Free Calcium Concentration Research Articles

Canonical Transient Receptor Potential (TRPC) Channels in Cardiovascular Pathology and their Modulators.

Ion channels play a crucial role in various aspects of cardiac function, such as regulating rhythm and contractility. As a result, they serve as key targets for therapeutic interventions in cardiovascular diseases. Cell function is substantially influenced by the concentration of free cytosolic calcium (Ca2+) and the voltage across the plasma membrane. These characteristics are known to be regulated by Ca2+-permeable non-selective cationic channels, although our knowledge of these channels is still inadequate. The transient receptor potential (TRP) superfamily comprises of many non-selective cation channels with diverse Ca2+ permeability. Canonical or classical TRP (TRPC) channels are a subgroup of the TRP superfamily that are expressed ubiquitously in mammalian cells. TRPC channels are multidimensional signalling protein complexes that play essential roles in a variety of physiological and pathological processes in humans, including cancer, neurological disorders, cardiovascular diseases, and others. The objective of this article is to focus on the role that TRPC channels play in the cardiovascular system. The role of TRPC channels will be deeply discussed in cardiovascular pathology. Together, a critical element in developing novel treatments that target TRPC channels is comprehending the molecular mechanisms and regulatory pathways of TRPC channels in related cardiovascular diseases and conditions.

Myogenic response of the cerebral vessel: a mathematical model

The myogenic response is most typical for small arterial vessels and consists in constriction of the vessel with an increase in transmural pressure, the pressure difference between the inner and outer surfaces of the vascular wall. There are many models describing this response. In models of last decades, in addition to macrostretchings and macrostrains, their microanalogues are introduced into consideration, that significantly complicates the description. In the presented work a simple model of the wall of the small cerebral artery is described, which includes the main parameters that control the myogenic response and corresponds to physiological processes underlining this response. It is believed that active stresses depend on stretching and the concentration of free calcium ions in the cytoplasm of the smooth muscle cell. In turn, the calcium content depends on the membrane potential, determined by the pressure. An axisymmetric stationary problem is considered; the wall material in the non-activated state is assumed to be pseudoelastic. Calculations showed a qualitative difference in the dependence of active stress on stretching and its distribution in the radial direction before and after the development of the vascular response. The calculated diameter values before and after exposure to substances affecting the polarization of the smooth muscle cell membrane are within the range of experimental values. These results indicate that the introduction of the dependence of calcium concentration on membrane potential expands the area of applicability of simple models of this kind for evaluating the value of vascular diameter.

Deciphering simplified regional anticoagulation with citrate in intermittent hemodialysis: a clinical and computational study

Regional citrate anticoagulation use in intermittent hemodialysis is limited by the increased risk of metabolic complications due to faster solute exchanges than with continuous renal replacement therapies. Several simplifications have been proposed. The objective of this study was to validate a mathematical model of hemodialysis anticoagulated with citrate that was then used to evaluate different prescription scenarios on anticoagulant effectiveness (free calcium concentration in dialysis filter) and calcium balance. A study was conducted in hemodialyzed patients with a citrate infusion into the arterial line and a 1.25 mmol/L calcium dialysate. Calcium and citrate concentrations were measured upstream and downstream of the citrate infusion site and in the venous line. The values measured in the venous lines were compared with those predicted by the model using Bland and Altman diagrams. The model was then used with 22 patients to make simulations. The model can predict the concentration of free calcium, bound to citrate or albumin, accurately. Irrespective of the prescription scenario a decrease in free calcium below 0.4 mmol/L was obtained only in a fraction of the dialysis filter. A zero or slightly negative calcium balance was observed, and should be taken into account in case of prolonged use.

Purinergic agonists increase [Ca2+]i in rat conjunctival goblet cells through ryanodine receptor type 3.

