Changes In Calcium Concentration Research Articles

Impact and accumulation of calcium on soft-shell mud crab Scylla paramamosain in recirculating aquaculture system

Soft-shell crabs are highly valued for their commercial significance and culinary appeal. To enhance production of soft-shell crabs, careful attention should be paid to the water calcium concentration in their culture. This study investigated the influences of different levels of calcium concentrations on the molting and growth of mud crab (Scylla paramamosain) within a meticulously controlled recirculating aquaculture system (RAS). We established three distinct water calcium concentration gradients (200, 300, and 450 mg/L) to comprehensively assess feeding habits, molting patterns, growth, and tissue calcium levels of mud crab. Initially, we observed a gradual reduction in water calcium concentrations with increments of crab molting events in RAS. Throughout one molting cycle, the feeding incidence of the crabs remained consistently high from 4 days to 37 days post-molt, irrespective of different calcium concentrations. Nonetheless, high water calcium levels (300 and 450 mg/L) significantly shortened molt intervals and accelerated exoskeletal calcification. Specifically, the molt intervals were 52.30 ± 2.72 d, 41.65 ± 5.24 d, and 37.70 ± 3.02 d, while the carapace hardening durations were 21.51 ± 4.33 h, 17.23 ± 3.38 h, and 13.46 ± 2.76 h at water calcium concentrations of 200, 300, and 450 mg/L, respectively. Moreover, elevated calcium concentrations also significantly enhanced weight gain of the mud crab during molting. Noteworthy, regardless of fluctuations in water calcium concentration, the condition factor of the molting crabs was approximately 63%, which could be an indicator of crab selection in soft-shell crab cultivation. Furthermore, the dynamic changes in calcium concentrations were disparate among different tissues in the post-molt stage. Gill and exoskeleton tissues exhibited an upward trend in calcium concentrations, while hemolymph and hepatopancreas tissues displayed distinct patterns of decrease followed by increase. Gill and exoskeleton tissues were identified as primary sites for calcium absorption and utilization, evidenced by a positive correlation between high calcium concentrations and increased calcium content in these tissues at 6 h and 3 d post-molt. Overall, our findings provide practical evidence for optimizing aquaculture management strategies through adjustments in water calcium concentration, thus to enhance production efficiency and profitability in soft-shell mud crab aquaculture.

A Mitochondrial Basis for Heart Failure Progression.

In health, the human heart is able to match ATP supply and demand perfectly. It requires 6kg of ATP per day to satisfy demands of external work (mechanical force generation) and internal work (ion movements and basal metabolism). The heart is able to link supply with demand via direct responses to ADP and AMP concentrations but calcium concentrations within myocytes play a key role, signalling both inotropy, chronotropy and matched increases in ATP production. Calcium/calmodulin-dependent protein kinase (CaMKII) is a key adapter to increased workload, facilitating a greater and more rapid calcium concentration change. In the failing heart, this is dysfunctional and ATP supply is impaired. This review aims to examine the mechanisms and pathologies that link increased energy demand to this disrupted situation. We examine the roles of calcium loading, oxidative stress, mitochondrial structural abnormalities and damage-associated molecular patterns.

Shear Stress Induces Morphology Changes in Human Lung Microvascular Endothelial Cells via Calcium Influx

