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Systemic glucose metabolism and insulin activity oscillate in response to diurnal rhythms and nutrient availability with the necessary involvement of adipose tissue to maintain metabolic homeostasis. However, the adipose‐intrinsic regulatory mechanism remains elusive. Here, the dynamics of PPARγ acetylation in adipose tissue are shown to orchestrate metabolic oscillation in daily rhythms. Acetylation of PPARγ displays a diurnal rhythm in young healthy mice, with the peak at zeitgeber time 0 (ZT0) and the trough at ZT18. This rhythmic pattern is deranged in pathological conditions such as obesity, aging, and circadian disruption. The adipocyte‐specific acetylation‐mimetic mutation of PPARγ K293Q (aKQ) restrains adipose plasticity during calorie restriction and diet‐induced obesity, associated with proteolysis of a core circadian component BMAL1. Consistently, the rhythmicity in glucose tolerance and insulin sensitivity is altered in aKQ and the complementary PPARγ deacetylation‐mimetic K268R/K293R (2KR) mouse models. Furthermore, the PPARγ acetylation‐sensitive downstream target adipsin is revealed as a novel diurnal factor that destabilizes BMAL1 and mediates metabolic rhythms. These findings collectively signify that PPARγ acetylation is a hinge connecting adipose plasticity and metabolic rhythms, the two determinants of metabolic health. [ABSTRACT FROM AUTHOR]

Dopamine has been implicated in circadian timing underlying the food entrainable oscillator (FEO) circuitry and overexpression of the dopamine D2 receptor (D2R) in the striatum has been reported to reduce motivation to obtain food rewards in operant tasks. In the present study, we explored both of these mechanisms by examining food anticipatory activity (FAA) in dopamine D2 receptor‐overexpressing (D2R‐OE) mice under various durations of food availability. First, we noted that at baseline, there were no differences between D2R‐OE mice and their littermates in activity level, food intake, and body weight or in circadian activity. Under conditions of very restricted food availability (4 or 6 hr), both genotypes displayed FAA. In contrast, under 8‐hr food availability, control mice showed FAA, but D2R‐OE mice did not. Normalization of D2R by administration of doxycycline, a tetracycline analogue, rescued FAA under 8‐hr restricted food. We next tested for circadian regulation of FAA. When given ad libitum access to food, neither D2R‐OE nor controls were active during the daytime. However, after an interval of food restriction, all mice showed elevated locomotor activity at the time of previous food availability in the day, indicating circadian timing of anticipatory activity. In summary, motivation is reduced in D2R‐OE mice but circadian timing behavior is not affected. We conclude that an increase in striatal D2R reduces FAA by modulating motivation and not by acting on a clock mechanism. [ABSTRACT FROM AUTHOR]

The hypothalamic suprachiasmatic nucleus ( SCN), locus of the master circadian clock, bears many neuronal types. At the cellular-molecular level, the clock is comprised of feedback loops involving 'clock' genes including Period1 and Period2, and their protein products, PERIOD1 and PERIOD2 ( PER1/2). In the canonical model of circadian oscillation, the PER1/2 proteins oscillate together. While their rhythmic expression in the SCN as a whole has been described, the possibility of regional differences remains unknown. To explore these clock proteins in distinct SCN regions, we assessed their expression through the rostro-caudal extent of the SCN in sagittal sections. We developed an automated method for tracking three fluorophores in digital images of sections triply labeled for PER1, PER2, and gastrin-releasing peptide (used to locate the core). In the SCN as a whole, neurons expressing high levels of PER2 were concentrated in the rostral, rostrodorsal, and caudal portions of the nucleus, and those expressing high levels of PER1 lay in a broad central area. Within these overall patterns, adjacent cells differed in expression levels of the two proteins. The results demonstrate spatially distinct localization of high PER1 vs. PER2 expression, raising the possibility that their distribution is functionally significant in encoding and communicating temporal information. The findings provoke the question of whether there are fundamental differences in PER1/2 levels among SCN neurons and/or whether topographical differences in protein expression are a product of SCN network organization rather than intrinsic differences among neurons. [ABSTRACT FROM AUTHOR]

