Feeding-entrainable Oscillator Research Articles

FEEDING ENTRAINMENT OF DAILY RHYTHMS OF LOCOMOTOR ACTIVITY AND CLOCK GENE EXPRESSION IN ZEBRAFISH BRAIN

Light and feeding cycles strongly synchronize daily rhythms in animals, which may, as a consequence, develop food anticipatory activity (FAA). However, the light/food entraining mechanisms of the central circadian oscillator remain unknown. In this study, we investigate the existence of FAA in seven groups of zebrafish subjected to a light/dark (LD) cycle or constant light (LL) and different feeding regimes (random, fasting, and feeding in the middle of the light phase or dark phase). The aim was to ascertain whether the daily rhythm of behavior and clock gene (per1 and cry1) expression in the zebrafish brain was entrained by the light and feeding regime. The results revealed that FAA developed in zebrafish fed daily at a fixed time, under LD and under LL. Zebrafish displayed locomotor activity mostly during the daytime, although the percentage of activity during the light phase varied depending on feeding time (ranging from 93.2% to 63.1% in the mid-light and mid-dark fed groups, respectively). However, the different feeding regimes failed to modify the daily rhythm of per1 and cry1 expression in the zebrafish brain under LD (approximate acrophases [peak times] at ZT22 and ZT4, respectively; lights-on = ZT0). Under LL, per1 and cry1 expression did not show significant daily rhythmicity, regardless of the feeding regime. These findings indicate that, although schedule-fed zebrafish developed FAA as regards locomotor activity, feeding had little effect on clock gene expression in whole brain homogenates, suggesting the feeding-entrainable oscillator may be located elsewhere or at specific brain sites. (Author correspondence: jasanchez@um.es)

Feeding Entrainment of Daily Rhythms of Locomotor Activity and Clock Gene Expression in Zebrafish Brain

Light and feeding cycles strongly synchronize daily rhythms in animals, which may, as a consequence, develop food anticipatory activity (FAA). However, the light/food entraining mechanisms of the central circadian oscillator remain unknown. In this study, we investigate the existence of FAA in seven groups of zebrafish subjected to a light/dark (LD) cycle or constant light (LL) and different feeding regimes (random, fasting, and feeding in the middle of the light phase or dark phase). The aim was to ascertain whether the daily rhythm of behavior and clock gene (per1 and cry1) expression in the zebrafish brain was entrained by the light and feeding regime. The results revealed that FAA developed in zebrafish fed daily at a fixed time, under LD and under LL. Zebrafish displayed locomotor activity mostly during the daytime, although the percentage of activity during the light phase varied depending on feeding time (ranging from 93.2% to 63.1% in the mid-light and mid-dark fed groups, respectively). However, the different feeding regimes failed to modify the daily rhythm of per1 and cry1 expression in the zebrafish brain under LD (approximate acrophases [peak times] at ZT22 and ZT4, respectively; lights-on = ZT0). Under LL, per1 and cry1 expression did not show significant daily rhythmicity, regardless of the feeding regime. These findings indicate that, although schedule-fed zebrafish developed FAA as regards locomotor activity, feeding had little effect on clock gene expression in whole brain homogenates, suggesting the feeding-entrainable oscillator may be located elsewhere or at specific brain sites. (Author correspondence: jasanchez@um.es)

Feeding entrainment of locomotor activity rhythms, digestive enzymes and neuroendocrine factors in goldfish

The existence of food anticipatory activity (FAA) in animals subjected to daily feeding schedules seems to be mediated by a feeding-entrainable oscillator (FEO). Such an FEO may help in anticipating meal time and so optimizing food acquisition and nutrient utilization. In this study we investigated the existence of FAA and whether digestive enzymes, plasma cortisol, hypothalamic NPY and gastrointestinal tract (GIT) and plasma melatonin were entrained by periodic feeding in goldfish. We observed that periodically fed goldfish showed FAA in locomotor activity as well as in amylase and NPY. Alkaline protease and GIT melatonin were higher after feeding, whereas plasma cortisol levels were reduced. Plasma melatonin remained unmodified before and after meal time. These results suggested that scheduled feeding entrained both behavioral and certain physiological patterns in goldfish, FAA being of adaptive value to anticipate a meal and prepare the digestive physiology of fish.

