Calcium and SOL Protease Mediate Temperature Resetting of Circadian Clocks.

Circadian clocks integrate light and temperature input to remain synchronized with the day/night cycle. Although light input to the clock is well studied, the molecular mechanisms by which circadian clocks respond to temperature remain poorly understood. We found that temperature phase shifts Drosophila circadian clocks through degradation of the pacemaker protein TIM. This degradation is mechanistically distinct from photic CRY-dependent TIM degradation. Thermal TIM degradation is triggered by cytosolic calcium increase and CALMODULIN binding to TIM and is mediated by the atypical calpain protease SOL. This thermal input pathway and CRY-dependent light input thus converge on TIM, providing a molecular mechanism for the integration of circadian light and temperature inputs. Mammals use body temperature cycles to keep peripheral clocks synchronized with their brain pacemaker. Interestingly, downregulating the mammalian SOL homolog SOLH blocks thermal mPER2 degradation and phase shifts. Thus, we propose that circadian thermosensation in insects and mammals share common principles.

A New Nephrometry Score for Predicting Positive Surgical Margin After Laparoscopic Partial Nephrectomy.

PURPOSE: To introduce the KESKIN ratio as a novel predictor of positive surgical margin (PSM) after laparoscopic partial nephrectomy (PN) and to evaluate other clinical characteristics and nephrometry scores (including RENAL, PADUA, and C-index) for predicting PSM. METHODS: We retrospectively analyzed 95 patients who underwent laparoscopic PN between June 2020 and April 2023. The KESKIN ratio was defined for all patients. The KESKIN ratio, tumor and patient-related paramaters, and nephrometry scores were analyzed to predict PSM. RESULTS: Positive surgical margin was found in 12 of 95 patients (12.6%). There was no statistical difference between the PSM and negative surgical margin (NSM) groups in RENAL, PADUA, and C-index scores. Only the KESKIN ratio was found to be a statistically significant predictor of PSM in both univariate and multivariate regression analysis (p = 0.007 and p = 0.043, respectively). Mean endophytic diameter and endophytic percentage were found to be statistically significant predictors of PSM in only univariate analysis (p = 0.005 and p = 0.01, respectively). The value of 0.5 was determined as the cut-off value for the KESKIN ratio. Values higher than 0.5 indicate an increase in PSM. CONCLUSIONS: The KESKIN ratio is a novel, easily measurable, and calculable image-based parameter that can be used to predict PSM after laparascopic PN. If externally validated in a larger patient population, the KESKIN ratio may be used in future versions of the current nephrometry scoring systems for predicting the PSM.

Risk Factors of Open Surgery Conversion in Laparoscopic Partial Nephrectomy to Achieve Nephron Sparing.

OBJECTIVES: We aimed to evaluate the risk factors for the conversion from laparoscopic partial nephrectomy (LPN) to open surgery to achieve partial nephrectomy (PN). METHODS: Data from patients who underwent LPN between June 2020 and September 2023 were analyzed retrospectively. Patients in whom the PN procedure could be completed laparoscopically were recorded as the 'Fully Laparoscopic' (FL) group (n = 97), and those converted to open surgery from laparoscopy were recorded as the 'Conversion to Open' (CTO) group (n = 10). The demographic and pathologic variables were compared between groups. Regression analyses were used to define predictor factors, and receiver operating characteristic analysis was used to define the cut-off value of the surgical bleeding volume. RESULTS: Conversion to open surgery was found in 10/107 patients (9.3%). There was no statistical difference between groups in demographic and pathologic variables. Intraoperative blood loss volume, upper pole localized tumor, and posterior localized tumor were found to be statistically higher in the CTO group (p = 0.001, p = 0.001, and p = 0.043, respectively). Furthermore, these factors were only found to be statistically significant predictors of conversion to open surgery in both univariate and multivariate regression analyses. 235 cc was found to be the cut-off value of intraoperative blood loss volume for predicting conversion to open surgery (p = 0.001). CONCLUSION: Using these predictive factors in clinical practice, treatment planning will lead to the possibility of starting the treatment directly with open surgery instead of minimally invasive options, and it may also provide a chance of being prepared for the possibility of conversion to open surgery peroperatively.

Studying circadian rhythms in Drosophila melanogaster.

Circadian rhythms have a profound influence on most bodily functions: from metabolism to complex behaviors. They ensure that all these biological processes are optimized with the time-of-day. They are generated by endogenous molecular oscillators that have a period that closely, but not exactly, matches day length. These molecular clocks are synchronized by environmental cycles such as light intensity and temperature. Drosophila melanogaster has been a model organism of choice to understand genetically, molecularly and at the level of neural circuits how circadian rhythms are generated, how they are synchronized by environmental cues, and how they drive behavioral cycles such as locomotor rhythms. This review will cover a wide range of techniques that have been instrumental to our understanding of Drosophila circadian rhythms, and that are essential for current and future research.

Glycogen synthase kinase is a regulator of the circadian clock of Neurospora crassa.

Timekeeping by circadian clocks relies upon precise adjustment of expression levels of clock proteins. Here we identify glycogen synthase kinase (GSK) as a novel and critical component of the circadian clock of Neurospora crassa that regulates the abundance of its core transcription factor white collar complex (WCC) on a post-transcriptional level. We show that GSK specifically binds and phosphorylates both subunits of the WCC. Reduced expression of GSK promotes an increased accumulation of WC-1, the limiting factor of the WCC, causing an acceleration of the circadian clock and a shorter free-running period.

Of switches and hourglasses: regulation of subcellular traffic in circadian clocks by phosphorylation.