ATP and benzoylbenzoyl-ATP (BzATP) increase free cytosolic Ca2+ concentration ([Ca2+]i) in conjunctival goblet cells (CGCs) resulting in mucin secretion. The purpose of this study was to investigate the source of the Ca2+i mobilized by ATP and BzATP. First-passage cultured rat CGCs were incubated with Fura-2/AM, and [Ca2+]i was measured under several conditions with ATP and BzATP stimulation. The following conditions were used: 1) preincubation with the Ca2+ chelator EGTA, 2) preincubation with the SERCA inhibitor thapsigargin (10-6 M), which depletes ER Ca2+ stores, 3) preincubation with phospholipase C (PLC) or protein kinase A (PKA) inhibitor, or 4) preincubation with the voltage-gated calcium channel antagonist nifedipine (10-5 M) and the ryanodine receptor (RyR) antagonist dantrolene (10-5 M). Immunofluorescence microscopy (IF) and quantitative reverse transcription polymerase chain reaction (RT-qPCR) were used to investigate RyR presence in rat and human CGCs. ATP-stimulated peak [Ca2+]i was significantly lower after chelating Ca2+i with 2 mM EGTA in Ca2+-free buffer. The peak [Ca2+]i increase in CGCs preincubated with thapsigargin, the PKA inhibitor H89, nifedipine, and dantrolene, but not the PLC inhibitor, was reduced for ATP at 10-5 M and BzATP at 10-4 M. Incubating CGCs with dantrolene alone decreased [Ca2+]i and induced CGC cell death at a high concentration. RyR3 was detected in rat and human CGCs with IF and RT-qPCR. We conclude that ATP- and BzATP-induced Ca2+i increases originate from the ER and that RyR3 may be an essential regulator of CGC [Ca2+]i. This study contributes to the understanding of diseases arising from defective Ca2+ signaling in nonexcitable cells.NEW & NOTEWORTHY ATP and benzoylbenzoyl-ATP (BzATP) induce mucin secretion through an increase in free cytosolic calcium concentration ([Ca2+]i) in conjunctival goblet cells (CGCs). The mechanisms through which ATP and BzATP increase [Ca2+]i in CGCs are unclear. Ryanodine receptors (RyRs) are fundamental in [Ca2+]i regulation in excitable cells. Herein, we find that ATP and BzATP increase [Ca2+]i through the activation of protein kinase A, voltage-gated calcium channels, and RyRs, and that RyRs are crucial for nonexcitable CGCs' Ca2+i homeostasis.

The Influence of Polygonatum King ilium on the Activity of Calcium-Regulating Enzymes and Calcium Ion Concentration in SH-SY5Y Cells Damaged by Aβ25-35 In Vitro

Objective To explore the effects of Polygonatum King ilium and Earthworm Formula (RPEWM) on the activity of calcium-regulating enzymes and free calcium ion (Ca2+) concentration in SH-SY5Y cells with Aβ damage in vitro. Methods SH-SY5Y cells were induced to differentiate with retinoic acid, then damaged by Aβ to mimic Alzheimer's disease (AD) injury. RPEWM extract was added during the AD injury to protect the cells, and its effects on cell proliferation, apoptosis, and the activity of cellular Ca2+-ATPase､Ca2+-Mg2+- ATPase ,as well as free calcium ion concentration were observed. Results RPEWM extract significantly protected cells from AD injury, resisted apoptosis, and promoted cell proliferation; it increased the activity of intracellular Ca2+-ATPase､Ca2+-Mg2+- ATPase, reduced Ca2+ concentration, and showed a dose-dependent relationship. Conclusion RPEWM has a good protective effect on differentiated SH-SY5Y cells with Aβ25-35damage and has a certain therapeutic effect on AD.

Sex-dependent intra-islet structural rearrangements affecting alpha-to-beta cell interactions lead to adaptive enhancements of Ca2+ dynamics in prediabetic beta cells

