Metabolic Oscillations Research Articles

Period-1 encodes an ATP-dependent RNA helicase that influences nutritional compensation of the Neurospora circadian clock

Mutants in the period-1 (prd-1) gene, characterized by a recessive allele, display a reduced growth rate and period lengthening of the developmental cycle controlled by the circadian clock. We refined the genetic location of prd-1 and used whole genome sequencing to find the mutation defining it, confirming the identity of prd-1 by rescuing the mutant circadian phenotype via transformation. PRD-1 is an RNA helicase whose orthologs, DDX5 [DEAD (Asp-Glu-Ala-Asp) Box Helicase 5] and DDX17 in humans and DBP2 (Dead Box Protein 2) in yeast, are implicated in various processes, including transcriptional regulation, elongation, and termination, ribosome biogenesis, and mRNA decay. Although prd-1 mutants display a long period (∼25 h) circadian developmental cycle, they interestingly display a WT period when the core circadian oscillator is tracked using a frq-luciferase transcriptional fusion under conditions of limiting nutritional carbon; the core oscillator in the prd-1 mutant strain runs with a long period under glucose-sufficient conditions. Thus, PRD-1 clearly impacts the circadian oscillator and is not only part of a metabolic oscillator ancillary to the core clock. PRD-1 is an essential protein, and its expression is neither light-regulated nor clock-regulated. However, it is transiently induced by glucose; in the presence of sufficient glucose, PRD-1 is in the nucleus until glucose runs out, which elicits its disappearance from the nucleus. Because circadian period length is carbon concentration-dependent, prd-1 may be formally viewed as a clock mutant with defective nutritional compensation of circadian period length.

Mathematical modeling of gap junction coupling and electrical activity in human β-cells

Coordinated insulin secretion is controlled by electrical coupling of pancreatic β-cells due to connexin-36 gap junctions. Gap junction coupling not only synchronizes the heterogeneous β-cell population, but can also modify the electrical behavior of the cells. These phenomena have been widely studied with mathematical models based on data from mouse β-cells. However, it is now known that human β-cell electrophysiology shows important differences to its rodent counterpart, and although human pancreatic islets express connexin-36 and show evidence of β-cell coupling, these aspects have been little investigated in human β-cells. Here we investigate theoretically, the gap junction coupling strength required for synchronizing electrical activity in a small cluster of cells simulated with a recent mathematical model of human β-cell electrophysiology. We find a lower limit for the coupling strength of approximately 20 pS (i.e., normalized to cell size, ∼2 pS pF−1) below which spiking electrical activity is asynchronous. To confront this theoretical lower bound with data, we use our model to estimate from an experimental patch clamp recording that the coupling strength is approximately 100–200 pS (10–20 pS pF−1), similar to previous estimates in mouse β-cells. We then investigate the role of gap junction coupling in synchronizing and modifying other forms of electrical activity in human β-cell clusters. We find that electrical coupling can prolong the period of rapid bursting electrical activity, and synchronize metabolically driven slow bursting, in particular when the metabolic oscillators are in phase. Our results show that realistic coupling conductances are sufficient to promote synchrony in small clusters of human β-cells as observed experimentally, and provide motivation for further detailed studies of electrical coupling in human pancreatic islets.

MYC Disrupts the Circadian Clock and Metabolism in Cancer Cells

The MYC oncogene encodes MYC, a transcription factor that binds the genome through sites termed E-boxes (5'-CACGTG-3'), which are identical to the binding sites of the heterodimeric CLOCK-BMAL1 master circadian transcription factor. Hence, we hypothesized that ectopic MYC expression perturbs the clock by deregulating E-box-driven components of the circadian network in cancer cells. Wereport here that deregulated expression of MYC or N-MYC disrupts the molecular clock invitro by directly inducing REV-ERBα to dampen expression and oscillation of BMAL1, and this could be rescued by knockdown of REV-ERB. REV-ERBα expression predicts poor clinical outcome for N-MYC-driven human neuroblastomas that have diminished BMAL1 expression, and re-expression of ectopic BMAL1 in neuroblastoma cell lines suppresses their clonogenicity. Further, ectopic MYC profoundly alters oscillation of glucose metabolism and perturbs glutaminolysis. Our results demonstrate an unsuspected link between oncogenic transformation and circadian and metabolic dysrhythmia, which we surmise to be advantageous for cancer.

Endogenous rhythmic growth in oak trees is regulated by internal clocks rather than resource availability.

