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1.
Cell Mol Neurobiol ; 44(1): 51, 2024 Jun 22.
Artículo en Inglés | MEDLINE | ID: mdl-38907776

RESUMEN

The circadian system is a conserved time-keeping machinery that regulates a wide range of processes such as sleep/wake, feeding/fasting, and activity/rest cycles to coordinate behavior and physiology. Circadian disruption can be a contributing factor in the development of metabolic diseases, inflammatory disorders, and higher risk of cancer. Glioblastoma (GBM) is a highly aggressive grade 4 brain tumor that is resistant to conventional therapies and has a poor prognosis after diagnosis, with a median survival of only 12-15 months. GBM cells kept in culture were shown to contain a functional circadian oscillator. In seeking more efficient therapies with lower side effects, we evaluated the pharmacological modulation of the circadian clock by targeting the cytosolic kinases glycogen synthase kinase-3 (GSK-3) and casein kinase 1 ε/δ (CK1ε/δ) with specific inhibitors (CHIR99021 and PF670462, respectively), the cryptochrome protein stabilizer (KL001), or circadian disruption after Per2 knockdown expression in GBM-derived cells. CHIR99021-treated cells had a significant effect on cell viability, clock protein expression, migration, and cell cycle distribution. Moreover, cultures exhibited higher levels of reactive oxygen species and alterations in lipid droplet content after GSK-3 inhibition compared to control cells. The combined treatment of CHIR99021 with temozolomide was found to improve the effect on cell viability compared to temozolomide therapy alone. Per2 disruption affected both GBM migration and cell cycle progression. Overall, our results suggest that pharmacological modulation or molecular clock disruption severely affects GBM cell biology.


Asunto(s)
Neoplasias Encefálicas , Glioblastoma , Glioblastoma/patología , Glioblastoma/metabolismo , Glioblastoma/tratamiento farmacológico , Humanos , Línea Celular Tumoral , Neoplasias Encefálicas/patología , Neoplasias Encefálicas/metabolismo , Neoplasias Encefálicas/tratamiento farmacológico , Piridinas/farmacología , Supervivencia Celular/efectos de los fármacos , Citosol/metabolismo , Citosol/efectos de los fármacos , Glucógeno Sintasa Quinasa 3/metabolismo , Pirimidinas/farmacología , Movimiento Celular/efectos de los fármacos , Relojes Circadianos/efectos de los fármacos , Relojes Circadianos/fisiología , Proteínas CLOCK/metabolismo , Proteínas CLOCK/genética , Proteínas Circadianas Period/metabolismo , Proteínas Circadianas Period/genética , Especies Reactivas de Oxígeno/metabolismo
2.
Neurobiol Aging ; 126: 77-90, 2023 06.
Artículo en Inglés | MEDLINE | ID: mdl-36958103

RESUMEN

Aging impairs both circadian rhythms and memory, though the relationship between these impairments is not fully understood. Circadian rhythms are largely dictated by clock genes within the body's central pacemaker, the suprachiasmatic nucleus (SCN), though these genes are also expressed in local clocks throughout the body. As circadian rhythms can directly affect memory performance, one possibility is that memory deficits observed with age are downstream of global circadian rhythm disruptions stemming from the SCN. Here, we demonstrate that expression of clock gene Period1 within a memory-relevant cortical structure, the retrosplenial cortex (RSC), is necessary for incidental learning, and that age-related disruption of Period1 within the RSC-but not necessarily the SCN-contributes to cognitive decline. These data expand the known functions of clock genes beyond maintaining circadian rhythms and suggests that age-associated changes in clock gene expression modulates circadian rhythms and memory performance in a brain region-dependent manner.


Asunto(s)
Relojes Circadianos , Giro del Cíngulo , Ratones , Animales , Masculino , Giro del Cíngulo/metabolismo , Núcleo Supraquiasmático/metabolismo , Ritmo Circadiano/genética , Encéfalo/metabolismo , Factores de Transcripción/metabolismo , Envejecimiento/genética , Relojes Circadianos/genética , Proteínas Circadianas Period/genética , Proteínas Circadianas Period/metabolismo
3.
Biomolecules ; 12(7)2022 06 25.
Artículo en Inglés | MEDLINE | ID: mdl-35883448

RESUMEN

The molecular circadian clock is based on a transcriptional/translational feedback loop in which the stability and half-life of circadian proteins is of importance. Cysteine residues of proteins are subject to several redox reactions leading to S-thiolation and disulfide bond formation, altering protein stability and function. In this work, the ability of the circadian protein period 2 (PER2) to undergo oxidation of cysteine thiols was investigated in HEK-293T cells. PER2 includes accessible cysteines susceptible to oxidation by nitroso cysteine (CysNO), altering its stability by decreasing its monomer form and subsequently increasing PER2 homodimers and multimers. These changes were reversed by treatment with 2-mercaptoethanol and partially mimicked by hydrogen peroxide. These results suggest that cysteine oxidation can prompt PER2 homodimer and multimer formation in vitro, likely by S-nitrosation and disulphide bond formation. These kinds of post-translational modifications of PER2 could be part of the redox regulation of the molecular circadian clock.