Investigation of the phosphorylation of circadian clock proteins has shown that this modification contributes to circadian timing in all model organisms. Phosphorylation alters the stability, transcriptional activity and subcellular localization of clock proteins during the course of a day, such that time-of-day-specific phosphorylation encodes information for measuring time and is crucial for the establishment of an approximately 24-h period. One main feature of molecular timekeeping is the daytime-specific nuclear accumulation of clock proteins, which can be regulated by phosphorylation. Here, we discuss increasing knowledge of how subcellular shuttling is regulated in circadian clocks, on the basis of recent observations in Neurospora crassa showing that clock proteins undergo maturation through sequential phosphorylation. In this model organism, clock proteins are regulated by the phosphorylation-dependent modulation of rapid shuttling cycles that alter their subcellular localization in a time-of-day-specific manner.

Lithium and genetic inhibition of GSK3beta enhance the effect of methamphetamine on circadian rhythms in the mouse.

Lithium, a drug commonly used to treat mood disorders, and the psychostimulant methamphetamine are both capable of altering circadian rhythmicity. Although the actions of lithium on the circadian system are thought to occur through inhibition of glycogen synthase kinase-3beta (GSK3beta), the mechanism by which methamphetamine alters circadian rhythms is unknown. We tested the effects of concurrent methamphetamine and lithium treatment on the circadian wheel-running behavior of mice. Methamphetamine alone lengthened both the active duration and the free-running period of locomotor activity in animals housed in constant conditions. Administering lithium enhanced the period-lengthening effects of methamphetamine in animals housed in constant darkness. This effect was even more pronounced when animals were housed in constant light. Lithium increased both methamphetamine intake and serum levels of methamphetamine, possibly contributing to the effects on circadian behavior. We also tested the effect of methamphetamine in mutant mice possessing only one allele for Gsk3beta. These animals, when treated with methamphetamine, responded like wild-type mice treated with a combination of methamphetamine and lithium, displaying long, free-running rhythms. These data, together with many others in the literature, point to a complicated interaction between the circadian system and the development and possible treatment of psychopathologies such as bipolar disorder and drug addiction.

The methamphetamine-sensitive circadian oscillator (MASCO) in mice.

The suprachiasmatic nucleus (SCN) orchestrates synchrony among many peripheral oscillators and is required for circadian rhythms of locomotor activity and many physiological processes. However, the unique effects of methamphetamine (MAP) on circadian behavior suggest the presence of an SCN-independent, methamphetamine-sensitive circadian oscillator (MASCO). Substantial data collected using rat models show that chronic methamphetamine dramatically lengthens circadian period of locomotor activity rhythms and induces rhythms in animals lacking an SCN. However, the anatomical substrate and the molecular components of the MASCO are unknown. The response to MAP is less well studied in mice, a model that would provide the genetic tools to probe the molecular components of this extra-SCN oscillator. The authors tested the effects of chronic MAP on 2 strains of intact and SCN-lesioned mice in constant dark and constant light. Furthermore, they applied various MAP availability schedules to SCN-lesioned mice to confirm the circadian nature of the underlying oscillator. The results indicate that this oscillator has circadian properties. In intact mice, the MASCO interacts with the SCN in a manner that is strain, sex, and dose dependent. In SCN-lesioned mice, it induces robust free-running locomotor rhythmicity, which persists for up to 14 cycles after methamphetamine is withdrawn. In the future, localization of the MASCO and characterization of its underlying molecular mechanism, as well as its interactions with other oscillators in the body, will be essential to a complete understanding of the organization of the mammalian circadian system.

The molecular ticks of the Drosophila circadian clock.

Drosophila is a powerful model to understand the mechanisms underlying circadian rhythms. The Drosophila molecular clock is comprised of transcriptional feedback loops. The expressions of the critical transcriptional activator CLK and its repressors PER and TIM are under tight transcriptional control. However, posttranslational modification of these proteins and regulation of their stability are critical to their function and to the generation of 24-hr period rhythms. We review here recent progress made in our understanding of PER, TIM and CLK posttranslational control. We also review recent studies that are uncovering the importance of novel regulatory mechanisms that affect mRNA stability and translation of circadian pacemaker proteins and their output.

Effect of lesioning the suprachiasmatic nuclei on behavioral despair in rats.

The suprachiasmatic nucleus (SCN) is involved in regulating many biological rhythms. Several lines of research implicate the SCN in affective behavior. The SCN is directly involved in regulating the daily rhythms of the hypothalamo-pituitary-adrenal (HPA) axis hormones involved in stress. Bilateral lesions of the SCN disrupt both the rhythms and the basal levels of the HPA axis hormones involved in coping with stress. Moreover, stress can affect the biological rhythms regulated by the SCN, and disruption of biological rhythms in turn can cause stress. The present study assessed the effect of bilateral destruction of the SCN on behavioral despair, an animal model of depression sensitive to antidepressant treatment. The results indicate that bilateral destruction of the SCN results in reduced immobility in the second forced swimming test (FST) compared to sham controls and animals with incomplete lesions. These results indicate that bilateral destruction of the SCN has a protective effect in the induction of behavioral despair which may arise out of disruption of the secretion of the HPA axis hormones and/or of the neural connections between the SCN and the limbic structures that modulate the response to swim stress.

Circadian effects of timed meals (and other rewards).

Mammals organize many of their activities around rhythmic events in their environments. Primary among these events is the daily light-dark cycle. However, for many animals, food availability is rhythmic or quasi-rhythmic and is therefore a potential synchronizing cue. While circadian rhythms in both behavior and physiological activity can be entrained in animals via meal-feeding schedules, the mechanism by which this occurs remains poorly understood. Similarities between the circadian effects of restricted feeding and the effects of chronic methamphetamine treatment may be indicative of a common mechanism. This article argues that reward (or the arousal that accompanies it) may be the final common pathway for such nonphotic circadian inputs.