Common oak trees display endogenous rhythmic growth with alternating shoot and root flushes. To explore the mechanisms involved, microcuttings of the Quercus robur L. clone DF159 were used for (13)C/(15)N labelling in combination with RNA sequencing (RNASeq) transcript profiling of shoots and roots. The effect of plant internal resource availability on the rhythmic growth of the cuttings was tested through inoculation with the ectomycorrhizal fungus Piloderma croceum. Shoot and root flushes were related to parallel shifts in above- and below-ground C and, to a lesser extent, N allocation. Increased plant internal resource availability by P. croceum inoculation with enhanced plant growth affected neither the rhythmic growth nor the associated resource allocation patterns. Two shifts in transcript abundance were identified during root and shoot growth cessation, and most concerned genes were down-regulated. Inoculation with P. croceum suppressed these transcript shifts in roots, but not in shoots. To identify core processes governing the rhythmic growth, functions [Gene Ontology (GO) terms] of the genes differentially expressed during the growth cessation in both leaves and roots of non-inoculated plants and leaves of P. croceum-inoculated plants were examined. Besides genes related to resource acquisition and cell development, which might reflect rather than trigger rhythmic growth, genes involved in signalling and/or regulated by the circadian clock were identified. The results indicate that rhythmic growth involves dramatic oscillations in plant metabolism and gene regulation between below- and above-ground parts. Ectomycorrhizal symbiosis may play a previously unsuspected role in smoothing these oscillations without modifying the rhythmic growth pattern.

Modeling of Glucose-Induced cAMP Oscillations in Pancreatic β Cells: cAMP Rocks when Metabolism Rolls

Recent advances in imaging technology have revealed oscillations of cyclic adenosine monophosphate (cAMP) in insulin-secreting cells. These oscillations may be in phase with cytosolic calcium oscillations or out of phase. cAMP oscillations have previously been modeled as driven by oscillations in calcium, based on the known dependence of the enzymes that generate cAMP (adenylyl cyclase) and degrade it (phosphodiesterase). However, cAMP oscillations have also been reported to occur in the absence of calcium oscillations. Motivated by similarities between the properties of cAMP and metabolic oscillations in pancreatic β cells, we propose here that in addition to direct control by calcium, cAMP is controlled by metabolism. Specifically, we hypothesize that AMP inhibits adenylyl cyclase. We incorporate this hypothesis into the dual oscillator model for β cells, in which metabolic (glycolytic) oscillations cooperate with modulation of ion channels and metabolism by calcium. We show that the combination of oscillations in AMP and calcium in the dual oscillator model can account for the diverse oscillatory patterns that have been observed, as well as for experimental perturbations of those patterns. Predictions to further test the model are provided.

Cyanohydrin reactions enhance glycolytic oscillations in yeast

Synchronous metabolic oscillations can be induced in yeast by addition of glucose and removal of extracellular acetaldehyde (ACAx). Compared to other means of ACAx removal, cyanide robustly induces oscillations, indicating additional cyanide reactions besides ACA to lactonitrile conversion. Here, 13C NMR is used to confirm our previous hypothesis, that cyanide directly affects glycolytic fluxes through reaction with carbonyl-containing compounds. Intracellularly, at least 3 cyanohydrins were identified. Extracellularly, all signals could be identified and lactonitrile was found to account for ~66% of total cyanide removal. Simulations of our updated computational model show that intracellular cyanide reactions increase the amplitude of oscillations and that cyanide addition lowers [ACA] instantaneously. We conclude that cyanide provides the following means of inducing global oscillations: a) by reducing [ACAx] relative to oscillation amplitude, b) by targeting multiple intracellular carbonyl compounds during fermentation, and c) by acting as a phase resetting stimulus.

Entrainment of heterogeneous glycolytic oscillations in single cells.

Cell signaling, gene expression, and metabolism are affected by cell-cell heterogeneity and random changes in the environment. The effects of such fluctuations on cell signaling and gene expression have recently been studied intensively using single-cell experiments. In metabolism heterogeneity may be particularly important because it may affect synchronisation of metabolic oscillations, an important example of cell-cell communication. This synchronisation is notoriously difficult to describe theoretically as the example of glycolytic oscillations shows: neither is the mechanism of glycolytic synchronisation understood nor the role of cell-cell heterogeneity. To pin down the mechanism and to assess its robustness and universality we have experimentally investigated the entrainment of glycolytic oscillations in individual yeast cells by periodic external perturbations. We find that oscillatory cells synchronise through phase shifts and that the mechanism is insensitive to cell heterogeneity (robustness) and similar for different types of external perturbations (universality).