Asunto(s)
Relojes Circadianos , Proteínas Circadianas Period , Ritmo Circadiano/fisiología , Cisteína/metabolismo , Dimerización , Oxidación-Reducción , Proteínas Circadianas Period/química , Proteínas Circadianas Period/genética , Proteínas Circadianas Period/metabolismo , Proteínas/metabolismo
4.
Front Endocrinol (Lausanne) ; 12: 652733, 2021.
Artículo en Inglés | MEDLINE | ID: mdl-34504470

RESUMEN

The important involvement of the suprachiasmatic nucleus (SCN) and the activity of vasopressinergic neurons in maintaining the rhythmicity of the female reproductive system depends on the mRNA transcription-translation feedback loops. Therefore, circadian clock function, like most physiological processes, is involved in the events that determine reproductive aging. This study describes the change of mRNA expression of clock genes, Per2, Bmal1, and Rev-erbα, in the hypothalamus-pituitary-gonadal axis (HPG) of female rats with regular cycle (RC) and irregular cycle (IC), and the vasopressinergic neurons activity in the SCN and kisspeptin neurons in the arcuate nucleus (ARC) of these animals. Results for gonadotropins and the cFos/AVP-ir neurons in the SCN of IC were higher, but kisspeptin-ir was minor. Change in the temporal synchrony of the clock system in the HPG axis, during the period prior to the cessation of ovulatory cycles, was identified. The analysis of mRNA for Per2, Bmal1, and Rev-erbα in the reproductive axis of adult female rodents shows that the regularity of the estrous cycle is guaranteed by alternation in the amount of expression of Bmal1 and Per2, and Rev-erbα and Bmal1 between light and dark phases, which ceases to occur and contributes to determining reproductive senescence. These results showed that the desynchronization between the central and peripheral circadian clocks contributes to the irregularity of reproductive events. We suggest that the feedback loops of clock genes on the HPG axis modulate the spontaneous transition from regular to irregular cycle and to acyclicity in female rodents.


Asunto(s)
Envejecimiento , Ritmo Circadiano , Gónadas/metabolismo , Sistema Hipotálamo-Hipofisario/metabolismo , ARN Mensajero/metabolismo , Núcleo Supraquiasmático/metabolismo , Vasopresinas/farmacología , Factores de Transcripción ARNTL/genética , Factores de Transcripción ARNTL/metabolismo , Animales , Relojes Circadianos , Femenino , Gónadas/efectos de los fármacos , Sistema Hipotálamo-Hipofisario/efectos de los fármacos , Miembro 1 del Grupo D de la Subfamilia 1 de Receptores Nucleares/genética , Miembro 1 del Grupo D de la Subfamilia 1 de Receptores Nucleares/metabolismo , Proteínas Circadianas Period/genética , Proteínas Circadianas Period/metabolismo , ARN Mensajero/genética , Ratas , Ratas Wistar , Núcleo Supraquiasmático/efectos de los fármacos
5.
Obes Res Clin Pract ; 15(4): 334-338, 2021.
Artículo en Inglés | MEDLINE | ID: mdl-34215556

RESUMEN

BACKGROUND: Obesity has reached epidemic proportions worldwide, affecting life quality and span. Susceptibility to obesity is partly mediated by genetic differences. Indeed, several genes from the clock gene family have already been shown to be intimately associated with obesity in diverse ethnic groups. In the present study, an association between BMI and the rs707467, rs228697 and rs228729 PER3 (Period Circadian Clock 3) polymorphisms in subjects with class II (BMI ≥ 35.0-39.9 kg/m2) and class III obesity (>40 kg/m2, extreme obesity) were carried out using TaqMan real-time PCR. Overall, 259 Brazilian adults were genotyped, of whom 122 had class II or III obesity (BMI ≥ 35.0 kg/m2) and 137 were controls having normal weight (BMI > 18.5 and <24.9 kg/m2). RESULTS: PER3 tag SNP (rs228729) shows a significant association with extreme obesity (1000 permutation p = 0.03 and p = 0.04), for genotype and allele frequency respectively) and a haplotype among the three assessed SNPs (alleles G/T/A, rs228697, rs228729, and rs707467, respectively, 1000 permutation p = 0.03) was significantly more prevalent in the group with obesity. CONCLUSION: This exploratory association study suggests that PER3 rs228729 may be associated with extreme obesity in Brazilian adults, however, replication is needed.


Asunto(s)
Obesidad Mórbida/genética , Proteínas Circadianas Period , Polimorfismo de Nucleótido Simple , Adulto , Alelos , Proteínas CLOCK/genética , Ritmo Circadiano , Frecuencia de los Genes , Genotipo , Humanos , Proteínas Circadianas Period/genética , Proteínas Circadianas Period/metabolismo
6.
Curr Biol ; 30(24): 5040-5048.e5, 2020 12 21.
Artículo en Inglés | MEDLINE | ID: mdl-33065014