Modifications of local cerebral glucose utilization during circadian food-anticipatory activity

Food-anticipatory activity that animals express before a daily timed meal is considered as the behavioral output of a feeding-entrainable oscillator whose functional neuroanatomy is still unknown. In order to identify the possible brain areas involved in that timing mechanism, we investigated local cerebral metabolic rate for glucose during food-anticipatory activity produced either by a 4-h daily access to food starting 4 h after light onset or by a hypocaloric feeding provided at the same time. Local cerebral metabolic rate for glucose measured by the labeled 2-[ 14C]-deoxyglucose technique was quantified in 40 structures. In both groups of food-restricted rats, three brain regions (the nucleus of the solitary tract, the cerebellar cortex and the medial preoptic area) showed a decrease in local cerebral metabolic rate for glucose, compared with control ad libitum animals. In addition, only one structure, the paraventricular thalamic nucleus, was affected by temporal restricted feeding, and not by hypocaloric feeding, compared with ad libitum rats. By contrast, three brain regions, i.e. the intergeniculate leaflets, the paraventricular hypothalamic and the arcuate nuclei, showed specifically metabolic decreases during anticipation of hypocaloric feeding, and not during anticipation of temporal restricted feeding, compared with the ad libitum group. Expression of food-anticipatory activity appears to be regulated by an integrated neural circuit of brainstem and hypothalamic pathways, with hypocaloric feeding involving more extensive forebrain areas than temporal restricted feeding.

Demand-feeding rhythms and feeding-entrainment of locomotor activity rhythms in tench ( Tinca tinca)

Tench ( Tinca tinca) has been described as a strictly nocturnal species whose locomotor activity rhythms, albeit strongly synchronised by light, have an endogenous nature. Aside from light, a number of other environmental factors, such as mealtime, can act as circadian system synchronisers in fish; however, there is a scarcity of information on tench feeding rhythms. This study describes daily self-feeding rhythms in tench, and analyses the role of feeding time on synchronisation of locomotor activity rhythms. Tench were able to operate string sensor-activated self-feeders, and they displayed a strictly nocturnal behavior, both under indoor and outdoor conditions. Locomotor activity remained strictly nocturnal irrespective of whether tench were fed only during the scotophase (D-feeding) or the photophase (L-feeding). However, no statistically significant differences were detected between both groups in terms of food intake or growth performance. Furthermore, unlike L-feeding, D-feeding elicited a clear anticipatory activity (FAA). When tench were given the possibility of feeding at both times of the day, they showed a clear preference for D-feeding. Finally, in fish exposed to constant darkness (DD), feeding time acted as a true zeitgeber and FAA was observed. When animals were fasted under DD conditions, locomotor activity free-run and 6 out of 12 individuals yielded significant results in the periodogram analysis. Under DD, fish resynchronised when regular food was resumed, with some tench displaying FAA. The obtained results indicated the existence of a feeding-entrainable oscillator (FEO) in tench.

Differential effects of meal size and food energy density on feeding entrainment in goldfish.