Non-transcriptional/translational regulations of the circadian system

Circadian clocks in organisms drive physiology and behavior, helping with adjustment to earth’s environment and human society. The generation and regulation mechanisms of circadian rhythms are starting to be understood. The transcriptional/post-transcriptional delayed feedback loop has been widely studied as the main regulation mechanism. But it cannot fully account for all circadian rhythms in cells. It has been reported that the “proto-clock” may have been a cytosolic metabolic oscillation and that the transcriptional translational regulation system, which provides robustness and amplifies circadian outputs, was developed during evolution. Research on circadian rhythms in cyanobacteria Synechococcus elongatus, algae Ostreococcus tauri, and human red blood cells in the absence of transcription and translation indicates the existence of a non-transcriptional/translational regulation mechanism, in addition to the classical transcriptional/translational regulation mechanism. It revealed a close link between the two regulation systems of circadian rhythms, in which they might complement each other. And NAD, involved in energy metabolism, may be involved in their interaction.

Tight coupling of metabolic oscillations and intracellular water dynamics in Saccharomyces cerevisiae.

We detected very strong coupling between the oscillating concentration of ATP and the dynamics of intracellular water during glycolysis in Saccharomyces cerevisiae. Our results indicate that: i) dipolar relaxation of intracellular water is heterogeneous within the cell and different from dilute conditions, ii) water dipolar relaxation oscillates with glycolysis and in phase with ATP concentration, iii) this phenomenon is scale-invariant from the subcellular to the ensemble of synchronized cells and, iv) the periodicity of both glycolytic oscillations and dipolar relaxation are equally affected by D2O in a dose-dependent manner. These results offer a new insight into the coupling of an emergent intensive physicochemical property of the cell, i.e. cell-wide water dipolar relaxation, and a central metabolite (ATP) produced by a robustly oscillating metabolic process.

Small heterodimer partner/neuronal PAS domain protein 2 axis regulates the oscillation of liver lipid metabolism.

In mammals, circadian rhythms are essential for coordinating the timing of various metabolic processes. The Clock gene regulates diurnal plasma triglyceride fluctuation through nuclear receptor small heterodimer partner (Shp; Nr0b2). Given that SHP is a critical regulator of metabolism in the liver, it is unknown whether SHP is necessary to coordinate metabolism and circadian rhythms. Shp(+/+) and Shp(-/-) mice on a C57BL/6 background (n = 3-5/group) were fed a standard chow diet and water ad libitum. Serum and livers were collected at zeitgeber time 2, 6, 10, 14, 18, and 22. In vivo and in vitro assays included RNA sequencing, quantitative polymerase chain reaction, very-low-density lipoprotein production, adenovirus overexpression and small interfering RNA knockdown, serum parameters, circadian locomotor activity, Oil Red O staining, transient transfection, luciferase reporter assay, chromatin immunoprecipitation assay, gel-shift assay, coimmunoprecipitation, and western blottings. Shp deficiency had a robust global impact on major liver metabolic genes. Several components of the liver clock, including peroxisome proliferator-activated receptor-γ, coactivator 1 (Pgc-1α), neuronal PAS domain-containing protein 2 (Npas2), and retinoic acid-related orphan receptor (Ror)α/γ were sharply induced in Shp(-/-) liver. At the molecular level, SHP inhibited Npas2 gene transcription and promoter activity through interaction with Rorγ to repress Rorγ transactivation and by interacting with Rev-erbα to enhance its inhibition of Rorα activity. Conversely, Npas2 controlled the circadian rhythm of Shp expression by binding rhythmically to the Shp promoter, which was enhanced by nicotinamide adenine dinucleotide, but not nicotinamide adenine dinucleotide phosphate. Phenotypically, Npas2 deficiency induced severe steatosis in Shp(-/-) mice, which was attributed to the dysregulation of lipoprotein metabolism. Shp and Npas2 crosstalk is essential to maintain hepatic lipid homeostasis.