RESUMEN

Behavioral outputs arise as a result of highly regulated yet flexible communication among neurons. The Drosophila circadian network includes 150 neurons that dictate the temporal organization of locomotor activity; under light-dark (LD) conditions, flies display a robust bimodal pattern. The pigment-dispersing factor (PDF)-positive small ventral lateral neurons (sLNv) have been linked to the generation of the morning activity peak (the "M cells"), whereas the Cryptochrome (CRY)-positive dorsal lateral neurons (LNds) and the PDF-negative sLNv are necessary for the evening activity peak (the "E cells") [1, 2]. While each group directly controls locomotor output pathways [3], an interplay between them along with a third dorsal cluster (the DN1ps) is necessary for the correct timing of each peak and for adjusting behavior to changes in the environment [4-7]. M cells set the phase of roughly half of the circadian neurons (including the E cells) through PDF [5, 8-10]. Here, we show the existence of synaptic input provided by the evening oscillator onto the M cells. Both structural and functional approaches revealed that E-to-M cell connectivity changes across the day, with higher excitatory input taking place before the day-to-night transition. We identified two different neurotransmitters, acetylcholine and glutamate, released by E cells that are relevant for robust circadian output. Indeed, we show that acetylcholine is responsible for the excitatory input from E cells to M cells, which show preferential responsiveness to acetylcholine during the evening. Our findings provide evidence of an excitatory feedback between circadian clusters and unveil an important plastic remodeling of the E cells' synaptic connections.


Asunto(s)
Relojes Biológicos/fisiología , Drosophila melanogaster/fisiología , Locomoción/fisiología , Terminales Presinápticos/metabolismo , Acetilcolina/metabolismo , Animales , Animales Modificados Genéticamente , Ritmo Circadiano/fisiología , Proteínas de Drosophila/genética , Proteínas de Drosophila/metabolismo , Retroalimentación Fisiológica , Ácido Glutámico/metabolismo , Masculino , Proteínas Circadianas Period/genética , Proteínas Circadianas Period/metabolismo , Fotoperiodo
7.
Horm Behav ; 120: 104683, 2020 04.
Artículo en Inglés | MEDLINE | ID: mdl-31930968

RESUMEN

Circadian (~24 h) rhythms in behavior and physiological functions are under control of an endogenous circadian pacemaker in the suprachiasmatic nucleus (SCN) of the hypothalamus. The SCN directly drives some of these rhythms or serves as a coordinator of peripheral oscillators residing in other tissues and organs. Disruption of the circadian organization may contribute to disease, including stress-related disorders. Previous research indicates that the master clock in the SCN is resistant to stress, although it is unclear whether stress affects rhythmicity in other tissues, possibly mediated by glucocorticoids, released in stressful situations. In the present study, we examined the effect of uncontrollable social defeat stress and glucocorticoid hormones on the central and peripheral clocks, respectively in the SCN and liver. Transgenic PERIOD2::LUCIFERASE knock-in mice were used to assess the rhythm of the clock protein PERIOD2 (PER2) in SCN slices and liver tissue collected after 10 consecutive days of social defeat stress. The rhythmicity of PER2 expression in the SCN was not affected by stress exposure, whereas in the liver the expression showed a delayed phase in defeated compared to non-defeated control mice. In a second experiment, brain slices and liver samples were collected from transgenic mice and exposed to different doses of corticosterone. Corticosterone did not affect PER2 rhythm of the SCN samples, but caused a phase shift in PER2 expression in liver samples. This study confirms earlier findings that the SCN is resistant to stress and shows that clocks in the liver are affected by social stress, which might be due to the direct influence of glucocorticoids released from the adrenal gland.


Asunto(s)
Ritmo Circadiano/efectos de los fármacos , Ritmo Circadiano/genética , Glucocorticoides/farmacología , Hígado/metabolismo , Proteínas Circadianas Period/genética , Estrés Psicológico , Núcleo Supraquiasmático/metabolismo , Glándulas Suprarrenales/metabolismo , Animales , Encéfalo/efectos de los fármacos , Encéfalo/metabolismo , Ritmo Circadiano/fisiología , Corticosterona/metabolismo , Dominación-Subordinación , Técnicas de Sustitución del Gen , Hígado/efectos de los fármacos , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Proteínas Circadianas Period/metabolismo , Conducta Social , Estrés Psicológico/genética , Estrés Psicológico/metabolismo , Estrés Psicológico/fisiopatología , Núcleo Supraquiasmático/efectos de los fármacos
8.
Phys Rev E ; 100(2-1): 022409, 2019 Aug.
Artículo en Inglés | MEDLINE | ID: mdl-31574627

RESUMEN

Organisms have evolved in a daily cyclic environment, developing circadian cell-autonomous clocks that temporally organize a wide range of biological processes. Translation is a highly regulated process mainly associated with the activity of microRNAs (miRNAs) at the translation initiation step that impacts on the molecular circadian clock dynamics. Recently, a molecular titration mechanism was proposed to explain the interactions between some miRNAs and their target mRNAs; new evidence also indicates that regulation by miRNA is a nonlinear process such that there is a threshold level of target mRNA below which protein production is drastically repressed. These observations led us to use a theoretical model of the circadian molecular clock to study the effect of miRNA-mediated translational thresholds on the molecular clock dynamics. We model the translational threshold by introducing a phenomenological Hill equation for the kinetics of PER translation and show how the parameters associated with translation kinetics affect the period, amplitude, and time delays between clock mRNA and clock protein expression. We show that our results are useful for analyzing experiments related to the translational regulation of negative elements of transcriptional-translational feedback loops. We also provide new elements for thinking about the translational threshold as a mechanism that favors the emergence of circadian rhythmicity, the tuning of the period-delay relationship and the cell capacity to control the protein oscillation amplitude with almost negligible changes in the mRNA amplitudes.