The synchronizing stimulus, its transduction site, and the afferent pathways responsible for feeding entrainment remain unknown. In fish, the role of the diet in the development of feeding anticipatory activity (FAA) is not well understood and fundamental questions on the mechanisms of feeding entrainment, such as the meal characteristics required to develop FAA, remain unexplored. To test the entraining properties of daily meals with different sizes and energy densities, activity rhythms were studied after a 12-h shift of the feeding cycle in individual goldfish under constant light. In the 1st experiment, the energy content of a control diet (16.7 kJ/g) was diluted by replacing 50% (8.3 kJ/g) or 90% (1.7 kJ/g) of the diet with cellulose. However, the number of days required to stabilize FAA after the shift did not differ statistically between diets. In the 2nd experiment, meal size was modified by reducing the daily feeding ration to 0.5% and 0.1% b.wt.d(-1). In this case, differences in the entraining properties of the two feeding rations appeared because goldfish fed at 0.1% b.wt.d(-1) resynchronized faster than those fed at 0.5% b.wt.d(-1). These results revealed that the dilution of the dietary energy up to 1.7 kJ/g had no significant effect on the entraining properties of the feeding-entrainable oscillator (FEO), whereas the reduction of the meal size to 0.1% b.wt.d(-1) provoked a faster resynchronization after shifting the daily meal cycle. Taken together, these results suggest that gut distension may be involved in feeding entrainment, as a reduction in meal size but not in the amount of dietary energy supplied significantly shortened the time required for resynchronization and highlighted the different synchronizing properties of meal size and energy density as zeitgebers for the FEO.

Food-anticipatory activity persists after olfactory bulb ablation in the rat

Restricted daily food access acts as an entraining stimulus (zeitgeber) for a circadian clock, the feeding-entrainable oscillator (FEO). There are many properties of a daily meal that could potentially convey timing information to the FEO. Olfactory cues associated with feeding are one such property. In order to rule out olfaction as a necessary entraining stimulus, olfactory bulbectomized and sham-operated male Sprague–Dawley rats had access to food for 2 h each day. Food bin approach behavior was monitored as an index of food-anticipatory activity (FAA). Both groups entrained to the daily meal with an increase in feeder approach time several hours before meal onset. There were no significant differences in the timing or the amount of FAA between groups. Furthermore, FAA was maintained during 3 days of food deprivation in both groups. In accordance with previous studies, the results show that olfactory cues are not necessary for the entrainment of FAA.

Circadian thermoregulation in suckling rabbit pups.

The rabbit pup is well suited to track the age-dependent development of periodic thermoregulation during the suckling period. Since the litters are regularly nursed once per day for a total of 3 to 4 min, an exogenous, metabolic, nonphotic periodic variable is supposed to have an impact on the 24-h rhythm of body temperature. The authors monitored the course of core body temperature during the suckling period of 20 pups by means of a transmitter implanted intraperitoneally on day 3 postpartum. The 24-h mean rose from an average of 37.8+/-0.3 degrees C on day 4 of life to 39.5+/-0.2 degrees C at weaning on day 27, for 2 out of 20 pups, and day 28, for 18 out of 20 pups. In constant dim illumination, the pups exhibited a 24-h rhythm even on postnatal day 4, which consolidated around days 5 to 7. The rhythm consisted of a significant anticipatory rise of 0.4 to 0.6 degrees C above the respective 24-h mean commencing 2.5 to 3.5 h prior to nursing. Milk intake was followed by a further increase of temperature for an additional 0.3 to 0.6 degrees C. Then the temperature dropped for 1.2 to 1.5 degrees C within 1 to 3 h and returned to average 3 to 5 h later. During a 48-h fast, the rhythm continued to exist, though in a modified shape: the anticipatory component persisted almost unchanged; a further elevation of temperature, however, did not occur. Thus, the anticipatory component apparently is generated endogenously and the second surge represents an exogenous suckling-induced, thermogenic peak. When maternal nursing was advanced for 15 min/day for a total of 5 h, the temperature rhythm of the pups followed the shift of the zeitgeber in parallel. These data confirm the assumption that a circadian rhythm exists during the first postnatal days of the rabbit and that this rhythm is entrained by the 24-h nursing rhythm. The authors suggest that the biological significance of a feeding entrainable oscillator (FEO) in the rabbit might be to activate the pups prior to the periodic nursing visit of the rabbit doe. Thus, the pups are prepared to quantitatively use the one and only short nursing episode per day for maximal milk ingestion.