Letter to the Editor Pyoderma gangrenosum with monoclonal IgA gammopathy and pulmonary tuberculosis. Illustrative case and review

ENWEndNote BIBJabRef, Mendeley RISPapers, Reference Manager, RefWorks, Zotero AMA Romańska-Gocka K, Cieścińska C, Zegarska B, et al. Letter to the EditorPyoderma gangrenosum with monoclonal IgA gammopathy and pulmonary tuberculosis. Illustrative case and review. Advances in Dermatology and Allergology/Postępy Dermatologii i Alergologii. 2015;32(2):137-141. doi:10.5114/pdia.2014.40974. APA Romańska-Gocka, K., Cieścińska, C., Zegarska, B., Schwartz, R., Cieściński, J., & Olszewska-Słonina, D. et al. (2015). Letter to the EditorPyoderma gangrenosum with monoclonal IgA gammopathy and pulmonary tuberculosis. Illustrative case and review. Advances in Dermatology and Allergology/Postępy Dermatologii i Alergologii, 32(2), 137-141. https://doi.org/10.5114/pdia.2014.40974 Chicago Romańska-Gocka, Krystyna, Czanita Cieścińska, Barbara Zegarska, Robert A. Schwartz, Jakub Cieściński, Dorota Olszewska-Słonina, and Rafał Czajkowski. 2015. "Letter to the EditorPyoderma gangrenosum with monoclonal IgA gammopathy and pulmonary tuberculosis. Illustrative case and review". Advances in Dermatology and Allergology/Postępy Dermatologii i Alergologii 32 (2): 137-141. doi:10.5114/pdia.2014.40974. Harvard Romańska-Gocka, K., Cieścińska, C., Zegarska, B., Schwartz, R., Cieściński, J., Olszewska-Słonina, D., and Czajkowski, R. (2015). Letter to the EditorPyoderma gangrenosum with monoclonal IgA gammopathy and pulmonary tuberculosis. Illustrative case and review. Advances in Dermatology and Allergology/Postępy Dermatologii i Alergologii, 32(2), pp.137-141. https://doi.org/10.5114/pdia.2014.40974 MLA Romańska-Gocka, Krystyna et al. "Letter to the EditorPyoderma gangrenosum with monoclonal IgA gammopathy and pulmonary tuberculosis. Illustrative case and review." Advances in Dermatology and Allergology/Postępy Dermatologii i Alergologii, vol. 32, no. 2, 2015, pp. 137-141. doi:10.5114/pdia.2014.40974. Vancouver Romańska-Gocka K, Cieścińska C, Zegarska B, Schwartz R, Cieściński J, Olszewska-Słonina D et al. Letter to the EditorPyoderma gangrenosum with monoclonal IgA gammopathy and pulmonary tuberculosis. Illustrative case and review. Advances in Dermatology and Allergology/Postępy Dermatologii i Alergologii. 2015;32(2):137-141. doi:10.5114/pdia.2014.40974.

Glucose-Induced Cyclic-AMP Oscillations: Modeling Incretin Impact on Pancreatic Beta Cell Secretion

Pancreatic beta cells modulate insulin secretion via incretin-induced amplification pathways of cytosolic calcium (Ca2+)-induced exocytosis, which may be help compensate for reduced insulin sensitivity in Type II Diabetes Mellitus. One modulator is cAMP, which is activated by an incretin-triggered Gs protein cascade and sensitive to glucose levels. Interestingly, cAMP is found to be oscillatory, even in cases where Ca2+ is not, providing a possible mechanism for pulstile insulin secretion in the absence of Ca2+ oscillations. When Ca2+ and cAMP both oscillate, the two may be either in-phase or out-of-phase with one another. To understand the cAMP oscillations and their relationship to Ca2+ oscillations we use mathematical modeling. We model the cAMP production sensitivity to glucose metabolism with AMP sensitive adenylyl cyclases and degradation with Ca2+ sensitive phosphodiesterases. These modest additions to the Dual Oscillator Model for beta cells allow us to capture observed relationships between cAMP and Ca2+. This provides further evidence for metabolic oscillations in beta-cells that are affected by, but that do not require, oscillations in Ca2+.

Considerations for RNA-seq analysis of circadian rhythms.

