An intracerebroventricular injection of amyloid-beta peptide (1-42) aggregates modifies daily temporal organization of clock factors expression, protein carbonyls and antioxidant enzymes in the rat hippocampus.

Alzheimer disease (AD) is the most frequent form of dementia in the elderly. It is characterized by the deterioration of memory and learning. The histopathological hallmarks of AD include the presence of extracellular deposits of amyloid beta peptide, intracellular neurofibrillary tangles, neuron and synapse loss, in the brain, including the hippocampus. Accumulation of Aß peptide causes an increase in intracellular reactive oxygen species (ROS) and free radicals associated to a deficient antioxidant defense system. Besides oxidative stress and cognitive deficit, AD patients show alterations in their circadian rhythms. The objective of this work was to investigate the effects of an intracerebroventricular injection of amyloid beta peptide Aß(1-42) aggregates on temporal patterns of protein oxidation, antioxidant enzymes and clock factors in the rat hippocampus. Four-month-old male Holtzman rats divided into the groups control (CO) and Aß-injected (Aß), were maintained under 12 h-light12h-dark conditions and received water and food ad-libitum. Hippocampus samples were isolated every 6 h during a 24 h period. Our results showed daily patterns of protein carbonyls, catalase (CAT) and glutathione peroxidase (GPx) expression and activity, as well as Rorα and Rev-erbß mRNA, in the rat hippocampus. Interestingly, an intracerebroventricular injection of Aß aggregates modified daily oscillation of protein carbonyls levels, phase-shifted daily rhythms of clock genes and had a differential effect on the daily expression and activity of CAT and GPx. Thus, Aß aggregates might affect clock-mediated transcriptional regulation of antioxidant enzymes, by affecting the formation of BMAL1:CLOCK heterodimer, probably, as a consequence of the alteration of the redox state observed in rats injected with Aß.

Oleic acid restores the rhythmicity of the disrupted circadian rhythm found in gastrointestinal explants from patients with morbid obesity.

BACKGROUND & AIMS: We investigated whether oleic acid (OA), one of the main components of the Mediterranean diet, participates in the regulation of the intestinal circadian rhythm in patients with morbid obesity. METHODS: Stomach and jejunum explants from patients with morbid obesity were incubated with oleic acid to analyze the regulation of clock genes. RESULTS: Stomach explants showed an altered circadian rhythm in CLOCK, BMAL1, REVERBα, CRY1, and CRY2, and an absence in PER1, PER2, PER3 and ghrelin (p > 0.05). OA led to the emergence of rhythmicity in PER1, PER2, PER3 and ghrelin (p < 0.05). Jejunum explants showed an altered circadian rhythm in CLOCK, BMAL1, PER1 and PER3, and an absence in PER2, REVERBα, CRY1, CRY2 and GLP1 (p > 0.05). OA led to the emergence of rhythmicity in PER2, REVERBα, CRY1 and GLP1 (p < 0.05), but not in CRY2 (p > 0.05). OA restored the rhythmicity of acrophase and increased the amplitude for most of the genes studied in stomach and jejunum explants. OA placed PER1, PER2, PER3, REVERBα, CRY1 and CRY2 in antiphase with regard to CLOCK and BMAL1. CONCLUSIONS: There is an alteration in circadian rhythm in stomach and jejunum explants in morbid obesity. OA restored the rhythmicity of the genes related with circadian rhythm, ghrelin and GLP1, although with slight differences between tissues, which could determine a different behaviour of the explants from jejunum and stomach in obesity.

Rhythmic Release of Corticosterone Induces Circadian Clock Gene Expression in the Cerebellum.