The suprachiasmatic nucleus (SCN) is the locus of the master circadian clock, setting the daily rhythms in physiology and behavior and synchronizing these responses to the local environment. The most important of these phase-setting cues derive from the light-dark cycle and reach the SCN directly via the retinohypothalamic tract (RHT). The SCN contains anatomically and functionally heterogeneous populations of cells. Understanding how these neurons access information about the photic environment so as to set the phase of daily oscillation requires knowledge of SCN innervation by the RHT. While retinal innervation of the SCN has long been a topic of interest, the information is incomplete. In some instances, studies have focused on the caudal aspect of the nucleus, which contains the core region. In other instances, subregions of the nucleus have been delineated based on projections of where specific peptidergic cell types lie, rather than based on double or triple immunochemical staining of distinct populations of cells. Here, we examine the full extent of the mouse SCN using cholera toxin β (CTβ) as a tracer to analyze RHT innervation in triple-labeled sagittal sections. Using specific peptidergic markers to identify clusters of SCN cells, we find 3 distinct patterns. First is an area of dense RHT innervation to the core region, delineated by gastrin-releasing peptide (GRP) and vasoactive intestinal peptide (VIP) immunoreactive cells. Second is an area of moderate RHT fiber clusters, bearing arginine-vasopressin (AVP)–positive cells that lie close to the core. Finally, the outermost, shell, and rostral AVP-containing regions of the SCN have few to no detectable retinal fibers. These results point to a diversity of inputs to individual SCN cell populations and suggest variation in the responses that underlie photic phase resetting. [ABSTRACT FROM AUTHOR]

Research on the mechanisms underlying circadian rhythmicity and the response of brain and body clocks to environmental and physiological challenges requires assessing levels of circadian clock proteins. Too often, however, it is difficult to acquire antibodies that specifically and reliably label these proteins. Many of these antibodies also lack appropriate validation. The goal of this project was to generate and characterize antibodies against several circadian clock proteins. We examined mice and hamsters at peak and trough times of clock protein expression in the suprachiasmatic nucleus (SCN). In addition, we confirmed specificity by testing the antibodies on mice with targeted disruption of the relevant genes. Our results identify antibodies against PER1, PER2, BMAL1 and CLOCK that are useful for assessing circadian clock proteins in the SCN by immunocytochemistry. [ABSTRACT FROM AUTHOR]

Light intensity is an important determinant of diverse physiological and behavioral responses within the non-image-forming visual system. Thresholds differ among various photic responses, namely control of circadian rhythms, vigilance state, activity level and pupil constriction, but the mechanisms that regulate photosensitivity are not known. Calbindin D (CalB) is a calcium-binding protein associated with light processing in the mammalian circadian clock. Loss-of-function studies indicate that CalB-deficient mice (CalB) have deficits in their ability to entrain to light-dark cycles. To explore the role of CalB in modulating photosensitivity, thresholds for three behaviors mediated by the non-image-forming visual system (entrainment, masking and pupillary light reflex; PLR) were compared in CalB and wildtype mice, and the localization of CalB protein in these circuits was examined in adult and juvenile mice. The results reveal a divergence in how CalB affects thresholds to photic cues among these responses. Entrainment and masking were 40- to 60-fold less sensitive in CalB than in wildtype mice. On the other hand, the PLR in CalB mice was 80- to 200-fold more sensitive. Though CalB is expressed in the retina and in brain circuits regulating entrainment we found no CalB expression in any component of the PLR pathway, namely the olivary pretectal nucleus, Edinger-Westphal nucleus and ciliary ganglion. The behavioral and anatomical data together suggest that, in normal animals, the retinal response to light is blunted in the presence of CalB, but responsiveness of the higher order processes that transduce afferent retinal input is enhanced. [ABSTRACT FROM AUTHOR]

Because we can observe oscillation within individual cells and in the tissue as a whole, the suprachiasmatic nucleus (SCN) presents a unique system in the mammalian brain for the analysis of individual cells and the networks of which they are a part. While dispersed cells of the SCN sustain circadian oscillations in isolation, they are unstable oscillators that require network interactions for robust cycling. Using cluster analysis to assess bioluminescence in acute brain slices from PERIOD2::Luciferase (PER2::LUC) knockin mice, and immunochemistry of SCN from animals harvested at various circadian times, we assessed the spatiotemporal activation patterns of PER2 to explore the emergence of a coherent oscillation at the tissue level. The results indicate that circadian oscillation is characterized by a stable daily cycle of PER2 expression involving orderly serial activation of specific SCN subregions, followed by a silent interval, with substantial symmetry between the left and right side of the SCN. The biological significance of the clusters identified in living slices was confirmed by co-expression of LUC and PER2 in fixed, immunochemically stained brain sections, with the spatiotemporal pattern of LUC expression resembling that revealed in the cluster analysis of bioluminescent slices. We conclude that the precise timing of PER2 expression within individual neurons is dependent on their location within the nucleus, and that small groups of neurons within the SCN give rise to distinctive and identifiable subregions. We propose that serial activation of these subregions is the basis of robustness and resilience of the daily rhythm of the SCN. [ABSTRACT FROM AUTHOR]