Glucose, but not fat, phase shifts the feeding-entrained circadian clock

STEPHAN, F. K. AND A. J. DAVIDSON. Glucose, but not fat, phase shifts the feeding-entrained circadian clock. PHYSIOL BEHAV 65(2) 277–288, 1998.—To study the ability of single macronutrients to entrain or phase shift the feeding entrainable circadian oscillator, rats with lesions of the suprachiasmatic nucleus were first maintained on a single daily meal of lab chow until robust anticipatory approaches to the feeder or anticipatory wheel running was established. The meal time was then delayed by 8 h and chow was replaced with a 25-mL solution of 0.2% saccharin or 25 mL of saccharin plus 15 g of glucose. For other phase shifts, rats received either 6 mL of vegetable oil or mineral oil for 2 consecutive days. Consumption of about 6 g (24 kcal) or more of glucose resulted in robust delaying transients on the days after ingestion, whereas saccharin induced only small delays consistent with the initiation of a free-running rhythm with a period greater than 24 h. Surprisingly, consumption of 5.5 g of vegetable oil (47 kcal) did not result in delays greater than those in rats receiving mineral oil. The introduction of oil also produced a severe reduction in approaches to the feeder which could be alleviated by placing inaccessible chow in the feeders between oil meals. Phase shifts with oil were repeated with rats housed in wheels using anticipatory wheel running as a phase marker to assess whether the lack of phase shifts with fat was apparatus dependent. As was the case with approach behavior, anticipatory wheel running was not significantly delayed by vegetable oil consumption. These results indicate that a simple monosaccharide, glucose, has zeitgeber properties for the feeding entrainable oscillator. Vegetable oil, despite a higher caloric content, may be ineffective because of slower gastric emptying and nutrient absorption or because fat is not a good zeitgeber for the feeding entrained circadian oscillator.

Calories Affect Zeitgeber Properties of the Feeding Entrained Circadian Oscillator

Rats with suprachiasmatic (SCN) lesions readily entrain to daily meals by increasing their activity prior to food access. Although previous experiments suggest that entrainment requires a nutritive meal, it is not yet clear what the parameters of the entraining stimulus are. The first experiment investigated the role of caloric content in resetting the feeding entrainable oscillator (FEO). Rats were entrained to 20 g chow/day until anticipatory activity was stable. The food access time was then delayed by 8 h and rats received either 0, 2, 6, or 16 g of chow for two days. Sixteen g of chow produced large delays on the next two cycles, while 0 and 2 g produced no delays. Two of 8 rats receiving 6 g showed delays, indicating that 22 kcal is near the threshold. In a second experiment, the effects of bulk were investigated. After an 8 h phase delay, rats received 0, 6, 10 or 16 g of chow mixed with cellulose for a total of 21 g for all groups. The 10 and 16 g chow groups delayed while the 0 g chow group did not. In the 6 g groups some animals phase shifted while others did not. Thus, the addition of non-nutritive bulk to the phase shifted meal had little or no effect on resetting the FEO and it appears that caloric content rather than gastric distention provides an effective zeitgeber for the FEO.

Light-dark and food restriction cycles in sea bass: Effect of conflicting zeitgebers on demand-feeding rhythms

Sea bass, a fish species characterized by its dualistic feeding pattern, was investigated to study the synchronizing effect of light and food on the demand-feeding rhythm. Nocturnal and diurnal sea bass, both in groups and individually, were exposed to restricted-feeding (RF) and light-dark (LD) cycles of different periods. The phase relationship between both zeitgebers was also studied. The results show that food-demand rhythms synchronize to periodic food availability under constant light conditions (DD) and that there is a partial coupling between food-entrained and light-entrained activity under conflicting zeitgebers (LD 13:13 h and RF 4:20 h), suggesting the existence of a feeding entrainable oscillator (FEO) in addition to the master light entrainable oscillator (LEO). In some cases, food availability restricted to the light or dark phase contrary to that of the previous feeding phase changed a diurnal feeding pattern into nocturnal and viceversa, suggesting that food can be one of the switching factors that decides whether the circadian system of sea bass is nocturnal or diurnal. However, the fact that the feeding pattern of some fish was unrelated with the phase in which food was available suggests that other internal and/or external factors could be involved in the temporal flexibility of sea bass.