Introduction: The pulmonary vascular endothelium is in contact with laminar blood flow, resulting in a physiologic degree of shear stress exerted on endothelial cells (ECs). In vitro cell culture of ECs is typically performed under static conditions, leading to underappreciation of the impact of shear stress. Calcium signaling is among the most ubiquitous cell signaling mechanisms, and changes in intracellular calcium concentration ([Ca2+]i) have been linked to shear stress. The impact of shear stress on human lung microvascular endothelial cell (hLMVEC) morphology and cell signaling is not well described. We hypothesized that hLMVECs exposed to shear stress would undergo Ca2+-dependent morphologic changes. Methods: The effect of shear stress on [Ca2+]i was first tested using ratiometric measurement. Early passage (p≤8) primary hLMVECs from three unique donor lots were plated in gelatin-coated Ibidi 0.2 uM depth plates for 16 h, then incubated with the calcium indicator Fura-2 AM for 1 h. Measurement of [Ca2+]i was obtained over 15 minutes under static and physiologic shear stress (0 and 12 dyn/cm2, respectively), using validated flow rates from the Ibidi system. Measured [Ca2+]i was compared between cells treated vehicle or an inhibitor of the mechanosensitive Ca2+-permeable channel TRPV4 (HC-067047). HC treatment was for 1 h prior to and throughout the duration of shear exposure. Morphology changes were then assessed in hLMVECs from the same three cell lots grown in standard 6-well plates; one plate kept in static culture, while another was exposed to 24 h of 12 dyn/cm2 shear stress using an orbital shaker. Treatments in each plate were control, vehicle, and HC. Representative photos of each well were taken at 0 h and at 24 h. Average ratios of shortest to longest dimensions from 30 cells/field were measured using ImageJ and compared between treatments and shear conditions. A 2-way ANOVA with post-test was used to test for statistically significant changes in morphology. Results: Shear stress increased [Ca2+]i within 15 minutes, and both baseline [Ca2+]i and observed change in response to shear stress were decreased by TRPV4 inhibition with HC. Morphologic changes were observed after exposure to shear stress on microscopic examination, with cells grown in static culture having typical cobblestone appearance and shear-adapted cells having an elongated, spindle like appearance aligned in the direction of flow. The calculated ratio (shortest to longest dimension) of all wells at 0 h was ~0.70 with no significant differences between groups. 24 h of shear stress caused a nearly 80% decrease in ratio compared to static culture in untreated cells (p<0.05), with a decrease of only 30% in HC-treated cells after 24 h of shear (p<0.05). Conclusions: Exposure to shear stress at physiological levels results in a rapid increase [Ca2+]i through mechanosensitive, Ca2+-permeable TRPV4 channels as well as notable morphologic changes to hLMVECs with prolonged exposure to shear stress that appear to be mediated in large part by TRPV4. Further investigation into the impact of shear stress on hLMVECs is warranted to elucidate biochemical pathways and other downstream functional consequences of increased [Ca2+]i. NIH-T32HL007534. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Honeybee CaV4 has distinct permeation, inactivation, and pharmacology from homologous NaV channels.

DSC1, a Drosophila channel with sequence similarity to the voltage-gated sodium channel (NaV), was identified over 20 years ago. This channel was suspected to function as a non-specific cation channel with the ability to facilitate the permeation of calcium ions (Ca2+). A honeybee channel homologous to DSC1 was recently cloned and shown to exhibit strict selectivity for Ca2+, while excluding sodium ions (Na+), thus defining a new family of Ca2+ channels, known as CaV4. In this study, we characterize CaV4, showing that it exhibits an unprecedented type of inactivation, which depends on both an IFM motif and on the permeating divalent cation, like NaV and CaV1 channels, respectively. CaV4 displays a specific pharmacology with an unusual response to the alkaloid veratrine. It also possesses an inactivation mechanism that uses the same structural domains as NaV but permeates Ca2+ ions instead. This distinctive feature may provide valuable insights into how voltage- and calcium-dependent modulation of voltage-gated Ca2+ and Na+ channels occur under conditions involving local changes in intracellular calcium concentrations. Our study underscores the unique profile of CaV4 and defines this channel as a novel class of voltage-gated Ca2+ channels.

Adenosine receptors are the on-and-off switch of astrocytic cannabinoid type 1 (CB1) receptor effect upon synaptic plasticity in the medial prefrontal cortex.

The medial prefrontal cortex (mPFC) is involved in cognitive functions such as working memory. Astrocytic cannabinoid type 1 receptor (CB1R) induces cytosolic calcium (Ca2+) concentration changes with an impact on neuronal function. mPFC astrocytes also express adenosine A1 and A2A receptors (A1R, A2AR), being unknown the crosstalk between CB1R and adenosine receptors in these cells. We show here that a further level of regulation of astrocyte Ca2+ signaling occurs through CB1R-A2AR or CB1R-A1R heteromers that ultimately impact mPFC synaptic plasticity. CB1R-mediated Ca2+ transients increased and decreased when A1R and A2AR were activated, respectively, unveiling adenosine receptors as modulators of astrocytic CB1R. CB1R activation leads to an enhancement of long-term potentiation (LTP) in the mPFC, under the control of A1R but not of A2AR. Notably, in IP3R2KO mice, that do not show astrocytic Ca2+ level elevations, CB1R activation decreases LTP, which is not modified by A1R or A2AR. The present work suggests that CB1R has a homeostatic role on mPFC LTP, under the control of A1R, probably due to physical crosstalk between these receptors in astrocytes that ultimately alters CB1R Ca2+ signaling.