Calbindin-D28K (CalB)-containing cells form a distinct cluster within the core of the hamster suprachiasmatic nucleus (SCN). These cells are directly retinorecipient but lack detectable rhythms in clock gene expression or electrical activity. In studies exploring SCN connectivity using double-label immunochemistry, we previously reported an absence of contacts among CalB fibers and vasopressin (VP) cells in animals sacrificed during the day. Here, we explored circadian variations in CalB-immunoreactivity (-ir) and re-examined the connections between CalB and other SCN cell types at zeitgeber times (ZT) 4 and 14. The results reveal a circadian rhythm of CalB-ir in fibers of SCN cells with high expression during the night and subjective night and low expression during the day and subjective day. This circadian difference is not seen in the other brain regions studied. Significantly more appositions were detected between CalB fibers and VP cells during the night than during the day, while circadian variation in numbers of contacts was not seen between CalB fibers and vasoactive intestinal polypeptide (VIP), cholecystokinin (CCK), or gastrin-releasing peptide (GRP) cells. There was no detectable variation in appositions from any peptidergic fiber type onto CalB cells. The present findings suggest that CalB cells relay photic information to VP oscillator cells of the SCN shell in a temporally gated manner [ABSTRACT FROM AUTHOR]

The mechanisms by which animals adapt to an ever-changing environment have long fascinated scientists. Different forces, conveying information regarding various aspects of the internal and external environment, interact with each other to modulate behavioral arousal. These forces can act in concert or, at times, in opposite directions. These signals eventually converge and are integrated to influence a common arousal pathway which, depending on all the information received from the environment, supports the activation of the most appropriate behavioral response. In this review we propose that the ventromedial hypothalamic nucleus (VMN) is part of the circuitry that controls food anticipation. It is the first nucleus activated when there is a change in the time of food availability, silencing of VMN ghrelin receptors decreases food-anticipatory activity (FAA) and, although lesions of the VMN do not abolish FAA, parts of the response are often altered. In proposing this model it is not our intention to exclude parallel, redundant and possibly interacting pathways that may ultimately communicate with, or work in concert with, the proposed network, but rather to describe the neuroanatomical requirements for this circuit and to illustrate how the VMN is strategically placed and connected to mediate this complex behavioral adaptation. [ABSTRACT FROM AUTHOR]

The suprachiasmatic nucleus (SCN) is the principal circadian pacemaker in mammals. A salient feature of the SCN is that cells of a particular phenotype are topographically organized; this organization defines functionally distinct subregions that interact to generate coherent rhythmicity. In Syrian hamsters ( Mesocricetus auratus), a dense population of directly retinorecipient calbindin D28K (CalB) neurons in the caudal SCN marks a subregion critical for circadian rhythmicity. In mouse SCN, a dense cluster of CalB neurons occurs during early postnatal development, but in the adult CalB neurons are dispersed through the SCN. In the adult retina CalB colocalizes with melanopsin-expressing ganglion cells. In the present study, we explored the role of CalB in modulating circadian function and photic entrainment by investigating mice with a targeted mutation of the CalB gene (CalB−/− mice). In constant darkness (DD), CalB−/− animals either become arrhythmic (40%) or exhibit low-amplitude locomotor rhythms with marked activity during subjective day (60%). Rhythmic clock gene expression is blunted in these latter animals. Importantly, CalB−/− mice exhibit anomalies in entrainment revealed following transfer from a light : dark cycle to DD. Paradoxically, responses to acute light pulses measured by behavioral phase shifts, SCN FOS protein and Period1 mRNA expression are normal. Together, the developmental pattern of CalB expression in mouse SCN, the presence of CalB in photoresponsive ganglion cells and the abnormalities seen in CalB−/− mice suggest an important role for CalB in mouse circadian function. [ABSTRACT FROM AUTHOR]