Asunto(s)
Relojes Circadianos/genética , Modelos Genéticos , Biosíntesis de Proteínas , Cinética , Proteínas Circadianas Period/metabolismo , ARN Mensajero/genética
9.
Yale J Biol Med ; 92(2): 241-250, 2019 06.
Artículo en Inglés | MEDLINE | ID: mdl-31249485

RESUMEN

Circadian clocks drive biological rhythms in physiology and behavior, providing a selective advantage by enabling organisms to synchronize to the 24 h environmental day. This process depends on light-dark transitions as the main signal that shifts the phase of the clock. In mammals, the light input reaches the master circadian clock in the hypothalamic suprachiasmatic nucleus through glutamatergic afferents from the retina, resulting in phase-shifts of the overt rhythms which depend on the time of the day at which light is applied, leading to changes in the activity of circadian core clock genes (i.e., Per1). This circadian gating of the synchronizing effect of light is dependent on the specific activation of signal transduction pathways involving several kinases acting on protein effectors. Protein phosphorylation is also an important regulatory mechanism essential for the generation and maintenance of circadian rhythms and plays a crucial role in the degradation and the appropriate turnover of PER proteins. In this work, we review the role of the main kinases implicated in the function of the master clock, with emphasis in those involved in circadian photic entrainment.


Asunto(s)
Relojes Circadianos/fisiología , Fototransducción/fisiología , Mamíferos/fisiología , Proteínas Quinasas/metabolismo , Animales , Regulación de la Expresión Génica/efectos de la radiación , Humanos , Luz , Mamíferos/genética , Mamíferos/metabolismo , Proteínas Circadianas Period/genética , Proteínas Circadianas Period/metabolismo , Fosforilación/efectos de la radiación
10.
Addict Biol ; 24(6): 1179-1190, 2019 11.
Artículo en Inglés | MEDLINE | ID: mdl-30295391

RESUMEN

Shift-work and exposure to light at night lead to circadian disruption, which favors the use of alcohol and may be a risk factor for development of addictive behavior. This study evaluated in two experimental models of circadian disruption behavioral indicators of elevated alcohol intake and looked for ΔFosB, which is a transcription factor for neuronal plasticity in corticolimbic structures. Male Wistar rats were exposed to experimental shift-work (AR) or to constant light (LL) and were compared with a control group (LD). After 4 weeks in their corresponding conditions, control LD rats remained rhythmic, AR rats exhibited a loss of day-night patterns in the brain and the LL rats showed arrhythmicity in general activity and day-night PER1 patterns in corticolimbic structures. During 12 days of exposure to 10 percent alcohol solution, the AR group showed daily increased alcohol intake while LD and LL rats ingested similar amounts. After 72 h of alcohol deprivation, AR and LL rats increased alcohol intake in a binge-like test; this could be due not only to circadian disruption but also to stress and/or anxiety developed from the AR and LL manipulations. Associated to the increased alcohol intake, the AR and LL rats had significant accumulation of ΔFosB in the nucleus accumbens shell and decreased ΔFosB in the infralimbic cortex. Data here reported confirm that the disruption of temporal patterns favors the increased alcohol consumption and that this is associated with a differential accumulation of ΔFosB which may favor the development of addictive behavior.


Asunto(s)
Consumo de Bebidas Alcohólicas , Encéfalo/efectos de los fármacos , Depresores del Sistema Nervioso Central/farmacología , Ritmo Circadiano , Etanol/farmacología , Proteínas Proto-Oncogénicas c-fos/efectos de los fármacos , Horario de Trabajo por Turnos , Animales , Ansiedad/metabolismo , Conducta Animal , Encéfalo/metabolismo , Núcleo Amigdalino Central/efectos de los fármacos , Núcleo Amigdalino Central/metabolismo , Depresores del Sistema Nervioso Central/administración & dosificación , Corteza Cerebral/efectos de los fármacos , Corteza Cerebral/metabolismo , Etanol/administración & dosificación , Plasticidad Neuronal , Núcleo Accumbens/efectos de los fármacos , Núcleo Accumbens/metabolismo , Proteínas Circadianas Period/metabolismo , Corteza Prefrontal/efectos de los fármacos , Corteza Prefrontal/metabolismo , Proteínas Proto-Oncogénicas c-fos/metabolismo , Distribución Aleatoria , Ratas , Autoadministración , Estrés Psicológico/metabolismo , Núcleo Supraquiasmático/efectos de los fármacos , Núcleo Supraquiasmático/metabolismo
11.
Chronobiol Int ; 35(2): 147-159, 2018 02.
Artículo en Inglés | MEDLINE | ID: mdl-29111822

RESUMEN

Myocardial gene expression and metabolism fluctuate over the course of the day in association with changes in energy supply and demand. Time-of-day-dependent oscillations in myocardial processes have been linked to the intrinsic cardiomyocyte circadian clock. Triiodothyronine (T3) is an important modulator of heart metabolism and function. Recently, our group has reported time-of-day-dependent rhythms in cardiac T3 sensitivity, as well as, T3-mediated acute alterations on core clock components. Hypo and hyperthyroidism are the second most prevalent endocrine disease worldwide. Considering the importance of the cardiomyocyte circadian clock and T3 to cardiac physiology, the aim of this study was to investigate the consequences of chronic hypo and hyperthyroidism on 24-h rhythms of circadian clock genes in the heart. Hypo and hyperthyroidism was induced in rats by thyroidectomy (Tx) and i.p. injections of supraphysiological dose of T3, respectively. Here we report alterations in mRNA levels of the major core clock components (Bmal1, Per2, Nr1d1, and Rora) for both experimental conditions (with the exception of Per2 during hyperthyroid condition). Oscillations in mRNA levels of key glucose and fatty-acid metabolism genes known to be clock controlled (Pdk4, Ucp3, Acot1, and Cd36) were equally affected by the experimental conditions, especially during the hypothyroid state. These findings suggest that chronic alterations in thyroid status significantly impacts 24-h rhythms in circadian clock and metabolic genes in the heart. Whether these perturbations contribute toward the pathogenesis of cardiac dysfunction associated with hypo and hyperthyroidism requires further elucidation.