Changes in nutritional and technological properties of heat-treated milk and cream at dairy production scale during storage

Changes in nutritional and technological properties during storage of milk and cream subjected to different heat treatments at dairy production scale (high-temperature-short-time pasteurisation, extended shelf-life treatment and ultra-high temperature treatment) were investigated. Results indicate a decrease in pH with longer storage times, whereas no changes in ionic calcium concentration occurred during shorter storage. Vitamin losses ranged from 1 to 22% for different heat treatments and more pronounced effects of vitamin degradation was observed with more intense heat treatment and longer storage times. Vitamin B12 concentration tended to decrease more over time than vitamins B1, B2 and E and more losses in vitamin concentration were found after storage than after heat treatment. Significant effects on physical stability and colour changes in milk and cream were observed during prolonged storage. This suggests that larger changes in nutritional and technological quality of heat-treated milk and cream at dairy production scale occur during longer storage times.

Structure of mavacamten-free human cardiac thick filaments within the sarcomere by cryoelectron tomography

Heart muscle has the unique property that it can never rest; all cardiomyocytes contract with each heartbeat which requires a complex control mechanism to regulate cardiac output to physiological requirements. Changes in calcium concentration regulate the thin filament activation. A separate but linked mechanism regulates the thick filament activation, which frees sufficient myosin heads to bind the thin filament, thereby producing the required force. Thick filaments contain additional nonmyosin proteins, myosin-binding protein C and titin, the latter being the protein that transmits applied tension to the thick filament. How these three proteins interact to control thick filament activation is poorly understood. Here, we show using 3-D image reconstruction of frozen-hydrated human cardiac muscle myofibrils lacking exogenous drugs that the thick filament is structured to provide three levels of myosin activation corresponding to the three crowns of myosin heads in each 429Å repeat. In one crown, the myosin heads are almost completely activated and disordered. In another crown, many myosin heads are inactive, ordered into a structure called the interacting heads motif. At the third crown, the myosin heads are ordered into the interacting heads motif, but the stability of that motif is affected by myosin-binding protein C. We think that this hierarchy of control explains many of the effects of length-dependent activation as well as stretch activation in cardiac muscle control.

Observation of sarcomere chaos induced by changes in calcium concentration in cardiomyocytes.

Heating cardiomyocytes to 38-42°C induces hyperthermal sarcomeric oscillations (HSOs), which combine chaotic instability and homeostatic stability. These properties are likely important for achieving periodic and rapid ventricular expansion during the diastole phase of the heartbeat. Compared with spontaneous oscillatory contractions in cardiomyocytes, which are sarcomeric oscillations induced in the presence of a constant calcium concentration, we found that calcium concentration fluctuations cause chaotic instability during HSOs. We believe that the experimental fact that sarcomeres, autonomously oscillating, exhibit such instability due to the action of calcium concentration changes is important for understanding the physiological function of sarcomeres. Therefore, we have named this chaotic sarcomere instability that appears under conditions involving changes in calcium concentration as Sarcomere Chaos with Changes in Calcium Concentration (S4C). Interestingly, sarcomere instability that could be considered S4C has also been observed in the relaxation dynamics of EC coupling. Unlike ADP-SPOCs and Cell-SPOCs under constant calcium concentration conditions, fluctuations in oscillation amplitude indistinguishable from HSOs were observed. Additionally, like HSO, a positive Lyapunov exponent was measured. S4C is likely a crucial sarcomeric property supporting the rapid and flexible ventricular diastole with each heartbeat of the heart.