In the course of evolution, mechanisms have evolved to anticipate the timing of regularly occurring events. These mechanisms are encompassed in a circadian timing system that include a master clock localized to the suprachiasmatic nucleus of the hypothalamus and “slave” oscillators distributed throughout the body. This system serves multiple functions so as to ensure that various physiological processes occur at optimal and nonoverlapping times, to synchronize our activities to local environmental time, and to permit changes required to respond to new environmental circumstances. We suggest that a generalized concept of arousal (which includes alterations in responsiveness to homeostatic pressures, sensory stimuli and emotional reactivity, and to changes in motor activity) serves as a rubric in which to explore the multiple ways in which the circadian system modulates behavior. [ABSTRACT FROM AUTHOR]

The mechanisms underlying CNS arousal in response to homeostatic pressures are not known. In this study, we pitted two forces for CNS arousal against each other (circadian influences vs. restricted food availability) and measured the neuronal activation that occurs in a behaviorally defined group of animals that exhibited increased arousal in anticipation of feeding restricted to their normal sleeping time. The number of c-FOS+ neurons was significantly increased only in the ventromedial nucleus of the hypothalamus (VMH) in these mice, compared with control animals whose feeding was restricted to their normal active and feeding time (P < 0.01). Because the activation of VMH neurons coincides with the earliest signs of behavioral arousal preceding a change in mealtime, we infer that VMH activation is involved in the increased arousal in anticipation of food. [ABSTRACT FROM AUTHOR]

Abstract: The hamster suprachiasmatic nucleus (SCN) is anatomically and functionally heterogeneous. A group of cells in the SCN shell, delineated by vasopressin-ergic neurons, are rhythmic with respect to Period gene expression and electrical activity but do not receive direct retinal input. In contrast, some cells in the SCN core, marked by neurons containing calbindin-D28k, gastrin-releasing peptide (GRP), substance P (SP), and vasoactive intestinal polypeptide (VIP), are not rhythmic with respect to Period gene expression and electrical activity but do receive direct retinal input. Examination of the timing of neurogenesis using bromodeoxyuridine indicates that SCN cells are born between embryonic day 9.5 and 12.5. Calbindin, GRP, substance P, and VIP cells are born only during early SCN neurogenesis, between embryonic days 9.5–11.0. Vasopressin cells are born over the whole period of SCN neurogenesis, appearing as late as embryonic day 12.5. Examination of the ontogeny of peptide expression in these cell types reveals transient expression of calbindin in a cluster of dorsolateral SCN cells on postnatal days 1–2. The adult pattern of calbindin expression is detected in a different ventrolateral cell cluster starting on postnatal day 2. GRP and SP expression appear on postnatal day 8 and 10, respectively, after the retinohypothalamic tract has innervated the SCN. In summary, the present study describes the ontogeny-specific peptidergic phenotypes in the SCN and compares these developmental patterns to previously identified patterns in the appearance of circadian functions. These comparisons suggest the possibility that these coincident appearances may be causally related, with the direction of causation to be determined. [Copyright &y& Elsevier]

We investigated parvalbumin immunoreactivity (PA-IR) in the retinas of rats maintained on a 12:12 h light:dark cycle, or after being placed in constant darkness for 24–72 h. Retinas were harvested at zeitgeber and circadian times 02:00, 06:00, 10:00, 14:00, 18:00 and 22:00 h. PA-IR was found primarily in retinal amacrine cells of the AII subtype. In a light/dark cycle, PA-IR showed a clear rhythm, with a low near zeitgeber time (ZT) 10:00 h and a peak near ZT 18:00 h. The ratio of immunofluorescence intensities at these timepoints was >15-fold. When animals were kept in complete darkness for 1–3 days, the rhythm of PA-IR was still preserved, but was progressively reduced in amplitude. The rhythm of PA-IR inferred from immunohistochemical data was confirmed by Western blots. We conclude that PA-IR in the rat retina shows an underlying circadian rhythm that is enhanced by cyclic light. The regulation may involve translocation of the protein between cell compartments and/or new protein synthesis. [ABSTRACT FROM AUTHOR]