Asunto(s)
Relojes Circadianos/fisiología , Ritmo Circadiano/fisiología , Hipertiroidismo/metabolismo , Miocardio/metabolismo , Factores de Transcripción ARNTL/genética , Animales , Regulación de la Expresión Génica/fisiología , Glucosa/metabolismo , Masculino , Proteínas Circadianas Period/genética , Proteínas Circadianas Period/metabolismo , Ratas Wistar
12.
Sci Rep ; 7(1): 13977, 2017 10 25.
Artículo en Inglés | MEDLINE | ID: mdl-29070825

RESUMEN

Melanopsin (OPN4) is a photo-pigment found in a small subset of intrinsically photosensitive ganglion cells (ipRGCs) of the mammalian retina. These cells play a role in synchronizing the central circadian pacemaker to the astronomical day by conveying information about ambient light to the hypothalamic suprachiasmatic nucleus, the site of the master clock. We evaluated the effect of a heat stimulus (39.5 °C) on clock gene (Per1 and Bmal1) expression in cultured murine Melan-a melanocytes synchronized by medium changes, and in B16-F10 melanoma cells, in the presence of the selective OPN4 antagonist AA92593, or after OPN4 knockdown by small interfering RNA (siRNA). In addition, we evaluated the effects of heat shock on the localization of melanopsin by immunocytochemistry. In both cell lines melanopsin was found in a region capping the nucleus and heat shock did not affect its location. The heat-induced increase of Per1 expression was inhibited when melanopsin was pharmacologically blocked by AA92593 as well as when its protein expression was suppressed by siRNA in both Melan-a and B16-F10 cells. These data strongly suggest that melanopsin is required for thermo-reception, acting as a thermo-opsin that ultimately feeds the local circadian clock in mouse melanocytes and melanoma cells.


Asunto(s)
Proteínas CLOCK/metabolismo , Relojes Circadianos/genética , Calor , Melanocitos/metabolismo , Melanoma Experimental/genética , Proteínas Circadianas Period/metabolismo , Opsinas de Bastones/metabolismo , Animales , Proteínas CLOCK/genética , Células Cultivadas , Regulación de la Expresión Génica , Melanoma Experimental/metabolismo , Melanoma Experimental/patología , Ratones , Proteínas Circadianas Period/genética , ARN Interferente Pequeño/genética , Opsinas de Bastones/antagonistas & inhibidores , Opsinas de Bastones/genética
13.
Genetics ; 207(2): 593-607, 2017 10.
Artículo en Inglés | MEDLINE | ID: mdl-28801530

RESUMEN

Circadian clocks organize the metabolism, physiology, and behavior of organisms throughout the day-night cycle by controlling daily rhythms in gene expression at the transcriptional and post-transcriptional levels. While many transcription factors underlying circadian oscillations are known, the splicing factors that modulate these rhythms remain largely unexplored. A genome-wide assessment of the alterations of gene expression in a null mutant of the alternative splicing regulator SR-related matrix protein of 160 kDa (SRm160) revealed the extent to which alternative splicing impacts on behavior-related genes. We show that SRm160 affects gene expression in pacemaker neurons of the Drosophila brain to ensure proper oscillations of the molecular clock. A reduced level of SRm160 in adult pacemaker neurons impairs circadian rhythms in locomotor behavior, and this phenotype is caused, at least in part, by a marked reduction in period (per) levels. Moreover, rhythmic accumulation of the neuropeptide PIGMENT DISPERSING FACTOR in the dorsal projections of these neurons is abolished after SRm160 depletion. The lack of rhythmicity in SRm160-downregulated flies is reversed by a fully spliced per construct, but not by an extra copy of the endogenous locus, showing that SRm160 positively regulates per levels in a splicing-dependent manner. Our findings highlight the significant effect of alternative splicing on the nervous system and particularly on brain function in an in vivo model.