Aims/hypothesisPrediabetic pancreatic beta cells can adapt their function to maintain normoglycaemia for a limited period of time, after which diabetes mellitus will manifest upon beta cell exhaustion. Understanding sex-specific beta cell compensatory mechanisms and their failure in prediabetes (impaired glucose tolerance) is crucial for early disease diagnosis and individualised treatment. Our aims were as follows: (1) to determine the key time points of the progression from beta cells’ functional adaptations to their failure in vivo; and (2) to mechanistically explain in vivo sex-specific beta cell compensatory mechanisms and their failure in prediabetes.MethodsIslets from male and female transgenic Ins1CreERT2-GCaMP3 mice were transplanted into the anterior chamber of the eye of 10- to 12-week-old sex-matched C57BL/6J mice. Recipient mice were fed either a control diet (CD) or western diet (WD) for a maximum of 4 months. Metabolic variables were evaluated monthly. Beta cell cytoplasmic free calcium concentration ([Ca2+]i) dynamics were monitored in vivo longitudinally by image fluorescence of the GCaMP3 reporter islets. Global islet beta cell [Ca2+]i dynamics in line with single beta cell [Ca2+]i analysis were used for beta cell coordination studies. The glucagon receptor antagonist L-168,049 (4 mmol/l) was applied topically to the transplanted eyes to evaluate in vivo the effect of glucagon on beta cell [Ca2+]idynamics. Human islets from non-diabetic women and men were cultured for 24 h in either a control medium or high-fat/high-glucose medium in the presence or absence of the glucagon receptor antagonist L-168,049. [Ca2+]i dynamics of human islets were evaluated in vitro after 1 h exposure to Fura-10.ResultsMice fed a WD for 1 month displayed increased beta cell [Ca2+]i dynamics linked to enhanced insulin secretion as a functional compensatory mechanism in prediabetes. Recruitment of inactive beta cells in WD-fed mice explained the improved beta cell function adaptation observed in vivo; this occurred in a sex-specific manner. Mechanistically, this was attributable to an intra-islet structural rearrangement involving alpha cells. These sex-dependent cytoarchitecture reorganisations, observed in both mice and humans, induced enhanced paracrine input from adjacent alpha cells, adjusting the glucose setpoint and amplifying the insulin secretion pathway. When WD feeding was prolonged, female mice maintained the adaptive mechanism due to their intrinsically high proportion of alpha cells. In males, [Ca2+]i dynamics progressively declined subsequent to glucose stimulation while insulin secretion continue to increase, suggesting uncoordinated beta cell function as an early sign of diabetes.Conclusions/interpretationWe identified increased coordination of [Ca2+]i dynamics as a beta cell functional adaptation mechanisms in prediabetes. Importantly, we uncovered the mechanisms by which sex-dependent beta cell [Ca2+]i dynamics coordination is orchestrated by an intra-islet structure reorganisation increasing the paracrine input from alpha cells on beta cell function. Moreover, we identified reduced [Ca2+]i dynamics coordination in response to glucose as an early sign of diabetes preceding beta cell secretory dysfunction, with males being more vulnerable. Alterations in coordination capacity of [Ca2+]i dynamics may thus serve as an early marker for beta cell failure in prediabetes.Graphical

The influence of cation exchange on the possible mechanism of erionite toxicity: A synchrotron-based micro-X-ray fluorescence study on THP-1-derived macrophages exposed to erionite-Na

Fibrous erionite is the only zeolite classified as Group 1 carcinogen by the International Agency for Research on Cancer (IARC). Carcinogenesis induced by erionite is thought to involve several factors as biopersistence, the iron role and cation exchange processes. To better understand these mechanisms, a detailed investigation at the micro scale was performed, collecting elemental information on iron and cation release and their distribution in biological systems by synchrotron micro-X-ray fluorescence mapping (SR-micro-XRF) and synchrotron micro-X-ray absorption spectroscopy (SR-micro-XANES) at the TwinMic beamline (Elettra synchrotron) and at the ID21 beamline of the European Synchrotron Radiation Facility (ESRF). By microscopy and chemical mapping, highly detailed maps of the chemical and morphological interaction of biological systems with fibres could be produced. In detail, THP-1 cell line derived macrophages, used as in vitro model, were analysed during erionite-Na phagocytosis at different time intervals, after single dose exposure. For comparison, cellular fluorescent probes were also used to evaluate the intracellular free sodium and calcium concentrations. Synchrotron analyses visualised the spatial distribution of both fibre and mineral particle associated metals during the phagocytosis, describing the mechanism of internalisation of erionite-Na and its accessory mineral phases. The intracellular distribution of metals and other cations was mapped to evaluate metal release, speciation changes and/or cation exchange during phagocytosis. The fluorescent probes complemented microchemical data clarifying, and confirming, the cation distribution observed in the SR-micro-XRF maps. The significant cytoplasmic calcium decrease, and the concomitant sodium increase, after the fibre phagocytosis seemed due to activation of plasma membrane cations exchangers triggered by the internalisation while, surprisingly, the ion-exchange capacity of erionite-Na could play a minor role in the disruption of the two cations intracellular homeostasis. These results help to elucidate the role of cations in the toxicity of erionite-treated THP-1 macrophages and add knowledge to its carcinogenicity process.