Circadian rhythms are daily endogenous oscillations of behavior, metabolism, and physiology. At a molecular level, these oscillations are generated by transcriptional-translational feedback loops composed of core clock genes. In turn, core clock genes drive the rhythmic accumulation of downstream outputs-termed clock-controlled genes (CCGs)-whose rhythmic translation and function ultimately underlie daily oscillations at a cellular and organismal level. Given the circadian clock's profound influence on human health and behavior, considerable efforts have been made to systematically identify CCGs. The recent development of next-generation sequencing has dramatically expanded our ability to study the expression, processing, and stability of rhythmically expressed mRNAs. Nevertheless, like any new technology, there are many technical issues to be addressed. Here, we discuss considerations for studying circadian rhythms using genome scale transcriptional profiling, with a particular emphasis on RNA sequencing. We make a number of practical recommendations-including the choice of sampling density, read depth, alignment algorithms, read-depth normalization, and cycling detection algorithms-based on computational simulations and our experience from previous studies. We believe that these results will be of interest to the circadian field and help investigators design experiments to derive most values from these large and complex data sets.

In Vivo Single-Cell Detection of Metabolic Oscillations in Stem Cells

Through the use of bulk measurements in metabolic organs, the circadian clock was shown to play roles in organismal energy homeostasis. However, the relationship between metabolic and circadian oscillations has not been studied invivo at a single-cell level. Also, it is unknown whether the circadian clock controls metabolism in stem cells. We used a sensitive, noninvasive method to detect metabolic oscillations and circadian phase within epidermal stem cells in live mice at the single-cell level. We observe a higher NADH/NAD+ ratio, reflecting an increased glycolysis/oxidative phosphorylation ratio during the night compared to the day. Furthermore, we demonstrate that single-cell metabolic heterogeneity within the basal cell layer correlates with the circadian clock and that diurnal fluctuations in NADH/NAD+ ratio are Bmal1 dependent. Our data show that, in proliferating stem cells, the circadian clock coordinates activities of oxidative phosphorylation and glycolysis with DNA synthesis, perhaps as a protective mechanism against genotoxicity.

Hepatocyte circadian clock controls acetaminophen bioactivation through NADPH-cytochrome P450 oxidoreductase.

The diurnal variation in acetaminophen (APAP) hepatotoxicity (chronotoxicity) reportedly is driven by oscillations in metabolism that are influenced by the circadian phases of feeding and fasting. To determine the relative contributions of the central clock and the hepatocyte circadian clock in modulating the chronotoxicity of APAP, we used a conditional null allele of brain and muscle Arnt-like 1 (Bmal1, aka Mop3 or Arntl) allowing deletion of the clock from hepatocytes while keeping the central and other peripheral clocks (e.g., the clocks controlling food intake) intact. We show that deletion of the hepatocyte clock dramatically reduces APAP bioactivation and toxicity in vivo and in vitro because of a reduction in NADPH-cytochrome P450 oxidoreductase gene expression, protein, and activity.

The effects of different light-dark cycles on the metabolism of the diazotrophic, unicellular cyanobacteria Cyanothece sp. ATCC 51142, and Cyanothecesp. PCC 7822.

The diazotrophic unicellular cyanobacterium Cyanothece sp. ATCC 51142 demonstrates circadian patterns in nitrogenase activity, H2 production and glycogen storage when grown under nitrogen-fixing, 12:12 light:dark (L:D) conditions. In this study, we grew Cyanothece sp. ATCC 51142, and another strain in this genus, Cyanothece sp. PCC 7822, under long-day (16:8 L:D) and short-day (8:16 L:D) nitrogen-fixing conditions to determine if they continued to display circadian rhythms. Both strains demonstrated similar circadian patterns for all three metabolic parameters when grown under long-day conditions. However, the strains responded differently to short-day growth conditions. Cyanothece sp. ATCC 51142 retained reasonable circadian patterns under 8:16 L:D conditions, whereas Cyanothece sp. PCC 7822 had quite damped patterns without a clear circadian pattern. In particular, glycogen storage changed very little throughout the day and we ascribe this to the difference in the type of glycogen granules in Cyanothece sp. PCC 7822 which has small β-granules, compared to the large, starch-like granules in Cyanothece sp. ATCC 51142. The results suggested that both mechanistic and regulatory processes play a role in establishing the basis for these metabolic oscillations.