Electrochemical cellular biosensor combined with fluorescence microscopy: An investigation of subtle changes in response of cells to a drug

AbstractThis study presents the combination of electrochemical cell based biosensor and fluorescence microscopy on a single platform to track biomolecular processes contributing to the morphology changes of the cells. Initial experiments demonstrate that using interdigitated electrodes where the gold microfingers were comparable in width and spacing to a single cell have the optimal sensitivity in the final electrochemical cell based‐sensing device. This was determined by measuring the cell index, based on the impedance analysis of bare and cell‐covered microelectrodes. The fabrication of the electrodes on a glass substrate enabled the capture of high‐resolution fluorescence microscopy images of single cells and related intracellular calcium release inside the HeLa cells via a window incorporated into the gold microelectrode design. As an illustration of the enhanced capability of the combined approach over traditional impedance cellular assay, the opto‐electric assay was utilized as a functional readout for G protein couple receptor activation. The simultaneous examinations of cells stimulated with histamine demonstrated an association between time courses of changes in cytosolic calcium concentration and reductions in cell‐cell adhesions.

Recording changes in biochemical parameters <i>in vivo</i> in the ischemic stroke model

Stroke is a significant and socially insidious disease that ranks second among fatal diseases according to the World Health Organization [1]. Understanding the molecular mechanisms behind the pathogenesis of this ailment will enable the development of more effective preventative measures and treatment strategies to minimize the negative consequences of stroke. Despite the abundance of experimental data, most of which were acquired indirectly, the study of the dynamics of biochemical parameters in brain tissue in real time during the acute phase of ischemic stroke is difficult. The use of genetically-encoded sensors creates novel possibilities for monitoring alterations in different biochemical and metabolic parameters in vivo tissues. In this study, we evaluated pH changes, hydrogen peroxide production (an important type of biologically active ROS), and polysulfide synthesis in various types of brain tissue cells of SHR rats during the development of ischemic stroke in real time using sensors such as SypHer3s (for pH detection), HyPer7 (for H2O2 detection), and PersIc (for polysulfide detection). Middle cerebral artery occlusion was used to simulate an ischemic stroke. The in vivo sensor signals were registered with a fiber optic setup that was created in the laboratory of spectroscopy and nonlinear optics at Moscow State University. The studies revealed that in the acute phase of stroke, acidosis occurred in the cytoplasm of neurons in the caudate nucleus, the epicenter of ischemia. The pH mutated from 7.25±0.08 to 6.7±0.15 within the first few seconds after arterial occlusion initiation. A gradual increase in pH was observed after the initial drop, which persisted throughout reperfusion but did not return to the original value in all animals. In the penumbra zone, a wave-like shift in sensor signal was detected, whereas no change in sensor signal was noted in the healthy hemisphere. Investigation of the dynamics of H2O2 formation in the mitochondrial matrix of caudate neurons revealed minimal sensor oxidation during ischemia/reperfusion in the acute phase of stroke, indicating low ROS production. Nevertheless, a substantial increase in the sensor signal was detected after 24 hours following the surgery. Thus, the confirmation of oxidative stress development in the affected hemisphere differed from the commonly accepted view in terms of its dynamics. Previously, it was believed that excessive production of H2O2 leading to oxidative stress and related brain cell death occurred primarily in the acute phase. However, a comparison of hydrogen peroxide production dynamics in neurons and astrocytes revealed differences between these cell populations. It was discovered that as early as 12 hours after middle cerebral artery occlusion, the sensor signal in astrocytes increased more intensely than in neurons. This trend persisted until the end of the measurements, 40 hours after surgery. The observed distinctions may stem from glial cells’ protective function in counteracting the harmful consequences of hydrogen peroxide on neurons, along with their contribution to maintaining the myelin structure in the brain. Additionally, the role of astrocytes in neuroinflammation development is noteworthy. Reactive sulfur species, in addition to reactive oxygen species, appear to be significant contributors to the development of pathological processes. The PersIc sensor signal measurement did not show any disparities between the caudate nucleus of the healthy hemisphere and the hemisphere affected by stroke development in terms of polysulfide and persulfide appearance detection. However, the area surrounding the core infarction is noteworthy due to the observed bouts of acidosis using the SypHer3s sensor. Our findings suggest a potential association between these bouts, spreading depolarization, changes in calcium concentration, and the development of neuroinflammation. These reactions may ultimately lead to the synthesis of polysulfides, known modulators of inflammatory reactions. Thus, our data provides valuable additions to the existing knowledge on metabolic changes that take place during the progression of ischemic brain injury.