Progress of researches on mechanisms of acupuncture underlying improvement of cerebral ischemia-reperfusion injury by regulating calcium overload.

Ischemic stroke is currently the most common type of stroke, and the key pathological link is cerebral ischemia-reperfusion injury (CIRI), while the key factor leading to apoptosis and necrosis of ischemic nerve cells is calcium overload. Current studies have confirmed that acupuncture therapy has a good modulating effect on calcium homeostasis and can reduce cerebral ischemia-reperfusion induced damage of neuronal cells by inhibiting calcium overload. After reviewing the relevant literature published in the past 15 years, we find that acupuncture plays a role in regulating the pathological mechanism of calcium overload after CIRI by inhibiting the opening of connexin 43 hemichannels, regulating the intracellular free calcium ion concentration, suppressing the expression of calmodulin, and blocking the function of L-type voltage-gated calcium channels, thereby inhibiting calcium overload, regulating calcium homeostasis and antagonizing neuronal damage resulted from cerebral ischemia-reperfusion, which may provide ideas for future research.

Adsorption Isotherm and Mechanism of Ca2+ Binding to Polyelectrolyte.

Polyelectrolytes, such as poly(acrylic acid) (PAA), can effectively mitigate CaCO3 scale formation. Despite their success as antiscalants, the underlying mechanism of binding of Ca2+ to polyelectrolyte chains remains unresolved. Through all-atom molecular dynamics simulations, we constructed an adsorption isotherm of Ca2+ binding to sodium polyacrylate (NaPAA) and investigated the associated binding mechanism. We find that the number of calcium ions adsorbed [Ca2+]ads to the polymer saturates at moderately high concentrations of free calcium ions [Ca2+]aq in the solution. This saturation value is intricately connected with the binding modes accessible to Ca2+ ions when they bind to the polyelectrolyte chain. We identify two dominant binding modes: the first involves binding to at most two carboxylate oxygens on a polyacrylate chain, and the second, termed the high binding mode, involves binding to four or more carboxylate oxygens. As the concentration of free calcium ions [Ca2+]aq increases from low to moderate levels, the polyelectrolyte chain undergoes a conformational transition from an extended coil to a hairpin-like structure, enhancing the accessibility to the high binding mode. At moderate concentrations of [Ca2+]aq, the high binding mode accounts for at least one-third of all binding events. The chain's conformational change and its consequent access to the high binding mode are found to increase the overall Ca2+ ion binding capacity of the polyelectrolyte chain.

Model-level Weight Update Domain Adaptive Dynamic CNN Soft Sensor for Free Calcium Ion Concentration in Cement Clinker

The paramount determinant of clinker quality within the cement clinker production process resides in the precise prediction of F-CAO (free calcium ion) content. Given the intricate nature of cement clinker production, a profound coupling and temporal progression are inherently intertwined between the process data, as working conditions undergo gradual transformations over time. This research suggests a Model-level Weight Update Domain Adaptive Dynamic CNN Soft Sensor for Free Calcium Ion Concentration in Cement Clinker to overcome the aforementioned issues. We introduce a graph-structured two-dimensional methodology aimed at transforming both labeled and unlabeled data within the original process dataset into two-dimensional process data blocks. This innovative approach serves to enhance the spatial and temporal attributes of the labeled samples within the comprehensive sample set. Importantly, this methodology effectively addresses the intricate challenges posed by robust data coupling and the substantial presence of unlabeled samples. In response to the challenge posed by the frequent temporal fluctuations in working conditions, we introduce the Model-level Weight Update Domain Adaptive (MWUDA) approach. MWUDA leverages the hidden layer outputs from both the source and target domains to optimize the weight updates of the fully-connected layer within the CNN model. This strategic weight optimization is aimed at ameliorating the performance degradation observed in static models when confronted with shifting patterns in sample distributions. Subsequently, a limited subset of source domain samples exhibiting analogous regularities to those in the test samples is identified, enabling the acquisition of refined weights through the MWUDA method. Employing this strategy safeguards the precision of model updates throughout the MWUDA computation procedure, ensuring a robust and accurate adaptation process. The effectiveness of the proposed method is verified in an experimental study with real cement production data.