Calcium and Metabolic Oscillations in Pancreatic Islets: Who's Driving the Bus?*

Pancreatic islets exhibit bursting oscillations in response to elevated blood glucose. These oscillations are accompanied by oscillations in the free cytosolic Ca2+ concentration (Cac ), which drives pulses of insulin secretion. Both islet Ca2+ and metabolism oscillate, but there is some debate about their interrelationship. Recent experimental data show that metabolic oscillations in some cases persist after the addition of diazoxide (Dz), which opens K(ATP) channels, hyperpolarizing β-cells and preventing Ca2+ entry and Ca2+ oscillations. Further, in some islets in which metabolic oscillations were eliminated with Dz, increasing the cytosolic Ca2+ concentration by the addition of KCl could restart the metabolic oscillations. Here we address why metabolic oscillations persist in some islets but not others, and why raising Cac restarts oscillations in some islets but not others. We answer these questions using the dual oscillator model (DOM) for pancreatic islets. The DOM can reproduce the experimental data and shows that the model supports two different mechanisms for slow metabolic oscillations, one that requires calcium oscillations and one that does not.

Circadian Metabolic Oscillations in the Epidermis Stem Cells by Fluorescence Lifetime Microscopy of NADH in Vivo

138-Plat Circadian Metabolic Oscillations in the Epidermis Stem Cells by Fluores- cence Lifetime Microscopy of NADH in Vivo Chiara Stringari 1,2 , Mikhail Geyfman 1 , Hong Wang 1,3 , Viera Crosignani 1 , Vivek Kumar 4 , Joseph S. Takahashi 4 , Bogi Andersen 1 , Enrico Gratton 1 . University California Irvine, Irvine, CA, USA, 2 Laboratory for Optics and Biosciences, E´cole Polytechnique, Paris, France, 3 China Agricultural University, Beijing, China, 4 University of Texas Southwestern Medical Center, Dallas, TX, USA. There is a lack of non-invasive methods to monitor circadian metabolic oscil- lations of single cells in their native environment. Here we implement a label- free method using NADH as an intrinsic biomarker and the Phasor approach to Fluorescence Lifetime microscopy to measure the metabolic optical finger- print of single cells during the day-night cycle. For the first time we detect in vivo metabolic circadian oscillations within the stem cells of the epidermis layer. We observe higher ratios of free/bound NADH, i.e. NADH/NAD þ , in the night with respect to the day. This difference indicates a glycolytic pheno- type associated with high proliferation during the night and an oxidative phos- phorylation phenotype associated with low proliferation during the day. We demonstrate that cell-to cell metabolic heterogeneity correlates with circadian phase as measured within the basal epidermal layer by Per1-Venus reporter assay. Finally, we show that NADH metabolic oscillations are Bmal1 dependent. Work supported with NIH grants P50 GM076516 and P41 GM103540.

Ca2+ has a Permissive Effect on Glycolytic Oscillations in Pancreatic Beta Cells

The secretion of insulin from pancreatic islet beta cells is pulsatile (∼5 min period) and reflects upstream oscillations in metabolism and Ca2+. Of the oscillating pathways in the beta cell, glycolytic oscillations are the least well studied. We recently developed a FRET biosensor for pyruvate kinase M2 activity (PKAR, Pyruvate Kinase Activity Reporter), which is activated by fructose 1,6-bisphosphate. We used PKAR FRET to measure oscillations in glycolysis stimulated by 10 mM glucose. After abolishing Ca2+ oscillations with diazoxide (Dz), the oscillations in PKAR were terminated. However, in some cases the oscillations could be restored by raising extracellular KCl, which elevated the intracellular Ca2+ levels but did not restore Ca2+ oscillations. These results indicate that glycolytic oscillations can persist in the absence of Ca2+ oscillations, and suggest the presence of a Ca2+ threshold that is permissive of glycolytic oscillations. To address this question, we varied the extracellular Ca2+ from 1.25 to 5 mM, which increased the amplitude of intracellular Ca2+ oscillations in a dose-dependent manner. Parallel measurements using PKAR indicated that the amplitude of glycolytic oscillations in pyruvate kinase M2 activity are augmented by intracellular Ca2+, however PKAR oscillations were terminated if extracellular Ca2+ dropped too low. Taken together, these results indicate that glycolytic oscillations in beta cells are dependent upon on a threshold level of intracellular Ca2+ level, and represent a distinct oscillating compartment of the cell. Supported by R01DK46409 (L.S.).

Electrical potentials and its fluctuations in nodal cells of nitellopsis obtusa

Using the liquid microelectrode technique, the electrical potentials of nodal cells and the with them internodal cells neighbouring, of <i>Nitellopsis obtusa</i> plants of the <i>Characeae</i> family were determined. A specific variation in the potential of nodal cells in the form of cacophony-type irregular disturbances was also found, indicating that the node is not a passive mechanical element controlled by internodal cells, but is a complex system of coupled-, and different metabolic oscillators.