Activation Of Ryanodine Receptors Research Articles

Juxtaglomerular cell CaSR stimulation decreases renin release via activation of the PLC/IP3pathway and the ryanodine receptor

The calcium-sensing receptor (CaSR) is a G-coupled protein expressed in renal juxtaglomerular (JG) cells. Its activation stimulates calcium-mediated decreases in cAMP content and inhibits renin release. The postreceptor pathway for the CaSR in JG cells is unknown. In parathyroids, CaSR acts through G(q) and/or G(i). Activation of G(q) stimulates phospholipase C (PLC), and inositol 1,4,5-trisphosphate (IP(3)), releasing calcium from intracellular stores. G(i) stimulation inhibits cAMP formation. In afferent arterioles, the ryanodine receptor (RyR) enhances release of stored calcium. We hypothesized JG cell CaSR activation inhibits renin via the PLC/IP(3) and also RyR activation, increasing intracellular calcium, suppressing cAMP formation, and inhibiting renin release. Renin release from primary cultures of isolated mouse JG cells (n = 10) was measured. The CaSR agonist cinacalcet decreased renin release 56 ± 7% of control (P < 0.001), while the PLC inhibitor U73122 reversed cinacalcet inhibition of renin (104 ± 11% of control). The IP(3) inhibitor 2-APB also reversed inhibition of renin from 56 ± 6 to 104 ± 11% of control (P < 0.001). JG cells were positively labeled for RyR, and blocking RyR reversed CaSR-mediated inhibition of renin from 61 ± 8 to 118 ± 22% of control (P < 0.01). Combining inhibition of IP(3) and RyR was not additive. G(i) inhibition with pertussis toxin plus cinacalcet did not reverse renin inhibition (65 ± 12 to 41 ± 8% of control, P < 0.001). We conclude stimulating JG cell CaSR activates G(q), initiating the PLC/IP(3) pathway, activating RyR, increasing intracellular calcium, and resulting in calcium-mediated renin inhibition.

Synthesis and insecticidal evaluation of novel N-pyridylpyrazolecarboxamides containing cyano substituent in the ortho-position

In an attempt to search for potent insecticides targeting the ryanodine receptor (RyR), a series of novel N-pyridylpyrazolecarboxamides containing cyano substituent in the ortho-position were designed and synthesized. Their insecticidal activities of target compounds against oriental armyworm (Mythimna separata) and diamondback moth (Plutella xylostella) indicated that most of the compounds showed moderate to high activities at the tested concentrations. In particular, compound 6l and 6o showed 86% larvicidal activities against Plutella xylostella at the concentration of 0.1mg/L, while the activity of compound 6h against Mythimna separate was 80% at 1mg/L. The calcium imaging technique was applied to investigate the effects of some title compounds on the intracellular calcium ion concentration ([Ca2+]i), experimental results demonstrated that compound 6h stimulates a transient elevation in [Ca2+]i in the absence of external calcium after the central neurons dye loading with fluo-3 AM. However, when the central neurons were dyed with fluo-5N and incubated with 2-APB, [Ca2+]i decreased transiently by treated of compound 6h. All of the calcium imaging technique experiments demonstrated that these novel compounds deliver calcium from endoplasmic reticulum to cytoplasm, which proved that the title compounds were the possible activators of insect RyR.

Ryanodine receptors are uncoupled from contraction in rat vena cava

Ryanodine receptors (RyR) are Ca2+-sensitive ion channels in the sarcoplasmic reticulum (SR) membrane, and are important effectors of SR Ca2+ release and smooth muscle excitation–contraction coupling. While the relationship between RyR activation and contraction is well characterized in arteries, little is known about the role of RyR in excitation–contraction coupling in veins. We hypothesized that RyR are present and directly coupled to contraction in rat aorta (RA) and vena cava (RVC). RA and RVC expressed mRNA for all 3 RyR subtypes, and immunofluorescence showed RyR protein was present in RA and RVC smooth muscle cells. RA and RVC rings contracted when Ca2+ was re-introduced after stores depletion with thapsigargin (1μM), indicating both tissues contained intracellular Ca2+ stores. To assess RyR function, contraction was then measured in RA and RVC exposed to the RyR activator caffeine (20mM). In RA, caffeine caused contraction that was attenuated by the RyR antagonists ryanodine (10μM) and tetracaine (100μM). However, caffeine (20mM) did not contract RVC. We next measured contraction and intracellular Ca2+ (Ca2+i) simultaneously in RA and RVC exposed to caffeine. While caffeine increased Ca2+i and contracted RA, it had no significant effect on Ca2+i or contraction in RVC. These data suggest that ryanodine receptors, while present in both RA and RVC, are inactive and uncoupled from Ca2+ release and contraction in RVC.