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.

Phosphorus balance calculator: an individualized tool for treatment of hyperphosphatemia in hemodialysis patients.

Lack of evaluations of the dietary phosphorus and dialysis phosphorus removal in daily clinical practice are the common obstacle to assess phosphorus balance and control phosphorus in hemodialysis patients. We aimed to investigate whether the individualized therapy using phosphorus balance calculator improves phosphorus control. A randomized, open-label, multicenter, 4-week clinical trial was conducted. 119 maintenance hemodialysis patients aged 18 to 85 years old and with serum phosphorus level higher than 1.45mmol/l from 3 university teaching hospitals in Shanghai were enrolled. Patients were randomized in a 1:1 ratio to individualized therapy (n=60), or conventional therapy (n=59). The primary outcome was the serum phosphorus concentration after 4-week treatment. Secondary outcomes included the serum calcium and parathyroid hormone (PTH) concentrations, changes in serum phosphorus, calcium and PTH concentrations, and the proportion of patients achieving target ranges of serum phosphorus, calcium and PTH after 4-week treatment. Among 119 randomized participants (mean age, 62 years; 68 male[57%]), 116 completed the trial. By using the phosphorus balance calculator, the individualized group achieved a better phosphorus balance state, significantly reduced the serum phosphorus (1.62±0.45mmol/l versus 1.85±0.45 mmol/l, P=0.006), increased the proportions of patients achieving target serum phosphorus range (41% versus 18%, P=0.006), and had greater adjusted mean difference in change in serum phosphorus over the 4 weeks (-0.47 versus -0.23mmol/l, P=0.010) when compared to conventional therapy. No significant changes were observed in serum calcium and PTH levels, the proportion of patients achieving target serum calcium or PTH levels, and adjusted mean difference of serum calcium and PTH levels over the treatment period. Phosphorus balance calculator was proved to improve serum phosphorus control in patients undergoing maintenance hemodialysis, offering a new tool for managing hyperphosphatemia.

Amyloid plaques and normal ageing have differential effects on microglial Ca2+ activity in the mouse brain

In microglia, changes in intracellular calcium concentration ([Ca2+]i) may regulate process motility, inflammasome activation, and phagocytosis. However, while neurons and astrocytes exhibit frequent spontaneous Ca2+ activity, microglial Ca2+ signals are much rarer and poorly understood. Here, we studied [Ca2+]i changes of microglia in acute brain slices using Fluo-4–loaded cells and mice expressing GCaMP5g in microglia. Spontaneous Ca2+ transients occurred ~ 5 times more frequently in individual microglial processes than in their somata. We assessed whether microglial Ca2+ responses change in Alzheimer's disease (AD) using AppNL−G−F knock-in mice. Proximity to Aβ plaques strongly affected microglial Ca2+ activity. Although spontaneous Ca2+ transients were unaffected in microglial processes, they were fivefold more frequent in microglial somata near Aβ plaques than in wild-type microglia. Microglia away from Aβ plaques in AD mice showed intermediate properties for morphology and Ca2+ responses, partly resembling those of wild-type microglia. By contrast, somatic Ca2+ responses evoked by tissue damage were less intense in microglia near Aβ plaques than in wild-type microglia, suggesting different mechanisms underlying spontaneous vs. damage-evoked Ca2+ signals. Finally, as similar processes occur in neurodegeneration and old age, we studied whether ageing affected microglial [Ca2+]i. Somatic damage-evoked Ca2+ responses were greatly reduced in microglia from old mice, as in the AD mice. In contrast to AD, however, old age did not alter the occurrence of spontaneous Ca2+ signals in microglial somata but reduced the rate of events in processes. Thus, we demonstrate distinct compartmentalised Ca2+ activity in microglia from healthy, aged and AD-like brains.