Asunto(s)
Ritmo Circadiano , Proteínas de Drosophila/genética , Drosophila melanogaster/genética , Locomoción , Factores de Empalme de ARN/metabolismo , Animales , Encéfalo/citología , Encéfalo/metabolismo , Encéfalo/fisiología , Proteínas de Drosophila/metabolismo , Drosophila melanogaster/fisiología , Neuronas/metabolismo , Proteínas Circadianas Period/genética , Proteínas Circadianas Period/metabolismo , Empalme del ARN , Factores de Empalme de ARN/genética
14.
eNeuro ; 4(2)2017.
Artículo en Inglés | MEDLINE | ID: mdl-28374011

RESUMEN

The suprachiasmatic nucleus (SCN) is generally considered the master clock, independently driving all circadian rhythms. We recently demonstrated the SCN receives metabolic and cardiovascular feedback adeptly altering its neuronal activity. In the present study, we show that microcuts effectively removing SCN-arcuate nucleus (ARC) interconnectivity in Wistar rats result in a loss of rhythmicity in locomotor activity, corticosterone levels, and body temperature in constant dark (DD) conditions. Elimination of these reciprocal connections did not affect SCN clock gene rhythmicity but did cause the ARC to desynchronize. Moreover, unilateral SCN lesions with contralateral retrochiasmatic microcuts resulted in identical arrhythmicity, proving that for the expression of physiological rhythms this reciprocal SCN-ARC interaction is essential. The unaltered SCN c-Fos expression following glucose administration in disconnected animals as compared to a significant decrease in controls demonstrates the importance of the ARC as metabolic modulator of SCN neuronal activity. Together, these results indicate that the SCN is more than an autonomous clock, and forms an essential component of a larger network controlling homeostasis. The present novel findings illustrate how an imbalance between SCN and ARC communication through circadian disruption could be involved in the etiology of metabolic disorders.


Asunto(s)
Núcleo Arqueado del Hipotálamo/fisiología , Ritmo Circadiano/fisiología , Núcleo Supraquiasmático/fisiología , Animales , Núcleo Arqueado del Hipotálamo/patología , Núcleo Arqueado del Hipotálamo/fisiopatología , Temperatura Corporal/fisiología , Corticosterona/metabolismo , Glucosa/administración & dosificación , Glucosa/metabolismo , Hígado/metabolismo , Modelos Animales , Actividad Motora/fisiología , Vías Nerviosas/fisiología , Vías Nerviosas/fisiopatología , Neuronas/metabolismo , Neuronas/patología , Proteínas Circadianas Period/metabolismo , Proteínas Proto-Oncogénicas c-fos/metabolismo , Ratas Wistar , Núcleo Supraquiasmático/patología , Núcleo Supraquiasmático/fisiopatología
15.
Endocrinology ; 157(1): 323-35, 2016 Jan.
Artículo en Inglés | MEDLINE | ID: mdl-26556532

RESUMEN

In rodents, kisspeptin neurons in the rostral periventricular area of the third ventricle (RP3V) of the preoptic area are considered to provide a major stimulatory input to the GnRH neuronal network that is responsible for triggering the preovulatory LH surge. Noradrenaline (NA) is one of the main modulators of GnRH release, and NA fibers are found in close apposition to kisspeptin neurons in the RP3V. Our objective was to interrogate the role of NA signaling in the kisspeptin control of GnRH secretion during the estradiol induced LH surge in ovariectomized rats, using prazosin, an α1-adrenergic receptor antagonist. In control rats, the estradiol-induced LH surge at 17 hours was associated with a significant increase in GnRH and kisspeptin content in the median eminence with the increase in kisspeptin preceding that of GnRH and LH. Prazosin, administered 5 and 3 hours prior to the predicted time of the LH surge truncated the LH surge and abolished the rise in GnRH and kisspeptin in the median eminence. In the preoptic area, prazosin blocked the increases in Kiss1 gene expression and kisspeptin content in association with a disruption in the expression of the clock genes, Per1 and Bmal1. Together these findings demonstrate for the first time that NA modulates kisspeptin synthesis in the RP3V through the activation of α1-adrenergic receptors prior to the initiation of the LH surge and indicate a potential role of α1-adrenergic signaling in the circadian-controlled pathway timing of the preovulatory LH surge.


Asunto(s)
Regulación de la Expresión Génica , Kisspeptinas/agonistas , Hormona Luteinizante/metabolismo , Neuronas/metabolismo , Norepinefrina/metabolismo , Área Preóptica/metabolismo , Regulación hacia Arriba , Factores de Transcripción ARNTL/agonistas , Factores de Transcripción ARNTL/antagonistas & inhibidores , Factores de Transcripción ARNTL/genética , Factores de Transcripción ARNTL/metabolismo , Antagonistas de Receptores Adrenérgicos alfa 1/farmacología , Animales , Estradiol/farmacología , Terapia de Reemplazo de Estrógeno , Femenino , Fase Folicular/efectos de los fármacos , Regulación de la Expresión Génica/efectos de los fármacos , Kisspeptinas/antagonistas & inhibidores , Kisspeptinas/genética , Kisspeptinas/metabolismo , Proteínas del Tejido Nervioso/agonistas , Proteínas del Tejido Nervioso/genética , Proteínas del Tejido Nervioso/metabolismo , Neuronas/efectos de los fármacos , Ovariectomía/efectos adversos , Proteínas Circadianas Period/agonistas , Proteínas Circadianas Period/antagonistas & inhibidores , Proteínas Circadianas Period/genética , Proteínas Circadianas Period/metabolismo , Prazosina/farmacología , Área Preóptica/efectos de los fármacos , Ratas Wistar , Receptores Adrenérgicos alfa 1/química , Receptores Adrenérgicos alfa 1/metabolismo , Transducción de Señal/efectos de los fármacos , Transmisión Sináptica/efectos de los fármacos , Regulación hacia Arriba/efectos de los fármacos
16.
Obes Rev ; 16(10): 871-82, 2015 Oct.
Artículo en Inglés | MEDLINE | ID: mdl-26214605

RESUMEN

The circadian system, headed by the suprachiasmatic nucleus, synchronizes behaviour and metabolism according to the external light-dark cycle through neuroendocrine and autonomic signals. Metabolic diseases, such as steatosis, obesity and glucose intolerance, have been associated with conditions of circadian misalignment wherein the feeding schedule has been moved to the resting phase. Here we describe the physiological processes involved in liver lipid accumulation and show how they follow a circadian pattern importantly regulated by both the autonomic nervous system and the feeding-fasting cycle. We propose that an unbalanced activity of the sympathetic-parasympathetic branches between organs induced by circadian misalignment provides the conditions for the development and progression of non-alcoholic fatty liver disease.


