Isolating neural correlates of the pacemaker for food anticipation.

Mice fed a single daily meal at intervals within the circadian range exhibit food anticipatory activity. Previous investigations strongly suggest that this behaviour is regulated by a circadian pacemaker entrained to the timing of fasting/refeeding. The neural correlate(s) of this pacemaker, the food entrainable oscillator (FEO), whether found in a neural network or a single locus, remain unknown. This study used a canonical property of circadian pacemakers, the ability to continue oscillating after removal of the entraining stimulus, to isolate activation within the neural correlates of food entrainable oscillator from all other mechanisms driving food anticipatory activity. It was hypothesized that continued anticipatory activation of central nuclei, after restricted feeding and a return to ad libitum feeding, would elucidate a neural representation of the signaling circuits responsible for the timekeeping component of the food entrainable oscillator. Animals were entrained to a temporally constrained meal then placed back on ad libitum feeding for several days until food anticipatory activity was abolished. Activation of nuclei throughout the brain was quantified using stereological analysis of c-FOS expressing cells and compared against both ad libitum fed and food entrained controls. Several hypothalamic and brainstem nuclei remained activated at the previous time of food anticipation, implicating them in the timekeeping mechanism necessary to track previous meal presentation. This study also provides a proof of concept for an experimental paradigm useful to further investigate the anatomical and molecular substrates of the FEO.

Circadian clock gene expression in brain regions of Alzheimer 's disease patients and control subjects.

Circadian oscillators have been observed throughout the rodent brain. In the human brain, rhythmic expression of clock genes has been reported only in the pineal gland, and little is known about their expression in other regions. The investigators sought to determine whether clock gene expression could be detected and whether it varies as a function of time of day in the bed nucleus of the stria terminalis (BNST) and cingulate cortex, areas known to be involved in decision making and motivated behaviors, as well as in the pineal gland, in the brains of Alzheimer's disease (AD) patients and aged controls. Relative expression levels of PERIOD1 (PER1 ), PERIOD2 (PER2), and Brain and muscle Arnt-like protein-1 (BMAL1) were detected by quantitative PCR in all 3 brain regions. A harmonic regression model revealed significant 24-h rhythms of PER1 in the BNST of AD subjects. A significant rhythm of PER2 was found in the cingulate cortex and BNST of control subjects and in all 3 regions of AD patients. In controls, BMAL1 did not show a diurnal rhythm in the cingulate cortex but significantly varied with time of death in the pineal and BNST and in all 3 regions for AD patients. Notable differences in the phase of clock gene rhythms and phase relationships between genes and regions were observed in the brains of AD compared to those of controls. These results indicate the presence of multiple circadian oscillators in the human brain and suggest altered synchronization among these oscillators in the brain of AD patients.

Circadian rhythms and clock genes in psychotic disorders.

Numerous lines of evidence suggest that a disordered circadian system contributes to the etiology and symptomatology of major psychiatric disorders. Sleep disturbances, particularly rapid eye movement (REM) sleep, have been observed in bipolar affective disorder (BPD) and schizophrenia. Therapies aimed at altering the timing and duration of sleep and realigning circadian rhythms, including sleep scheduling, wake extension, light therapy and drug therapies that alter sleep and circadian rhythms appear beneficial for affective disorders. Interventional studies aiming to correct sleep and circadian disturbances in schizophrenia are scarce, although exogenous melatonin has been shown to improve both sleep structure and psychotic symptoms. The study of molecular clock mechanisms in psychiatric disorders is also gaining interest. Genetics studies have found associations with CLOCK, PERIOD1, PERIOD3, and TIMELESS in schizophrenia. Most research on BPD has focused on polymorphisms of CLOCK, but the lithium target GSK-3 may also be significant. New research examining the role of circadian rhythms and clock genes in major mental illness is likely to produce rapid advances in circadian-based therapeutics.

The role of circadian clock genes in mental disorders.

The study of molecular clock mechanisms in psychiatric disorders is gaining significant interest due to data suggesting that a misalignment between the endogenous circadian system and the sleep-wake cycle might contribute to the clinical status of patients suffering from a variety of psychiatric disorders. Sleep disturbances in major depressive disorder (MDD) are characterized by increased sleep latency, poorer sleep efficiency reduced latency to the first rapid eye movement (REM) sleep episode, and early-morning awakening, but there is little data to indicate a role of circadian clock genes in MDD. There is also relatively little information regarding the role of clock genes in anxiety. In contrast, a significant amount of evidence gathered in bipolar disorder (BPD) patients suggests a circadian rhythm disorder, namely an advanced circadian rhythm and state-dependent alterations of REM sleep latency. Most research on the role of clock genes in BPD has focused on polymorphisms of CLOCK, but the lithium target GSK3 may also play a significant role. A circadian phase shift is also theorized to contribute to the pathophysiology of winter seasonal affective disorder (SAD). Certain allelic combinations of NPAS2, PER3, and BMAL1 appear to contribute to the risk of SAD. In chronic schizophrenia, disturbances of sleep including insomnia and reduced sleep efficiency have been observed. Genetic studies have found associations with CLOCK, PER1, PER3, and TIMELESS. Sleep and circadian changes associated with dementia due to Alzheimer's disease suggest a functional change in the circadian master clock, which is supported by postmortem studies of clock gene expression in the brain.