Involvement of the Calmodulin-like Protein Gene VaCML92 in Grapevine Abiotic Stress Response and Stilbene Production.

Calmodulin-like proteins (CMLs) are an important family of plant calcium sensor proteins that sense and decode changes in the intracellular calcium concentration in response to environmental and developmental stimuli. Nonetheless, the specific functions of individual CML family members remain largely unknown. This study aims to explore the role of the Vitis amurensis VaCML92 gene in the development of its high stress resistance and the production of stilbenes. The expression of VaCML92 was sharply induced in V. amurensis cuttings after cold stress. The VaCML92 gene was cloned and its role in the abiotic stress responses and stilbene production in grapevine was further investigated. The VaCML92-overexpressing callus cell cultures of V. amurensis and soil-grown plants of Arabidopsis thaliana exhibited enhanced tolerance to cold stress and, to a lesser extent, to the drought, while their tolerance to heat stress and high salinity was not affected. In addition, the overexpression of VaCML92 increased stilbene production in the V. amurensis cell cultures by 7.8-8.7-fold. Taken together, the data indicate that the VaCML92 gene is involved as a strong positive regulator in the rapid response to cold stress, the induction of cold stress resistance and in stilbene production in wild grapevine.

Effects of Remimazolam on Intracellular Calcium Dynamics in Myotubes Derived from Patients with Malignant Hyperthermia and Functional Analysis of Type 1 Ryanodine Receptor Gene Variants.

Remimazolam is a novel general anesthetic and its safety in patients with malignant hyperthermia (MH) is unknown. We used myotubes derived from the skeletal muscle of patients with MH to examine the response to ryanodine receptor 1 (RYR1) agonist and remimazolam in MH-susceptible patients. Patients underwent muscle biopsy for the Ca2+-induced Ca2+ release (CICR) rate test, a diagnostic tool for MH in Japan. Ten patients had myotubes obtained from skeletal muscle cultures, and the genes associated with malignant hyperthermia in these patients were analyzed. The EC50 of caffeine, cresol, and remimazolam to induce intracellular calcium concentration change were compared between myotubes from CICR-negative genetic test patients and myotubes from other patients. Eight of the ten were CICR-positive, five of whom had RYR1 causative gene mutations or variants. Two patients had CICR-negative genetic tests, and as expected had the highest EC50 (the concentration of a drug that gives a half-maximal response) in response to caffeine, 4CmC and remimazolam. Three patients had a positive CICR but no known variants in RYR1 or CACNA1S (voltage-gated calcium channel subunit alpha1S). Myotubes in these patients had significantly lower EC50s for all agents than myotubes in CICR-negative patients. When myotubes from a patient who was CICR-negative and had no gene variant were used as a control, myotubes from CICR-positive patients were more hyper-responsive than controls to all stimulants used. The EC50 for remimazolam was lowest for myotubes from CICR-positive, RYR1-mutant patients, at 206 µM (corresponding to 123 µg/mL). The concentration was more than 80-times higher than the clinical concentration. RYR1 gene variants in R4645Q and W5020G were shown to be causative gene mutations for MH. Intracellular calcium in myotubes from MH patients are elevated at high concentrations of remimazolam but not at clinically used concentrations of remimazolam. Remimazolam appears to be safe to use in patients with MH.

Neurovirulent cytokines increase neuronal excitability in a model of coronavirus-induced neuroinflammation

BackgroundNeurological manifestations of severe coronavirus infections, including SARS-CoV-2, are wide-ranging and may persist following virus clearance. Detailed understanding of the underlying changes in brain function may facilitate the identification of therapeutic targets. We directly tested how neocortical function is impacted by the specific panel of cytokines that occur in coronavirus brain infection. Using the whole-cell patch-clamp technique, we determined how the five cytokines (TNFα, IL-1β, IL-6, IL-12p40 and IL-15 for 22–28-h) at concentrations matched to those elicited by MHV-A59 coronavirus brain infection, affected neuronal function in cultured primary mouse neocortical neurons.ResultsWe evaluated how acute cytokine exposure affected neuronal excitability (propensity to fire action potentials), membrane properties, and action potential characteristics, as well as sensitivity to changes in extracellular calcium and magnesium (divalent) concentration. Neurovirulent cytokines increased spontaneous excitability and response to low divalent concentration by depolarizing the resting membrane potential and hyperpolarizing the action potential threshold. Evoked excitability was also enhanced by neurovirulent cytokines at physiological divalent concentrations. At low divalent concentrations, the change in evoked excitability was attenuated. One hour after cytokine removal, spontaneous excitability and hyperpolarization of the action potential threshold normalized but membrane depolarization and attenuated divalent-dependent excitability persisted.ConclusionsCoronavirus-associated cytokine exposure increases spontaneous excitability in neocortical neurons, and some of the changes persist after cytokine removal.