Asunto(s)
Sistema Nervioso Autónomo/fisiopatología , Ácidos Grasos no Esterificados/metabolismo , Hígado/metabolismo , Enfermedad del Hígado Graso no Alcohólico/metabolismo , Obesidad/metabolismo , Proteínas Circadianas Period/metabolismo , Tejido Adiposo , Ritmo Circadiano , Regulación de la Expresión Génica , Humanos , Lipólisis , Datos de Secuencia Molecular , Enfermedad del Hígado Graso no Alcohólico/etiología , Enfermedad del Hígado Graso no Alcohólico/fisiopatología , Obesidad/complicaciones , Obesidad/fisiopatología
17.
J Pineal Res ; 58(4): 490-9, 2015 May.
Artículo en Inglés | MEDLINE | ID: mdl-25807895

RESUMEN

This study investigated the maturational stage (immature and mature ovaries) differences of mRNA expression of melatonin-forming enzymes (Aanat and Asmt), melatonin membrane receptors (Mt1 and Mt2) and putative nuclear (Rorα) receptors, and clock genes (Clock, Bmal1, Per1, Per2, Cry1, Cry2) in cumulus-oocyte complexes (COC) from weaning Wistar rats. We also examined the effects of pinealectomy and of melatonin pharmacological replacement on the daily expression of these genes in COC. qRT-PCR analysis revealed that in oocytes, the mRNA expression of Asmt, Mt2, Clock, Bmal1, Per2, and Cry1 were higher (P < 0.05) in immature ovaries than in the mature ones. In cumulus cells, the same pattern of mRNA expression for Asmt, Aanat, Rorα, Clock, Per1, Cry1, and Cry2 genes was observed. In oocytes, pinealectomy altered the daily mRNA expression profiles of Asmt, Mt1, Mt2, Clock, Per1, Cry1, and Cry2 genes. In cumulus cells, removal of the pineal altered the mRNA expression profiles of Mt1, Mt2, Rorα, Aanat, Asmt, Clock, Bmal1, Per2, Cry1, and Cry2 genes. Melatonin treatment partially or completely re-established the daily mRNA expression profiles of most genes studied. The mRNA expression of melatonin-related genes and clock genes in rat COC varies with the maturational stage of the meiotic cellular cycle in addition to the hour of the day. This suggests that melatonin might act differentially in accordance with the maturational stage of cumulus/oocyte complex. In addition, it seems that circulating pineal melatonin is very important in the design of the daily profile of mRNA expression of COC clock genes and genes related to melatonin synthesis and action.


Asunto(s)
Células del Cúmulo/metabolismo , Melatonina/metabolismo , Oocitos/metabolismo , Glándula Pineal/cirugía , Animales , N-Acetiltransferasa de Arilalquilamina/metabolismo , Ritmo Circadiano/fisiología , Femenino , Proteínas Circadianas Period/metabolismo , Ratas , Ratas Wistar , Receptores de Melatonina/metabolismo
18.
J Pineal Res ; 58(3): 251-61, 2015 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-25626464

RESUMEN

Melatonin, the main hormone produced by the pineal gland, is secreted in a circadian manner (24-hr period), and its oscillation influences several circadian biological rhythms, such as the regulation of clock genes expression (chronobiotic effect) and the modulation of several endocrine functions in peripheral tissues. Assuming that the circadian synchronization of clock genes can play a role in the regulation of energy metabolism and it is influenced by melatonin, our study was designed to assess possible alterations as a consequence of melatonin absence on the circadian expression of clock genes in the epididymal adipose tissue of male Wistar rats and the possible metabolic repercussions to this tissue. Our data show that pinealectomy indeed has impacts on molecular events: it abolishes the daily pattern of the expression of Clock, Per2, and Cry1 clock genes and Pparγ expression, significantly increases the amplitude of daily expression of Rev-erbα, and affects the pattern of and impairs adipokine production, leading to a decrease in leptin levels. However, regarding some metabolic aspects of adipocyte functions, such as its ability to synthesize triacylglycerols from glucose along 24 hr, was not compromised by pinealectomy, although the daily profile of the lipogenic enzymes expression (ATP-citrate lyase, malic enzyme, fatty acid synthase, and glucose-6-phosphate dehydrogenase) was abolished in pinealectomized animals.