Neurons of the cerebellar cortex contain a circadian oscillator, with circadian expression of clock genes being controlled by the master clock of the suprachiasmatic nucleus (SCN). However, the signaling pathway connecting the SCN to the cerebellum is unknown. Glucocorticoids exhibit a prominent SCN-dependent circadian rhythm, and high levels of the glucocorticoid receptor have been reported in the cerebellar cortex; we therefore hypothesized that glucocorticoids may control the rhythmic expression of clock genes in the cerebellar cortex. We here applied a novel methodology by combining the electrolytic lesion of the SCN with implantation of a micropump programmed to release corticosterone in a circadian manner mimicking the endogenous hormone profile. By use of this approach, we were able to restore the corticosterone rhythm in SCN-lesioned male rats. Clock gene expression in the cerebellum was abolished in rats with a lesioned SCN, but exogenous corticosterone restored the daily rhythm in clock gene expression in the cerebellar cortex, as revealed by quantitative real-time PCR and radiochemical in situ hybridization for the detection of the core clock genes Per1, Per2, and Arntl. On the contrary, exogenous hormone did not restore circadian rhythms in body temperature and running activity. RNAscope in situ hybridization further revealed that the glucocorticoid receptor colocalizes with clock gene products in cells of the cerebellar cortex, suggesting that corticosterone exerts its actions by binding directly to receptors in neurons of the cerebellum. However, rhythmic clock gene expression in the cerebellum was also detectable in adrenalectomized rats, indicating that additional control mechanisms exist. These data show that the cerebellar circadian oscillator is influenced by SCN-dependent rhythmic release of corticosterone.

Suppressive effect of ghrelin on nicotine-induced clock gene expression in the mouse pancreas.

Those who smoke nicotine-based cigarettes have elevated plasma levels of ghrelin, a hormone secreted from the stomach. Ghrelin has various physiological functions and has recently been shown to be involved in regulating biological rhythms. Therefore, in this study, in order to clarify the significance of the plasma ghrelin increase in smokers, we sought to clarify how nicotine and ghrelin affect the expression dynamics of clock genes using a mouse model. A single dose of nicotine administered intraperitoneally increased plasma ghrelin concentrations transiently, whereas continuous administration of nicotine with an osmotic minipump did not induce any change in the plasma ghrelin concentration. Single administration of nicotine resulted in a transient increase in ghrelin gene expression in the pancreas but not in the stomach, which is the major producer of ghrelin. In addition, in the pancreas, the expression of clock genes was also increased temporarily. Therefore, in order to clarify the interaction between nicotine-induced ghrelin gene expression and clock gene expression in the pancreas, nicotine was administered to ghrelin gene-deficient mice. Administration of nicotine to ghrelin-gene deficient mice increased clock gene expression in the pancreas. However, upon nicotine administration to mice pretreated with octanoate to upregulate ghrelin activity, expression levels of nicotine-inducible clock genes in the pancreas were virtually the same as those in mice not administered nicotine. Thus, our findings indicate that pancreatic ghrelin may suppress nicotine-induced clock gene expression in the pancreas.

Cannabidiol affects circadian clock core complex and its regulation in microglia cells.

Cannabis is often used by consumers for sleep disorders. Studies show that circadian rhythm could be affected by a misuse of cannabis. Recent research has connected the role of microglial cells with psychiatric disorders such as substance abuse. The aim was to show the effect of two major components of cannabis on circadian genes regulation in microglial cells. In BV-2 microglial cells, cannabidiol (CBD) induces a deregulation of circadian genes with (P-value = 0.039) or without (P-value = 0.0015) lipopolisaccharides stimulation. CBD up regulated Arntl (P = 9.72E-5) and down regulated Clock (P = 0.0034) in BV-2 cells. Temporal expression of Arntl (light and dark P = 0.0054) and Clock (light and dark P = 0.047) was confirmed to have 24 hours light and dark rhythmic regulation in dissected suprachiasmatic nucleus as well as of Cb1 cannabinoid receptor (light and dark P = 0.019). In BV-2 microglia cells, CBD also up regulated CRY2 (P = 0.0473) and PER1 (P = 0.0131). Other nuclear molecules show a deregulation of circadian rhythm in microglial cells by CBD, such as RORA, RevErbα, RORB, CREBBP, AFT4, AFT5 and NFIL3. Our study suggests that circadian rhythm in microglial cells is deregulated by CBD but not by THC. It is consistent with clinical observations of the use of therapeutic cannabis to treat insomnia.

Naloxone precipitated morphine withdrawal and clock genes expression in striatum: A comparative study in three different protocols for the development of morphine dependence.