From circadian clock gene expression to pathologies.

In most organisms, circadian rhythms are generated by a molecular clockwork involving so-called clock genes. These circadian clock genes participate in regulatory feedback loops, in which proteins regulate their own expression. The outcome is that ribonucleic acids (RNAs) and proteins produced from many of these genes oscillate with a circadian rhythm. Here, we describe the regulation of clock genes and proteins, as deduced from work in rodents. Furthermore, we summarize the work done on human clock genes and their expression in peripheral tissues. Importantly, the research reviewed here points to an implication of clock gene defects in circadian rhythm disorders, including the advanced and delayed sleep phase disorders. Moreover, circadian clock gene dysfunction is likely to be of importance in the development of cancer as well as various other diseases.

The central and basolateral nuclei of the amygdala exhibit opposite diurnal rhythms of expression of the clock protein Period2.

There is considerable evidence that circadian rhythms in mammals can be modulated by emotional state, but how emotional state modulates specific circadian outputs is poorly understood. We analyzed the expression of the circadian clock protein Period2 (PER2) in three regions of the limbic forebrain known to play key roles in emotional regulation, the central nucleus of the amygdala (CEA), the basolateral amygdala (BLA), and the dentate gyrus (DG). We report here that cells in all three regions exhibit daily rhythms in expression of PER2 that are under the control of the master clock, the suprachiasmatic nucleus (SCN). The rhythm in the CEA and the rhythms in the BLA and DG are diametrically opposite in phase and are differentially affected by adrenalectomy. Adrenalectomy completely abolished the PER2 rhythm in the CEA but had no effect on the PER2 rhythms in the BLA and DG. We previously reported a rhythm in PER2 expression in the oval nucleus of the bed nucleus of the stria terminalis that is identical in phase and sensitivity to adrenalectomy to that found in the CEA. Together, these findings show that key structures of the limbic forebrain exhibit daily oscillations in clock gene expression that are controlled not only by input from the SCN but, importantly, by hormonal and neurochemical changes that normally accompany motivational and emotional states. Thus, cells within these areas are strategically positioned to integrate the inputs from the SCN and emotional states to modulate circadian rhythms downstream from the SCN clock.

A circadian rhythm in the expression of PERIOD2 protein reveals a novel SCN-controlled oscillator in the oval nucleus of the bed nucleus of the stria terminalis.

Circadian rhythms in mammals are regulated not only globally by the master clock in the suprachiasmatic nucleus (SCN), but also locally by widely distributed populations of clock cells in the brain and periphery that control tissue-specific rhythmic outputs. Here we show that the oval nucleus of the bed nucleus of the stria terminalis (BNST-OV) exhibits a robust circadian rhythm in expression of the Period2 (PER2) clock protein. PER2 expression is rhythmic in the BNST-OV in rats housed under a light/dark cycle or in constant darkness, in blind rats, and in mice, and is in perfect synchrony with the PER2 rhythm of the SCN. Constant light or bilateral SCN lesions abolish the rhythm of PER2 in the BNST-OV. Large abrupt shifts in the light schedule transiently uncouple the BNST-OV rhythm from that of the SCN. Re-entrainment of the PER2 rhythm is faster in the SCN than in the BNST-OV, and it is faster after a delay than an advance shift. Bilateral adrenalectomy blunts the PER2 rhythm in the BNST-OV. Thus, the BNST-OV contains circadian clock cells that normally oscillate in synchrony with the SCN, but these cells appear to require both input from the SCN and circulating glucocorticoids to maintain their circadian oscillation. Taken together with what is known about the functional organization of the connections of the BNST-OV with systems of the brain involved in stress and motivational processes, these findings place BNST-OV oscillators in a position to influence specific physiological and behavioral rhythms downstream from the SCN clock.

Melanopsin in the circadian timing system.

In mammals, circadian rhythms are generated by a light-entrainable oscillator located in the hypothalamic suprachiasmatic nucleus (SCN). Light signals reach the SCN via a dedicated retinal pathway, the retinohypothalamic tract (RHT). One question that continues to elude scientists is whether the circadian system has its own dedicated photoreceptor or photoreceptors. It is well established that conventional photoreceptors, rods and cones, are not required for circadian photoreception, suggesting that the inner retinal layer might contribute to circadian photoreception. Melanopsin, a novel photo pigment expressed in retinal ganglion cells (RGCs), has been proposed recently as a candidate circadian photoreceptor. Melanopsin-containing RGCs are intrinsically photosensitive, form part of the RHT, and contain neurotransmitters known to play a critical role in the circadian response to light. Furthermore, melanopsin-containing RGCs do not depend on inputs from rods and cones to transmit light signals to the SCN. However, based on a review of the available information about melanopsin and on new data from our laboratory, we propose that melanopsin, in itself, is not necessary for circadian photoreception. In fact, it appears that of the known photoreceptor systems, none, in and of itself, is necessary for circadian photoreception. Instead, it appears that within the photoreceptive systems there is some degree of redundancy, each contributing in some way to photic entrainment.