1:1-h light-dark (LD1:1) cycles for 50-90 days, and then they were released into constant darkness (DD). During LD1:1 cycles, behavioral rhythms were gradually disintegrated, and circadian rhythms of locomotor activity, drinking, and urine 6-sulfatoxymelatonin excretion were eventually abolished. After release into DD, 44 (49%) rats showed arrhythmic behavior for >10 days. Seven (8%) animals that remained arrhythmic for >50 days in DD were exposed to brief light pulses or 12:12-h light-dark cycles, and then they restored their circadian rhythms. These results indicate that the circadian clock was stopped, at least functionally, by LD1:1 cycles and was restarted by subsequent light stimulation.     

Light regulates the cell cycle in zebrafish

The timing of cell proliferation is a key factor contributing to the regulation of normal growth. Daily rhythms of cell cycle progression have been documented in a wide range of organisms. However, little is known about how environmental, humoral, and cell-autonomous factors contribute to these rhythms. Here, we demonstrate that light plays a key role in cell cycle regulation in the zebrafish. Exposure of larvae to light-dark (LD) cycles causes a range of different cell types to enter S phase predominantly at the end of the day. When larvae are raised in constant darkness (DD), a low level of arrhythmic S phase is observed. In addition, light-entrained cell cycle rhythms persist for several days after transfer to DD, both observations pointing to the involvement of the circadian clock. We show that the number of LD cycles experienced is essential for establishing this rhythm during larval development. Furthermore, we reveal that the same phenomenon exists in a zebrafish cell line. This represents the first example of a vertebrate cell culture system where circadian rhythms of the cell cycle are observed. Thus, we implicate the cell-autonomous circadian clock in the regulation of the vertebrate cell cycle by light.     

Circadian rhythms of heart rate in freely moving and restrained American lobsters,Homarus americanus

While circadian rhythms of locomotion have been reported in the American lobster, Homarus americanus, it is unclear whether heart rate is also modulated on a circadian basis. To address this issue, both heart rate and locomotor activity were continuously monitored in light-dark (LD) cycles and constant darkness (DD). Lobsters in running wheels exhibited significant nocturnal increases in locomotor activity and heart rates during LD, and these measures were significantly correlated. In DD, most lobsters exhibited persistent circadian rhythms of both locomotion and heart rate. When heart rate was monitored in restrained lobsters in LD and DD, most animals also demonstrated clear daily and circadian rhythms in heart rate. Overall, this is the first demonstration of circadian rhythms of heart rate in H. americanus, the expression of which does not appear to be dependent on the expression of locomotor activity.     

Period and phase control in a multioscillatory circadian system (Iguana iguana)

The circadian system of the lizard Iguana iguana is composed of several independent pacemakers that work in concert: the pineal gland, retinae of the lateral eyes, and a fourth oscillator presumed to be located in the hypothalamus. These pacemakers govern the circadian expression of multiple behaviors and physiological processes, including rhythms in locomotor activity, endogenous body temperature, electroretinogram, and melatonin synthesis. The numerous, easily measurable rhythmic outputs make the iguana an ideal organism for examining the contributions of individual oscillators and their interactions in governing the expression of overt circadian rhythms. The authors have examined the effects of pinealectomy and enucleation on the endogenous body temperature rhythm (BTR) and locomotor activity rhythm (LAR) of juvenile iguanas at constant temperature both in LD cycles and in constant darkness (DD). They measured the periods (tau) of the circadian rhythms of LAR and BTR, the phase relationships between them in DD (psiAT), and the phase relationship between each rhythm and the light cycle (psiRL). Pinealectomy lengthened tau of locomotor activity in all animals tested and abolished the BTR in two-thirds of the animals. In those animals in which the BTR did persist following pinealectomy, tau lengthened to the same extent as that of locomotor activity. Pinealectomy also delayed the onset of activity with respect to its normal phase relationship with body temperature in DD. Enucleation alone had no significant effect on tau of LAR or BTR; however, after enucleation, BTR became 180 degrees out of phase from LAR in DD. After both pinealectomy and enucleation, 4 of 16 animals became arrhythmic in both activity and body temperature. Their data suggest that rhythmicity, period, and phase of overt circadian behaviors are regulated through the combined output of multiple endogenous circadian oscillators.     