Cortical neurons obtained from patient-derived iPSCs with GNAO1 p.G203R variant show altered differentiation and functional properties

Pathogenic variants in the GNAO1 gene, encoding the alpha subunit of an inhibitory heterotrimeric guanine nucleotide-binding protein (Go) highly expressed in the mammalian brain, have been linked to encephalopathy characterized by different combinations of neurological symptoms, including developmental delay, hypotonia, epilepsy and hyperkinetic movement disorder with life-threatening paroxysmal exacerbations. Currently, there are only symptomatic treatments, and little is known about the pathophysiology of GNAO1-related disorders. Here, we report the characterization of a new in vitro model system based on patient-derived induced pluripotent stem cells (hiPSCs) carrying the recurrent p.G203R amino acid substitution in Gαo, and a CRISPR-Cas9-genetically corrected isogenic control line. RNA-Seq analysis highlighted aberrant cell fate commitment in neuronal progenitor cells carrying the p.G203R pathogenic variant. Upon differentiation into cortical neurons, patients’ cells showed reduced expression of early neural genes and increased expression of astrocyte markers, as well as premature and defective differentiation processes leading to aberrant formation of neuronal rosettes. Of note, comparable defects in gene expression and in the morphology of neural rosettes were observed in hiPSCs from an unrelated individual harboring the same GNAO1 variant. Functional characterization showed lower basal intracellular free calcium concentration ([Ca2+]i), reduced frequency of spontaneous activity, and a smaller response to several neurotransmitters in 40- and 50-days differentiated p.G203R neurons compared to control cells. These findings suggest that the GNAO1 pathogenic variant causes a neurodevelopmental phenotype characterized by aberrant differentiation of both neuronal and glial populations leading to a significant alteration of neuronal communication and signal transduction.

Angiotensin II-dependent aldosterone production in the adrenal cortex.

The adrenal cortex is responsible for production of adrenal steroid hormones and is anatomically divided into three distinct zones: zona glomerulosa secreting mineralocorticoids (mainly aldosterone), zona fasciculata secreting glucocorticoids (cortisol), and zona reticularis producing androgens. Importantly, due to their high lipophilicity, no adrenal steroid hormone (including aldosterone) is stored in vesicles but rather gets synthesized and secreted instantly upon cell stimulation with specific stimuli. Aldosterone is the most potent mineralocorticoid hormone produced from the adrenal cortex in response to either angiotensin II (AngII) or elevated K+ levels in the blood (hyperkalemia). AngII, being a peptide, cannot cross cell membranes and thus, uses two distinct G protein-coupled receptor (GPCR) types, AngII type 1 receptor (AT1R) and AT2R to exert its effects inside cells. In zona glomerulosa cells, AT1R activation by AngII results in aldosterone synthesis and secretion via two main pathways: (a) Gq/11 proteins that activate phospholipase C ultimately raising intracellular free calcium concentration; and (b) βarrestin1 and -2 (also known as Arrestin-2 and -3, respectively) that elicit sustained extracellular signal-regulated kinase (ERK) activation. Both pathways induce upregulation and acute activation of StAR (steroidogenic acute regulatory) protein, the enzyme that catalyzes the rate-limiting step in aldosterone biosynthesis. This chapter describes these two salient pathways underlying AT1R-induced aldosterone production in zona glomerulosa cells. We also highlight some pharmacologically important notions pertaining to the efficacy of the currently available AT1R antagonists, also known as angiotensin receptor blockers (ARBs) or sartans at suppressing both pathways, i.e., their inverse agonism efficacy at G proteins and βarrestins.

Zn2+ is essential for Ca2+ oscillations in mouse eggs.

Changes in the intracellular concentration of free calcium (Ca2+) underpin egg activation and initiation of development in animals and plants. In mammals, the Ca2+ release is periodical, known as Ca2+ oscillations, and mediated by the type 1 inositol 1,4,5-trisphosphate receptor (IP3R1). Another divalent cation, zinc (Zn2+), increases exponentially during oocyte maturation and is vital for meiotic transitions, arrests, and polyspermy prevention. It is unknown if these pivotal cations interplay during fertilization. Here, using mouse eggs, we showed that basal concentrations of labile Zn2+ are indispensable for sperm-initiated Ca2+ oscillations because Zn2+-deficient conditions induced by cell-permeable chelators abrogated Ca2+ responses evoked by fertilization and other physiological and pharmacological agonists. We also found that chemically or genetically generated eggs with lower levels of labile Zn2+ displayed reduced IP3R1 sensitivity and diminished ER Ca2+ leak despite the stable content of the stores and IP3R1 mass. Resupplying Zn2+ restarted Ca2+ oscillations, but excessive Zn2+ prevented and terminated them, hindering IP3R1 responsiveness. The findings suggest that a window of Zn2+ concentrations is required for Ca2+ responses and IP3R1 function in eggs, ensuring optimal response to fertilization and egg activation.