The suprachiasmatic nucleus (SCN) of the hypothalamus is the neural locus of the circadian clock. To explore the organization of the SCN, two strains of transgenic mice, each bearing jellyfish green fluorescent protein (GFP) reporter, were used. In one, GFP was driven by the promoter region of the mouse Period1 gene (mPer1) (Per1::GFP mouse), whereas in the other, GFP was inserted in the promoter region of calbindin-D[sub28k]--bacterial artificial chromosome (CalB::GFP mouse). In the latter mouse, GFP-containing SCN cells are immunopositive for gastrin-releasing peptide. In both mouse lines, light-induced Per1 mRNA and Fos are localized to the SCN subregion containing gastrin-releasing peptide. Double-label immunohistochemistry reveals that most gastrin-releasing peptide cells (&sim70%) contain Fos after a brief light pulse. To determine the properties of SCN cells in this light-responsive region, we examined the expression of rhythmic Period genes and proteins. Gastrin-releasing peptide-containing cells do not express detectable rhythms in these key components of the molecular circadian clock. The results support the view that the mammalian SCN is composed of functionally distinct cell groups, of which some are light induced and others are rhythmic with respect to dock gene expression. Furthermore, the findings suggest that gastrin-releasing peptide is a potential mediator of intercellular communication between light-induced and oscillator cells within the SCN. [ABSTRACT FROM AUTHOR]

It is well known that light resets the circadian clock only at specific times of day. The mechanisms mediating such gating of environmental input to the CNS are not well understood. We show that calbindin[subD28K] (CalB)-containing cells of the suprachiasmatic nucleus (SCN), which are directly retinorecipient, gate photic entrainment of cellular circadian oscillators and thereby determine the timing of locomotor rhythmicity. Specifically, we demonstrate a circadian rhythm of subcellular localization of CalB: whereas the protein is detected at all times in the cytoplasm, it is low or absent in the nucleus during the night. Under normal circumstances, light-induced behavioral phase shifts and Period (Per) gene expression in the SCN occur only during the subjective night. Surprisingly, both behavioral phase shifts and light-induced Per are blocked during the subjective night and enhanced during the subjective day after administration of CalB antisense oligodeoxynucleotides. These results suggest a cellular basis for temporal gating of photic input to the circadian clock. [ABSTRACT FROM AUTHOR]

Deals with a study which examined the cellular location and rhythmic expression of Period 1 circadian clock gene in the retina of a transgenic mouse. Materials and methods; Results; Discussion.

Period1 (Per1) is one of several clock genes driving the oscillatory mechanisms that mediate circadian rhythmicity. Per1 mRNA and protein are highly expressed in the suprachiasmatic nuclei, which contain oscillator cells that drive circadian rhythmicity in physiological and behavioral responses. We examined a transgenic mouse in which degradable green fluorescent protein (GFP) is driven by the mPer1 gene promoter. This mouse expresses precise free-running rhythms and characteristic light induced phase shifts. GFP protein (reporting Per1 mRNA) is expressed rhythmically as measured by either fluorescence or immunocytochemistry. In addition the animals show predicted rhythms of Per1 mRNA, PER1 and PER2 proteins. The localization of GFP overlaps with that of Per1 mRNA, PER1 and PER2 proteins. Together, these results suggest that GFP reports rhythmic Per1 expression. A surprising finding is that, at their peak expression time GFP, Per1 mRNA, PER1 and PER2 proteins are absent or not detectable in a subpopulation of SCN cells located in the core region of the nucleus. [Copyright &y& Elsevier]

Presents the results of a study in which the transplanted suprachiasmatic nuclei (SCN) of hamsters were able to sustain circadian activity rhythms by means of a diffusible signal. The similarity to the neural pacemakers of Drosophilia and silkmoths.

Examines the recovery of circadian locomotor rhythms in animals implanted with small grafts harvested by taking micropunches from vibratome-sectioned brain slices. Derivation of the micropunches from the suprachiasmatic nuclei, the subparaventricular zone and the supraoptic nucleus.