Light-dependent development of circadian gene expression in transgenic zebrafish

The roles of environmental stimuli in initiation and synchronization of circadian oscillation during development appear to vary among different rhythmic processes. In zebrafish, a variety of rhythms emerge in larvae only after exposure to light-dark (LD) cycles, whereas zebrafish period3 (per3) mRNA has been reported to be rhythmic from day 1 of development in constant conditions. We generated transgenic zebrafish in which expression of the firefly luciferase (luc) gene is driven by the zebrafish per3 promoter. Live larvae from these lines are rhythmically bioluminescent, providing the first vertebrate system for high-throughput measurement of circadian gene expression in vivo. Circadian rhythmicity in constant conditions was observed only after 5–6 d of development, and only if the fish were exposed to LD signals after day 4. Regardless of light exposure, a novel developmental profile was observed, with low expression during the first few days and a rapid increase when active swimming begins. Ambient temperature affected the developmental profile and overall levels of per3 and luc mRNA, as well as the critical days in which LD cycles were needed for robust bioluminescence rhythms. In summary, per3-luc zebrafish has revealed complex interactions among developmental events, light, and temperature in the expression of a clock gene.     

Role of suprachiasmatic nuclei in circadian and light-entrained behavioral rhythms of lizards

To establish whether the suprachiasmatic nuclei (SCN) of the Ruin lizard (Podarcis sicula) play a role in entrainment of circadian rhythms to light, we examined the effects of exposure to 24-h light-dark (LD) cycles on the locomotor behavior of lizards with SCN lesions. Lizards became arrhythmic in response to complete SCN lesion under constant temperature and constant darkness (DD), and they remained arrhythmic after exposure to LD cycles. Remnants of SCN tissue in other lesioned lizards were sufficient to warrant entrainment to LD cycles. Hence, the SCN of Ruin lizards are essential both to maintain locomotor rhythmicity and to mediate entrainment of these rhythms to light. We also asked whether light causes expression of Fos-like immunoreactivity (Fos-LI) in the SCN. Under LD cycles, the SCN express a daily rhythm in Fos-LI. Because Fos-LI is undetectable in DD, the rhythm seen in LD cycles is caused by light. We further showed that unilateral SCN lesions in DD induce dramatic period changes. Altogether, the present data support the existence of a strong functional similarity between the SCN of lizards and the SCN of mammals.     

Inconsistency in the expression of locomotor and ERG circadian rhythms in the German cockroach, Blattella germanica (L.).

ERG recordings from German cockroaches showed that the amplitude of light-evoked responses have a circadian rhythmicity in adult males that coincided with the locomotor circadian rhythm. The peak of the response occurred during the subjective night, and the circadian period was less than 24 h under DD condition. In contrast, although the locomotor circadian rhythm was masked by the development of ovaries and pregnancy in females, their visual responses displayed circadian rhythmicity. This inconsistency in expression of locomotor and visual sensitivity circadian rhythms in females implied separate pacemakers for these two overt rhythms. After severing the optic nerves, changes in ERG amplitude of the operated cockroaches still displayed a circadian rhythm under DD condition, demonstrating that the visual sensitive pacemaker was located in the eye and independent from the locomotor pacemaker.     

Behavioral and molecular analyses suggest that circadian output is disrupted by disconnected mutants in D. melanogaster.

Mutations in the disconnected (disco) gene act to disrupt neural cell patterning in the Drosophila visual system. These mutations also affect adult locomotor activity rhythms, as disco flies are arrhythmic under conditions of constant darkness (DD). To determine the state of the circadian pacemaker in disco mutants, we constructed with pers double mutants (a short period allele of the period gene) and assayed their behavioral rhythms in light-dark cycles (LD), and their biochemical rhythms of period gene expression under both LD and DD conditions. The results demonstrate that disco flies are rhythmic, indicating that they have an active circadian pacemaker that can be entrained by light. They also suggest that disco mutants block or interfere with elements of the circadian system located between the central pacemaker and its outputs that mediate overt rhythms.     

Stocking Density Affects Circadian Rhythms of Locomotor Activity in African Catfish,Clarias gariepinus

The effect of stocking density on the locomotor activity of African catfish C. gariepinus under different light regimes was investigated. C. gariepinus were stocked under different densities (1, 5, or 10 fish/tank), and their locomotor activity recorded under light-dark (LD), constant light (LL), constant darkness (DD), and LD-reversed (DL) regimens. Under the LD cycle, catfish showed a crepuscular activity pattern, irrespective of stocking density, with most of the daily activity concentrated around the light-onset and light-offset times. When fish were subjected to DD, all 4 tanks with medium (5 fish) and high (10 fish) stocking densities showed circadian rhythmicity, with an average period (τ) of 23.3?±?0.5 and 24.6?±?0.5?h, respectively. In contrast, only 2 low (1 fish) density tanks showed free-running rhythms. Under LL, activity levels decreased significantly in comparison with levels observed under LD and DD. Moreover, fish of 1, 2, and 3 out of the 4 tanks with low, medium, and high densities, respectively, showed free-running rhythms under these conditions. When the photocycle was reversed (DL), fish of 3, 2, and 4 out of the 4 tanks with low, medium, and high stocking densities, respectively, showed gradual resynchronization to the new phase, and transient cycles of activity were observed. These results suggest that stocking density of fish affected the display of circadian rhythmicity and the intensity of activity levels. Thus, fish kept in higher densities showed more robust rhythmicity and higher levels of daily activity, indicating that social interactions may have an influence on behavioral patterns in the African catfish. (Author correspondence: lmvera@um.es)     