Asunto(s)
Tejido Adiposo Blanco/metabolismo , Ritmo Circadiano/genética , Expresión Génica/genética , Proteínas Circadianas Period/metabolismo , Glándula Pineal , Animales , Ritmo Circadiano/fisiología , Expresión Génica/fisiología , Masculino , Proteínas Circadianas Period/genética , Glándula Pineal/enzimología , Glándula Pineal/fisiología , Glándula Pineal/cirugía , Ratas , Ratas Wistar
19.
J Comp Neurol ; 523(6): 982-96, 2015 Apr 15.
Artículo en Inglés | MEDLINE | ID: mdl-25504089

RESUMEN

Circadian rhythms are conserved across kingdoms and coordinate physiology and behavior for appropriate time-keeping. The neuronal populations that govern circadian rhythms are described in many animal models, and the current challenge is to understand how they interact to control overt rhythms, remaining plastic enough to respond and adapt to a changing environment. In Drosophila melanogaster, the circadian network comprises about 150 neurons, and the main synchronizer is the neuropeptide pigment-dispersing factor (PDF), released by the well-characterized central pacemaker neurons, the small ventral lateral neurons (sLNvs). However, the rules and properties governing the communication and coupling between this central pacemaker and downstream clusters are not fully elucidated. Here we genetically manipulate the speed of the molecular clock specifically in the central pacemaker neurons of Drosophila and provide experimental evidence of their restricted ability to synchronize downstream clusters. We also demonstrate that the sLNv-controlled clusters have an asymmetric entrainment range and were able to experimentally assess it. Our data imply that different clusters are subjected to different coupling strengths, and display independent endogenous periods. Finally, the manipulation employed here establishes a suitable paradigm to test other network properties as well as the cell-autonomous mechanisms running in different circadian-relevant clusters.


Asunto(s)
Encéfalo/anatomía & histología , Relojes Circadianos/fisiología , Drosophila melanogaster/anatomía & histología , Drosophila melanogaster/fisiología , Red Nerviosa/fisiología , Análisis de Varianza , Animales , Animales Modificados Genéticamente , Encéfalo/metabolismo , Proteínas de Drosophila/genética , Proteínas de Drosophila/metabolismo , Actividad Motora , Red Nerviosa/metabolismo , Neuronas/metabolismo , Proteínas Circadianas Period/genética , Proteínas Circadianas Period/metabolismo
20.
PLoS One ; 9(9): e106252, 2014.
Artículo en Inglés | MEDLINE | ID: mdl-25184495

RESUMEN

Melanopsin has been implicated in the mammalian photoentrainment by blue light. This photopigment, which maximally absorbs light at wavelengths between 470 and 480 nm depending on the species, is found in the retina of all classes of vertebrates so far studied. In mammals, melanopsin activation triggers a signaling pathway which resets the circadian clock in the suprachiasmatic nucleus (SCN). Unlike mammals, Drosophila melanogaster and Danio rerio do not rely only on their eyes to perceive light, in fact their whole body may be capable of detecting light and entraining their circadian clock. Melanopsin, teleost multiple tissue (tmt) opsin and others such as neuropsin and va-opsin, are found in the peripheral tissues of Danio rerio, however, there are limited data concerning the photopigment/s or the signaling pathway/s directly involved in light detection. Here, we demonstrate that melanopsin is a strong candidate to mediate synchronization of zebrafish cells. The deduced amino acid sequence of melanopsin, although being a vertebrate opsin, is more similar to invertebrate than vertebrate photopigments, and melanopsin photostimulation triggers the phosphoinositide pathway through activation of a G(q/11)-type G protein. We stimulated cultured ZEM-2S cells with blue light at wavelengths consistent with melanopsin maximal absorption, and evaluated the time course expression of per1b, cry1b, per2 and cry1a. Using quantitative PCR, we showed that blue light is capable of slightly modulating per1b and cry1b genes, and drastically increasing per2 and cry1a expression. Pharmacological assays indicated that per2 and cry1a responses to blue light are evoked through the activation of the phosphoinositide pathway, which crosstalks with nitric oxide (NO) and mitogen activated protein MAP kinase (MAPK) to activate the clock genes. Our results suggest that melanopsin may be important in mediating the photoresponse in Danio rerio ZEM-2S cells, and provide new insights about the modulation of clock genes in peripheral clocks.


Asunto(s)
Relojes Circadianos/genética , Fibroblastos/efectos de la radiación , Subunidades alfa de la Proteína de Unión al GTP Gq-G11/genética , Retina/efectos de la radiación , Opsinas de Bastones/genética , Proteínas de Pez Cebra/genética , Secuencia de Aminoácidos , Animales , Línea Celular , Criptocromos/genética , Criptocromos/metabolismo , Embrión no Mamífero , Proteínas del Ojo/genética , Proteínas del Ojo/metabolismo , Fibroblastos/citología , Fibroblastos/metabolismo , Subunidades alfa de la Proteína de Unión al GTP Gq-G11/metabolismo , Regulación de la Expresión Génica , Luz , Proteínas Quinasas Activadas por Mitógenos/genética , Proteínas Quinasas Activadas por Mitógenos/metabolismo , Datos de Secuencia Molecular , Óxido Nítrico/metabolismo , Proteínas Circadianas Period/genética , Proteínas Circadianas Period/metabolismo , Fosfatidilinositoles/metabolismo , Estimulación Luminosa , Retina/citología , Retina/metabolismo , Opsinas de Bastones/metabolismo , Transducción de Señal , Núcleo Supraquiasmático/citología , Núcleo Supraquiasmático/metabolismo , Pez Cebra , Proteínas de Pez Cebra/metabolismo
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