Zn2+ is essential for Ca2+ oscillations in mouse eggs

Changes in the intracellular concentration of free calcium (Ca2+) underpin egg activation and initiation of development in animals and plants. In mammals, the Ca2+ release is periodical, known as Ca2+ oscillations, and mediated by the type 1 inositol 1,4,5-trisphosphate receptor (IP3R1). Another divalent cation, zinc (Zn2+), increases exponentially during oocyte maturation and is vital for meiotic transitions, arrests, and polyspermy prevention. It is unknown if these pivotal cations interplay during fertilization. Here, using mouse eggs, we showed that basal concentrations of labile Zn2+ are indispensable for sperm-initiated Ca2+ oscillations because Zn2+-deficient conditions induced by cell-permeable chelators abrogated Ca2+ responses evoked by fertilization and other physiological and pharmacological agonists. We also found that chemically or genetically generated eggs with lower levels of labile Zn2+ displayed reduced IP3R1 sensitivity and diminished ER Ca2+ leak despite the stable content of the stores and IP3R1 mass. Resupplying Zn2+ restarted Ca2+ oscillations, but excessive Zn2+ prevented and terminated them, hindering IP3R1 responsiveness. The findings suggest that a window of Zn2+ concentrations is required for Ca2+ responses and IP3R1 function in eggs, ensuring optimal response to fertilization and egg activation.

Analyzing fuzzy boundary value problems: a study on the influence of mitochondria and ER fluxes on calcium ions in neuron cells.

Cytosolic-free calcium ions play an important role in various physical and physiological processes. A vital component of neural signaling is the free calcium ion concentration often known as the second messenger. There are many parameters that effect the cytosolic free calcium concentration like buffer, voltage-gated ion channels, Endoplasmic reticulum, Mitochondria, etc. Mitochondria are small organelles located within thenervous system that are involved in processes within cells such as calcium homeostasis management, energy generation, response to stress, and cell demise pathways. In this work, a mathematical model with fuzzy boundary values has been developed to study the effect of Mitochondria and ER fluxes on free Calcium ions. The intended findings are displayed utilizingthe physiological understanding that amyloid beta plaques and tangles of neurofibrillary fibers have been identified as the two main causes of AD. The key conclusion of the work is the investigation of [Formula: see text] for healthy cells and cells affected by Alzheimer's disease, which may aid in the study of such processes for computational scientists and medical practitioners. Also, it has been shown that when a unique solution is found for a specific precise problem, it also successfully deals with any underlying ambiguity within the problem by utilizinga technique based on the principles of linear transformation. Furthermore, the comparison between the analytical approach and the generalized hukuhara derivative approach is shown here, which illustrates the benefits of the analytical approach. The simulation is carried out in MATLAB.

Relations between Glucose and d-Amino Acids in the Modulation of Biochemical and Functional Properties of Rodent Islets of Langerhans.

The cell-to-cell signaling role of d-amino acids (d-AAs) in the mammalian endocrine system, particularly in the islets of Langerhans, has drawn growing interest for their potential involvement in modulating glucose metabolism. Previous studies found colocalization of serine racemase [produces d-serine (d-Ser)] and d-alanine (d-Ala) within insulin-secreting beta cells and d-aspartate (d-Asp) within glucagon-secreting alpha cells. Expressed in the islets, functional N-methyl-d-aspartate receptors are involved in the modulation of glucose-stimulated insulin secretion and have binding sites for several d-AAs. However, knowledge of the regulation of d-AA levels in the islets during glucose stimulation as well as the response of islets to different levels of extracellular d-AAs is limited. In this study, we determined the intracellular and extracellular levels of d-Ser, d-Ala, and d-Asp in cultures of isolated rodent islets exposed to different levels of extracellular glucose. We found that the intracellular levels of the enantiomers demonstrated large variability and, in general, were not affected by extracellular glucose levels. However, significantly lower levels of extracellular d-Ser and d-Ala were observed in the islet media supplemented with 20 mM concentration of glucose compared to the control condition utilizing 3 mM glucose. Glucose-induced oscillations of intracellular free calcium concentration ([Ca2+]i), a proxy for insulin secretion, were modulated by the exogenous application of d-Ser and d-Ala but not by their l-stereoisomers. Our results provide new insights into the roles of d-AAs in the biochemistry and function of pancreatic islets.