Feeding rhythms in constant light and constant darkness: the role of the eyes and the effect of melatonin infusion

Exposure to constant light abolishes circadian behavioral rhythms of locomotion and feeding as well as circulating melatonin rhythms in pigeons (Columba livia). To determine if feeding rhythmicity could be maintained in pigeons exposed to constant light, periodic infusions (10h/day) of melatonin were administered to pinealectomized and bilaterally retinectomized/pinealectomized pigeons under conditions of both constant darkness and constant light. The infusions were sufficient to entrain rhythmicity in pinealectomized pigeons in constant darkness and to restore and maintain rhythmicity in bilaterally retinectomized/pinealectomized pigeons in constant darkness. On subsequent exposure to constant light, rhythmicity remained phase locked to the melatonin infusions in bilaterally retinectomized/pinealectomized pigeons but was abolished in sighted pinealectomized birds. These results suggest that while endogenous melatonin rhythms are both necessary and sufficient to maintain behavioral rhythms in DD, their effect can be overridden by constant light but only if perceived by the eyes. Thus, constant light may abolish behavioral rhythmicity in intact pigeons (and perhaps in other species) by a mechanism other than suppression of endogenous melatonin rhythmicity. Such a mechanism might involve direct stimulation of locomotor or feeding activity by retinally perceived (but not by extra-retinally perceived) light, or alternatively by suppression of a hypothalamic oscillator that receives its major light input from the retinae.Abbreviations PX pinealectomized - EX bilaterally enucleated - LD light:dark cycle - LL constant light - DD constant darkness - DD_b constant darkness before exposure to constant light - DD_a constant darkness after exposure to constant light     

Locomotor activity rhythms in cave fishes from Chapada Diamantina, northeastern Brazil (Teleostei: Siluriformes)

Previous studies on the locomotor activity of troglobitic (exclusively subterranean) species have shown that circadian rhythmicity may be reduced in populations evolving in the absence of zeitgebers such as daily cycles of light and temperature; therefore, circadian activity rhythms, although not infradian nor ultradian rhythms, seem to have been selected by external, ecological factors. We studied the locomotor activity of a highly specialized Heptapteridae catfish (undescribed genus and species) from Chapada Diamantina, NE Brazil, compared to another specialized Brazilian troglobitic heptapterid, Taunayia sp. Locomotor activity was continuously measured in the laboratory with an infra-red photocell system. Seven specimens of the new genus were tested, each one during 14 consecutive days according to the following schedule: three days in DD → seven days in LD (12:12 h) → four days in DD. Data were submitted both to fast Fourier transform periodogram followed by Siegel's test of significance and Lombs - Scargle periodogram techniques in order to identify spectral composition of the time series. In general, results were similar to those obtained for Taunayia sp.: (a) for most specimens, absence of significant circadian components in locomotor activity under DD; (b) for all specimens, significant circadian components under LD, with higher levels of activity during the dark phase, as expected for species belonging to nocturnal epigean taxa; (c) for most specimens, no residual oscillations recorded when free-running conditions were reinstalled. Circadian locomotor activity detected under LD may thus be interpreted as a direct, masking effect of the LD cycle. This suggests a pattern for highly specialized troglobitic species, isolated for a long time in the subterranean habitat, with a progressive reduction of circadian time-keeping mechanisms. Our studies also demonstrate the potential of subterranean organisms for investigation of the origin, evolution, functioning and genetics of circadian rhthmicity.     

Circadian rhythms of locomotor activity and temperature selection in sleepy lizards, <Emphasis Type="Italic">Tiliqua rugosa</Emphasis>

This study examined whether the daily rhythms of locomotor activity and behavioural thermoregulation that have previously been observed in Australian sleepy lizards (Tiliqua rugosa) under field conditions are true circadian rhythms that persist in constant darkness (DD) and whether these rhythms show similar characteristics. Lizards held on laboratory thermal gradients in the Australian spring under the prevailing 12-hour light : dark (LD) cycle for 14 days displayed robust daily rhythms of behavioural thermoregulation and locomotor activity. In the 13-day period of DD that followed LD, most lizards exhibited free-running circadian rhythms of locomotor activity and behavioural thermoregulation. The predominant activity pattern displayed in LD was unimodal and this was retained in DD. While mean levels of skin temperature and locomotor activity were found to decrease from LD to DD, activity duration remained unchanged. The present results demonstrate for the first time that this species’ daily rhythm of locomotor activity is an endogenous circadian rhythm. Our results also demonstrate a close correlation between the circadian activity and thermoregulatory rhythms in this species indicating that the two rhythms are controlled by the same master oscillator(s). Future examination of seasonal aspects of these rhythms, may, however, cause this hypothesis to be modified.     