Ontogeny of the rabbit's circadian rhythms without an external zeitgeber

We have monitored parturition and different behavioral rhythms of does, the activity of suckling pups and behavioral rhythms of rabbits after weaning until adulthood. All animals were living in continuous light conditions (LL). As a consequence of the free-run of the does parturition, which takes place during the resting period of the animal, occurred at almost any time of the day. The once/day nursing was advanced during the first 7–10 days until it was in synchrony with the time of main activity. The pups significantly anticipated nursing by day 5: the 60 min bin prior to nursing exceeded the 24-h mean by 63 ± 30%. The anticipatory activity (AA) was entrained by an artificial nursing schedule of T = 24 h, the 60 min bin prior to nursing exceeding the 24-h mean by 123 ± 50%. Since AA persisted during a 48 h fast, its phase is controlled endogenously. At weaning behavioral rhythms of rabbits free-ran with a main tau <24 h (average 23:35 ± 17′), period length extending with increasing age. 7 10 animals had an additional component >24 h (average: 24:24 ± 16′) which became increasingly dominant, the components <24 h fading out within 30 days after weaning. Between days 340–350 a stable period length of 24:35 ± 8′ and a consolidation of the circadian rhythms was attained. These results confirm that during the first days of the rabbit's life the circadian system is controlled by a feeding entrainable oscillator, the light entrainable oscillators taking over control later.

Activity-stress ulcers in rats: The role of preentrainment to meal time

Rats housed in activity wheels (A-W) or hanging cages (HC) received food restriction to 4 h/day for 15 days. Food was given at fixed times to entrain the feeding entrainable oscillator or at irregular times to prevent entrainment. One half of the rats received food in the light phase, the other half in the dark phase of the light-dark cycle. Food was then reduced to 6 g/day for 4 days for A-W rats of 7 days for HC rats. Stomachs were removed 24 h after the last meal and the ulcerated area was quantified. Twenty one of 32 A-W rats and 19 of 32 HC rats had ulcers in the glandular stomach. Ulcers were significantly larger in A-W rats ( p < 0.01). Activity levels (in A-W rats), % body weight loss and terminal body weight (in both groups) were significantly correlated with the area of ulcers. There was no significant time of day effect on the ulcer severity. However, an unexpected finding was that for A-W rats, those fed at fixed times had larger ulcers than those fed at irregular times ( p < 0.01) while HC rats showed an expected trend in the opposite direction. One possible explanation for this result is that for A-W rats, the reduced food supply at an expected time of day generated a greater stress response than in rats whose feeding time was unpredictable.

Circadian rhythms of rabbits during restrictive feeding.

Without a zeitgeber the circadian rhythms of five physiological functions free-ran with a period length greater than 24 h. Restricted feeding time (RF) masked the free-running rhythms. In addition to masking, entrainment with RF occurred. This process was most evident in locomotor activity and visits to the food box. RF thus had zeitgeber properties in these rabbits. However, in most rabbits the RF zeitgeber was not strong enough to entrain the circadian rhythm completely. A small component free-ran during RF. Following return to continuous food access the whole circadian rhythm resumed to free-run again. In some animals its phase was determined by the RF zeitgeber and in others by the small free-running fraction present during RF. The results suggest that in addition to the light-dark-entrainable circadian oscillator system a feeding-entrainable oscillator exists that takes over phase control of the majority of the rhythm during RF.