Calcium regulates cortex contraction in Physarum polycephalum

The tubular network-forming slime mold Physarum polycephalum is able to maintain long-scale contraction patterns driven by an actomyosin cortex. The resulting shuttle streaming in the network is crucial for the organism to respond to external stimuli and reorganize its body mass giving rise to complex behaviors. However, the chemical basis of the self-organized flow pattern is not fully understood. Here, we present ratiometric measurements of free intracellular calcium in simple morphologies of Physarum networks. The spatiotemporal patterns of the free calcium concentration reveal a nearly anti-correlated relation to the tube radius, suggesting that calcium is indeed a key regulator of the actomyosin activity. We compare the experimentally observed phase relation between the radius and the calcium concentration to the predictions of a theoretical model including calcium as an inhibitor. Numerical simulations of the model suggest that calcium indeed inhibits the contractions in Physarum, although a quantitative difference to the experimentally measured phase relation remains. Unraveling the mechanism underlying the contraction patterns is a key step in gaining further insight into the principles of Physarum’s complex behavior.

Studying the role of random translocation of GLUT4 in cardiomyocytes on calcium oscillations

Defective excitation-contraction coupling (ECC) is one of the causes of contractile dysfunction linked to diabetic cardiomyopathy (DCM). This defect in ECC is mainly caused by the dysregulation of the calcium homeostasis in normal cardiomyocytes due to prolonged elevated blood glucose levels. Different types of channels inside the cardiomyocytes help maintain calcium homeostasis in the cell. Free cytosolic calcium concentration changes are one of several variables that affect complicated physiological cardiac processes and significantly impact proper heart functioning. Glucose concentration in the cardiac cell also plays a vital role in altering the calcium homeostasis in the DCM condition. In the current study, we develop a mathematical model that explicitly represents many of the known signalling components mediating translocation of the insulin-responsive glucose transporter type 4 (GLUT4) to gain insight into the complexities of metabolic insulin signalling pathways in cardiac cells. We examined the effect of the random movement of GLUT4 responsible for the entry of glucose in the cardiomyocytes. In non-paced cardiac cells calcium remains in the steady state in the absence of external pacing and avoid oscillation for healthy cardiomyocyte functioning. Intracellular calcium transients were observed during action potential. We investigated the circumstances under which the system transits from a stable state to an oscillatory state. We studied the effect of random translocation of GLUT4 on calcium oscillation. By altering system parameters, we induced insulin resistance in cardiomyocytes to mimic diabetic conditions and proposed some potential restoration strategies to restore normal calcium dynamics. Early bifurcation was observed with the introduction of randomness in the system.

Synergistic Interaction of 5-HT1B and 5-HT2B Receptors in Cytoplasmic Ca2+ Regulation in Human Umbilical Vein Endothelial Cells: Possible Involvement in Pathologies.