THERMOCYCLIC AND PHOTOCYCLIC ENTRAINMENT OF CIRCADIAN LOCOMOTOR ACTIVITY RHYTHMS IN SLEEPY LIZARDS,TILIQUA RUGOSA

Australian sleepy lizards (Tiliqua rugosa) exhibit marked locomotor activity rhythms in the field and laboratory. Light-dark (LD) and temperature cycles (TCs) are considered important for the entrainment of circadian locomotor activity rhythms and for mediating seasonal adjustments in aspects of these rhythms, such as phase, amplitude, and activity pattern. The relative importance of 24 h LD and TCs in entraining the circadian locomotor activity rhythm in T. rugosa was examined in three experiments. In the first experiment, lizards were held under LD 12:12 and subjected to either a TC of 33:15?°?C in phase with the LD cycle or a reversed TC positioned in antiphase to the LD cycle. Following LD 12:12, lizards were maintained under the same TCs but were subjected to DD. Activity was restricted to the thermophase in LD, irrespective of the lighting regime and during the period of DD that followed, suggesting entrainment by the TC. The amplitude of the TC was lowered by 8?°?C to reduce the intensity and possible masking effect of the TC zeitgeber in subsequent experiments. In the second experiment, lizards were held under LD 12.5:11.5 and subjected to one of three treatments: constant 30?°?C, normal TC (30:20?°?C) in phase with the LD cycle, or reversed TC. Following LD, all lizards were subjected to DD and constant 30?°?C. Post-entrainment free-run records revealed that LD cycles and TCs could both entrain the locomotor rhythms of T. rugosa. In LD, mean activity duration (α) of lizards in the normal TC group was considerably less than that in the constant 30?°?C group. Mean α also increased between LD and DD in lizards in the normal TC group. Although there was large variation in the phasing of the rhythm in relation to the LD cycle in reversed TC lizards, TCs presented in phase with the LD cycle most accurately synchronized the rhythm to the photocycle. In the third experiment, lizards were held in DD at constant 30?°?C before being subjected to a further period of DD and one of four treatments: normal TC (06:00 to 18:00 h thermophase), delayed TC (12:00 to 00:00 h thermophase), advanced TC (00:00 to 12:00 h thermophase), or control (no TC, constant 30?°?C). While control lizards continued to free-run in DD at constant temperature, the locomotor activity rhythms of lizards subjected to TCs rapidly entrained to TCs, whether or not the TC was phase advanced or delayed by 6 h. There was no difference in the phase relationships of lizard activity rhythms to the onset of the thermophase among the normal, delayed, and advanced TC groups, suggesting equally strong entrainment to the TC in each group. The results of this experiment excluded the possibility that masking effects were responsible for the locomotor activity responses of lizards to TCs. The three experiments demonstrated that TCs are important for entraining circadian locomotor activity rhythms of T. rugosa, even when photic cues are conflicting or absent, and that an interaction between LD cycles and TCs most accurately synchronizes this rhythm. (Author correspondence: david.j.ellis@adelaide.edu.au)     

Daily oscillation in melatonin synthesis in the Turkey pineal gland and retina: diurnal and circadian rhythms

The aim of the present study was to examine arylalkylamine N-acetyltransferase (AANAT) activity and melatonin content in the pineal gland and retina as well as the melatonin concentration in plasma of the turkey (Meleagris gallopavo), an avian species in which several physiological processes, including reproduction, are controlled by day length. In order to investigate whether the analyzed parameters display diurnal or circadian rhythmicity, we measured these variables in tissues isolated at regular time intervals from birds kept either under a regular light-dark (LD) cycle or under constant darkness (DD). The pineal gland and retina of the turkey rhythmically produced melatonin. In birds kept under a daily LD cycle, melatonin levels in the pineal gland and retina were high during the dark phase and low during the light phase. Rhythmic oscillations in melatonin, with high night-time concentrations, were also found in the plasma. The pineal and retinal melatonin rhythms mirrored oscillations in the activity of AANAT, the penultimate enzyme in the melatonin biosynthetic pathway. Rhythmic oscillations in AANAT activity in the turkey pineal gland and retina were circadian in nature, as they persisted under conditions of constant darkness (DD). Transferring birds from LD into DD, however, resulted in a potent decline in the amplitude of the AANAT rhythm from the first day of DD. On the sixth day of DD, pineal AANAT activity was still markedly higher during the subjective dark than during the subjective light phase; whereas, AANAT activity in the retina did not exhibit significant oscillations. The results indicate that melatonin rhythmicity in the turkey pineal gland and retina is regulated both by light and the endogenous circadian clock. The findings suggest that environmental light may be of primary importance in the maintenance of the high-amplitude melatonin rhythms in the turkey.