The present study has been designed to do a comparative study on the morphine treatment protocols for the development of morphine dependence. We have selected three different previously reported chronic morphine treatment protocols where mice were treated with different doses of morphine for different time intervals of varied number of days to develop morphine dependence. At first, animals were divided into four groups: control or saline treated and three morphine treated groups. Then we determined the differences in naloxone precipitated withdrawal behaviors and checked the expression level of the circadian clock genes in striatum. In addition we examined the level of pERK in brain tissues from cortex, hippocampus and striatum regions. Our studies showed differences in the severity of naloxone precipitated withdrawal behaviors in the three protocols. A significant increase of Period 2 gene was found in one of the morphine treated group which was absent in the animals subjected to other morphine dose regimen. Increased pERK was observed in the hippocampus of two morphine treated groups. These results support that the differences in the level of morphine dependence may not only reflect in somatic withdrawal behaviors but also have an impact on both naloxone precipitated ERK phosphorylation and clock gene expressions. Hence it can be stated that choice of chronic morphine treatment protocol may influence the research outcomes.

Historia de los terrenos del Hospital Clínico y la Facultad de Medicina de la Universidad de Chile / History of the University of Chile Faculty of Medicine and clinical hospital location

The history of the location of the University of Chile Faculty of Medicine North Campus is derived from a farm of Pedro de Valdivia founder of the city of Santiago de la Nueva Extremadura and governor of the “Reyno de Chile”. This work narrates succinctly the history of this particular location from the Spanish Conquest period to present days.

Ketamine influences CLOCK:BMAL1 function leading to altered circadian gene expression.

Major mood disorders have been linked to abnormalities in circadian rhythms, leading to disturbances in sleep, mood, temperature, and hormonal levels. We provide evidence that ketamine, a drug with rapid antidepressant effects, influences the function of the circadian molecular machinery. Ketamine modulates CLOCK:BMAL1-mediated transcriptional activation when these regulators are ectopically expressed in NG108-15 neuronal cells. Inhibition occurs in a dose-dependent manner and is attenuated after treatment with the GSK3ß antagonist SB21673. We analyzed the effect of ketamine on circadian gene expression and observed a dose-dependent reduction in the amplitude of circadian transcription of the Bmal1, Per2, and Cry1 genes. Finally, chromatin-immunoprecipitation analyses revealed that ketamine altered the recruitment of the CLOCK:BMAL1 complex on circadian promoters in a time-dependent manner. Our results reveal a yet unsuspected molecular mode of action of ketamine and thereby may suggest possible pharmacological antidepressant strategies.

Reduced expression of circadian clock genes in male alcoholic patients.

BACKGROUND: There are clear interactions between chronic alcohol consumption and circadian rhythmicity that is regulated by several circadian clock genes. The altered expressions of these genes have been mainly described in animals. The mammalian master clock in the suprachiasmatic nuclei orchestrates the biological rhythms in peripheral tissues. As peripheral blood mononuclear cells (PBMCs) are the most accessible tissue clinically, we assessed the mRNA levels of these genes in patients with alcohol dependence (AD) undergoing alcohol-withdrawal (AW) treatment. METHODS: Twenty-two male patients fulfilled the DSM-IV diagnostic criteria of AD, and 12 comparison healthy control subjects were recruited. The patients with AD were further divided by the presence of delirium tremens (DTs), the most severe form of AW syndrome, into DT group and non-DT group. All the participants received blood withdrawal at 9 am, while the patients with AD had blood collection twice: on the next morning of admission (baseline) and on the seventh day. PBMCs were isolated from whole blood, and the mRNA expression profiles of hClock1, hBmal1, hPer1, hPer2, hCry1, and hCry2 were determined by quantitative real-time PCR. RESULTS: The baseline mRNA levels of the target circadian clock genes were markedly lower in patients with AD than in control subjects. After 1 week of alcohol detoxification, there were very limited restorations of discrete circadian gene expressions. DT group did not differ in the expression patterns of circadian clock genes from non-DT group. CONCLUSIONS: This is the first study demonstrating the overall lowering of circadian clock genes among patients with AD. The expression pattern is comparable between patients with and without DTs. Although preliminary with data at only one single time point, the observation of strikingly reduced mRNA levels supports the association between circadian clock gene dysregulation and chronic alcohol intake.