Androgens act widely in the body in both central and peripheral sites. Prior studies indicate that in the mouse, suprachiasmatic nucleus (SCN) cells bear androgen receptors (ARs). The SCN of the hypothalamus in mammals is the locus of a brain clock that regulates circadian rhythms in physiology and behavior. Gonadectomy results in reduced AR expression in the SCN and in marked lengthening of the period of free-running activity rhythms. Both responses are restored by systemic administration of androgens, but the site of action remains unknown. Our goal was to determine whether intracranial androgen implants targeted to the SCN are sufficient to restore the characteristic free-running period in gonadectomized male mice. The results indicate that hypothalamic implants of testosterone propionate in or very near the SCN produce both anatomical and behavioral effects, namely increased AR expression in the SCN and restored period of free-running locomotor activity. The effect of the implant on the period of the free-running locomotor rhythm is positively correlated with the amount of AR expression in the SCN. There is no such correlation of period change with amount of AR expression in other brain regions examined, namely the preoptic area, bed nucleus of the stria terminalis and premammillary nucleus. We conclude that the SCN is the site of action of androgen effects on the period of circadian activity rhythmicity. [ABSTRACT FROM AUTHOR]

• Stem cell proteins MSI2 and OCT4 are expressed in SCN neurons and glial cells. • Cytoskeletal components SOX2, GFAP, vimentin, and nestin are co-expressed in SCN. • SCN astrocytes may promote the stem-like state through Notch and MSI2 signaling. • Circadian rhythms continue while SCN stem-like cells proliferate in vitro. • MSI2 and similar RNA-binding proteins may regulate stem-like circadian clocks cells. The suprachiasmatic nucleus (SCN) of the mammalian hypothalamus contains the master circadian clock of the body and an unusually large number of cells expressing stem cell-related proteins. These seemingly undifferentiated cells may serve in entrainment of the SCN circadian clock to light cycles or allow it to undergo neural plasticity important for modifying its rhythmic output signals. These cells may also proliferate and differentiate into neurons or glia in response to episodic stimuli or developmental events requiring alterations in the SCN's control of physiology and behavior. To characterize expression of stem cell related proteins in the SCN and the effects of stem-like cells on circadian rhythms. Explant cultures of mouse SCN were maintained in medium designed to promote survival and growth of stem cells but not neuronal cells. Several stem cell marker proteins including SRY-box containing gene 2 (SOX2), nestin, vimentin, octamer-binding protein 4 (OCT4), and Musashi RNA-binding protein 2 (MSI2) were identified by immunocytochemistry in histological sections from adult mouse SCN and in cultures of microdissected SCN. A bioinformatics analysis located potential SCN targets of MSI2 and related RNA-binding proteins. Cells expressing stem cell markers proliferated in culture. Immunostained brain sections and bioinformatics supported the view that MSI2 regulates immature properties of SCN neurons, potentially providing flexibility in SCN neural circuits. Explant cultures had ongoing mitotic activity, indicated by proliferating-cell nuclear antigen, and extensive cell loss shown by propidium iodide staining. Cells positive for vasoactive intestinal polypeptide (VIP) that are highly enriched in the SCN were diminished in explant cultures. Despite neuronal cell loss, tissue remained viable for over 7 weeks in culture, as shown by bioluminescence imaging of explants prepared from SCN of Per1::luc transgenic mice. The circadian rhythm in SCN gene expression persisted in brain slice cultures in stem cell medium. Prominent, widespread expression of RNA-binding protein MSI2 supported the importance of posttranscriptional regulation in SCN functions and provided further evidence of stem-like cells. The results show that the SCN retains properties of immature neurons and these properties persist in culture conditions suitable for stem cells, where the SCN stem-like cells also proliferate. These properties may allow adaptive circadian rhythm adjustments. Further exploration should examine stem-like cells of the SCN in vivo, how they may affect circadian rhythms, and whether MSI2 serves as a master regulator of SCN stem-like properties. [ABSTRACT FROM AUTHOR]

Abstract: Despite the importance of learning and circadian rhythms to feeding, there has been relatively little effort to integrate these separate lines of research. In this review, we focus on how light and food entrainable oscillators contribute to the anticipation of food. In particular, we examine the evidence for temporal conditioning of food entrainable oscillators throughout the body. The evidence suggests a shift away from previous notions of a single locus or neural network of food entrainable oscillators to a distributed system involving dynamic feedback among cells of the body and brain. Several recent advances, including documentation of peroxiredoxin metabolic circadian oscillation and anticipatory behavior in the absence of a central nervous system, support the possibility of conditioned signals from the periphery in determining anticipatory behavior. Individuals learn to detect changes in internal and external signals that occur as a consequence of the brain and body preparing for an impending meal. Cues temporally near and far from actual energy content can then be used to optimize responses to temporally predictable and unpredictable cues in the environment. [Copyright &y& Elsevier]