Efficacy and Safety of Ferric Citrate on Hyperphosphatemia among Chinese Patients with Chronic Kidney Disease Undergoing Hemodialysis: A Phase III Multicenter Randomized Open-Label Active-Drug-Controlled Study

Introduction: Hyperphosphatemia in chronic kidney disease (CKD) patients is positively associated with mortality. Ferric citrate is a potent phosphorus binder that lowers serum phosphorus level and improves iron metabolism. We compared its efficacy and safety with active drugs in Chinese CKD patients with hemodialysis. Methods: Chinese patients undergoing hemodialysis were randomized into two treatment groups in a 1:1 ratio, receiving either ferric citrate or sevelamer carbonate, respectively, for 12 weeks. Serum phosphorus levels, calcium concentration, and iron metabolism parameters were evaluated every 2 weeks. Frequency and severity of adverse events were recorded. Results: 217 (90.4%) patients completed the study with balanced demographic and baseline characteristics between two groups. Ferric citrate decreased the serum phosphorus level to 0.59 ± 0.54 mmol/L, comparable to 0.56 ± 0.62 mmol/L by sevelamer carbonate. There was no significant difference between two groups (p > 0.05) in the proportion of patients with serum phosphorus levels reaching the target range, the response rate to the study drug, and the changes of corrected serum calcium concentrations, and intact-PTH levels at the end of treatment. The change of iron metabolism indicators in the ferric citrate group was significantly higher than those in the sevelamer carbonate group. There are 47 (40.5%) patients in the ferric citrate group, and 26 (21.3%) patients in the sevelamer carbonate group experienced drug-related treatment emergent adverse events (TEAEs); most were mild and tolerable. Common drug-related TEAEs were gastrointestinal disorders, including diarrhea (12.9 vs. 2.5%), fecal discoloration (14.7 vs. 0%), and constipation (1.7 vs. 7.4%) in ferric citrate and sevelamer carbonate group. Conclusion: Ferric citrate capsules have good efficacy and safety in the control of hyperphosphatemia in adult patients with CKD undergoing hemodialysis. Efficacy is not inferior to sevelamer carbonate. The TEAEs were mostly mild and tolerated by the patients.

Method for Studying Calcium Homeostasis at the Level of Gastroenteral Environment in Minipigs

The stages of calcium metabolism at the enteric medium level were studied in experiments on minipigs. Atomic absorption spectrophotometry was used to determine the concentration of calcium in the endogenous and exogenous structures of the chyme while its passing through the small and large intestines, both in different layers of the intestinal wall and in the blood flowing in and out of the intestine. Regularities of changes in calcium concentration in chyme fractions and its predominant accumulation in the dense endogenous fraction in the small intestine were established. Taking into account the hydration of the glycoproteins of the abdominal mucus and its existence in the form of a kind of enteroplasm of the chyme, it is assumed that calcium is involved in the formation of endogenous structures and chyme as a whole by redistributing between soluble and fixed forms of the element. The dynamics of the increase in calcium concentration in the layer of mucous deposits from the distal to the proximal section of the intestinal wall was revealed. In the intestinal mucosa, the calcium concentration is generally lower, although the dynamics follows a similar pattern. The arterio-venous difference in the damage of the digestive tract indicates the predominant excretion of the element into the digestive cavity. The conclusion is made about the role of endogenous chyme structures in maintaining calcium homeostasis not only in the gastroenteric environment, but also in the body as a whole.

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.