The aim of this work was to explore the involvement of 5-HT1B and 5-HT2B receptors (5-HT1BR and 5-HT2BR) in the regulation of free cytoplasmic calcium concentration ([Ca2+]i) in human umbilical vein endothelial cells (HUVEC). We have shown by quantitative PCR analysis, that 5-HT1BR and 5-HT2BR mRNAs levels are almost equal in HUVEC. Immunofluorescent staining demonstrated, that 5-HT1BR and 5-HT2BR are expressed both in plasma membrane and inside the cells. Intracellular 5-HT1BR are localized mainly in the nuclear region, whereas 5-HT2BR receptors are almost evenly distributed in HUVEC. 5-HT, 5-HT1BR agonist CGS12066B, or 5-HT2BR agonist BW723C86 added to HUVEC caused a slight increase in [Ca2+]i, which was much lower than that of histamine, ATP, or SFLLRN, an agonist of protease-activated receptors (PAR1). However, activation of 5-HT1BR with CGS12066B followed by activation of 5-HT2BR with BW723C86 manifested a synergism of response, since several-fold higher rise in [Ca2+]i occurred. CGS12066B caused more than a 5-fold increase in [Ca2+]i rise in HUVEC in response to 5-HT. This 5-HT induced [Ca2+]i rise was abolished by 5-HT2BR antagonist RS127445, indicating that extracellular 5-HT acts through 5-HT2BR. Synergistic [Ca2+]i rise in response to activation of 5-HT1BR and 5-HT2BR persisted in a calcium-free medium. It was suppressed by the phospholipase C inhibitor U73122 and was not inhibited by the ryanodine and NAADP receptors antagonists dantrolene and NED-19. [Ca2+]i measurements in single cells demonstrated that activation of 5-HT2BR alone by BW723C86 caused single asynchronous [Ca2+]i oscillations in 19.8 ± 4.2% (n = 3) of HUVEC that occur with a long delay (66.1 ± 4.3 s, n = 71). On the contrary, histamine causes a simultaneous and almost immediate increase in [Ca2+]i in all the cells. Pre-activation of 5-HT1BR by CGS12066B led to a 3-4 fold increase in the number of HUVEC responding to BW723C86, to synchronization of their responses with a delay shortening, and to the bursts of [Ca2+]i oscillations in addition to single oscillations. In conclusion, to get a full rise of [Ca2+]i in HUVEC in response to 5-HT, simultaneous activation of 5-HT1BR and 5-HT2BR is required. 5-HT causes an increase in [Ca2+]i via 5-HT2BR while 5-HT1BR could be activated by the membrane-permeable agonist CGS12066B. We hypothesized that CGS12066B acts via intracellular 5-HT1BR inaccessible to extracellular 5-HT. Intracellular 5-HT1BR might be activated by 5-HT which could be accumulated in EC under certain pathological conditions.

Interaction of calcium responsive proteins and transcriptional factors with the PHO regulon in yeasts and fungi.

Phosphate and calcium ions are nutrients that play key roles in growth, differentiation and the production of bioactive secondary metabolites in filamentous fungi. Phosphate concentration regulates the biosynthesis of hundreds of fungal metabolites. The central mechanisms of phosphate transport and regulation, mediated by the master Pho4 transcriptional factor are known, but many aspects of the control of gene expression need further research. High ATP concentration in the cells leads to inositol pyrophosphate molecules formation, such as IP3 and IP7, that act as phosphorylation status reporters. Calcium ions are intracellular messengers in eukaryotic organisms and calcium homeostasis follows elaborated patterns in response to different nutritional and environmental factors, including cross-talking with phosphate concentrations. A large part of the intracellular calcium is stored in vacuoles and other organelles forming complexes with polyphosphate. The free cytosolic calcium concentration is maintained by transport from the external medium or by release from the store organelles through calcium permeable transient receptor potential (TRP) ion channels. Calcium ions, particularly the free cytosolic calcium levels, control the biosynthesis of fungal metabolites by two mechanisms, 1) direct interaction of calcium-bound calmodulin with antibiotic synthesizing enzymes, and 2) by the calmodulin-calcineurin signaling cascade. Control of very different secondary metabolites, including pathogenicity determinants, are mediated by calcium through the Crz1 factor. Several interactions between calcium homeostasis and phosphate have been demonstrated in the last decade: 1) The inositol pyrophosphate IP3 triggers the release of calcium ions from internal stores into the cytosol, 2) Expression of the high affinity phosphate transporter Pho89, a Na+/phosphate symporter, is controlled by Crz1. Also, mutants defective in the calcium permeable TRPCa7-like of Saccharomyces cerevisiae shown impaired expression of Pho89. This information suggests that CrzA and Pho89 play key roles in the interaction of phosphate and calcium regulatory pathways, 3) Finally, acidocalcisomes organelles have been found in mycorrhiza and in some melanin producing fungi that show similar characteristics as protozoa calcisomes. In these organelles there is a close interaction between orthophosphate, pyrophosphate and polyphosphate and calcium ions that are absorbed in the polyanionic polyphosphate matrix. These advances open new perspectives for the control of fungal metabolism.