Crystal structure and activating effect on RyRs of AhV_TL-I, a glycosylated thrombin-like enzyme from Agkistrodon halys snake venom

A snake venom thrombin-like enzyme (SVTLE) from Agkistrodon halys pallas venom was isolated by means of a two-step chromatographic procedure. The purified enzyme, named AhV_TL-I, showed fibrinogenolytic activity against both the Aα and Bβ chains of bovine fibrinogen. Unlike the other SVTLEs, AhV_TL-I has poor esterolytic activity upon BAEE substrate. The N-terminal sequence of AhV_TL-I was determined to be IIGGDEXNINEHRFLVALYT, and the molecular mass was confirmed to 29389.533Da by MALDI-TOF mass spectrometry. Its complete cDNA and derived amino acid sequence were obtained by RT-PCR. The crystal structure of AhV_TL-I was determined at a resolution of 1.75 Å. A disaccharide was clearly mapped in the structure, which involved in regulating the esterolytic activity of AhV_TL-I. The presence of the N-glycan deformed the 99-loop, and the resulting steric hindrances hindered the substrates to access the active site. Furthermore, with the carbohydrate moiety, AhV_TL-I could induce mouse thoracic aortic ring contraction with the EC(50) of 147nmol/L. Besides, the vasoconstrictor effects of AhV_TL-I were also independent of the enzymatic activity. The results of [Ca(2+)](i) measurement showed that the vasoconstrictor effects of AhV_TL-I were attributed to Ca(2+) releasing from Ca(2+) store. Further studies showed that it was related to the activation of ryanodine receptors (RyRs). These offer new insights into the snake SVTLEs functions and provide a novel pathogenesis of A. halys pallas venom.

Making Synapses Strong: Metaplasticity Prolongs Associativity of Long-Term Memory by Switching Synaptic Tag Mechanisms

One conceptual mechanism for the induction of associative long-term memory is that a synaptic tag, set by a weak event, can capture plasticity-related proteins from a nearby strong input, thus enabling associativity between the 2 (synaptic tagging and capture, STC). So far, STC has been observed for only a limited time of 60 min. Nevertheless, association of weak memory forms can occur beyond this period and its mechanism is not well understood. Here we report that metaplasticity induced by ryanodine receptor activation or synaptic activation of metabotropic glutamate receptors prolongs the durability of the synaptic tag, thus extending the time window for associative interactions mediating storage of long-term memory. We provide evidence that such metaplasticity alters the mechanisms of STC from a CaMKII-mediated (in non-primed STC) to a protein kinase Mzeta (PKMζ)-mediated process (in primed STC). Thus the association of weak synapses with strong synapses in the "late" stage of associative memory formation occurs only through metaplasticity. The results also reveal that the short-lived, CaMKII-mediated tag may contribute to a mechanism for a fragile form of memory while metaplasticity enables a PKMζ-mediated synaptic tag capable of prolonged interactions that induce a more stable form of memory that is resistant to reversal.

Maternal high-altitude hypoxia and suppression of ryanodine receptor-mediated Ca2+ sparks in fetal sheep pulmonary arterial myocytes

Ca(2+) sparks are fundamental Ca(2+) signaling events arising from ryanodine receptor (RyR) activation, events that relate to contractile and dilatory events in the pulmonary vasculature. Recent studies demonstrate that long-term hypoxia (LTH) can affect pulmonary arterial reactivity in fetal, newborn, and adult animals. Because RyRs are important to pulmonary vascular reactivity and reactivity changes with ontogeny and LTH we tested the hypothesis that RyR-generated Ca(2+) signals are more active before birth and that LTH suppresses these responses. We examined these hypotheses by performing confocal imaging of myocytes in living arteries and by performing wire myography studies. Pulmonary arteries (PA) were isolated from fetal, newborn, or adult sheep that lived at low altitude or from those that were acclimatized to 3,801 m for > 100 days. Confocal imaging demonstrated preservation of the distance between the sarcoplasmic reticulum, nucleus, and plasma membrane in PA myocytes. Maturation increased global Ca(2+) waves and Ca(2+) spark activity, with sparks becoming larger, wider, and slower. LTH preferentially depressed Ca(2+) spark activity in immature pulmonary arterial myocytes, and these sparks were smaller, wider, and slower. LTH also suppressed caffeine-elicited contraction in fetal PA but augmented contraction in the newborn and adult. The influence of both ontogeny and LTH on RyR-dependent cell excitability shed new light on the therapeutic potential of these channels for the treatment of pulmonary vascular disease in newborns as well as adults.

Luminal Ca2+ controls activation of the cardiac ryanodine receptor by ATP

The synergic effect of luminal Ca2+, cytosolic Ca2+, and cytosolic adenosine triphosphate (ATP) on activation of cardiac ryanodine receptor (RYR2) channels was examined in planar lipid bilayers. The dose–response of RYR2 gating activity to ATP was characterized at a diastolic cytosolic Ca2+ concentration of 100 nM over a range of luminal Ca2+ concentrations and, vice versa, at a diastolic luminal Ca2+ concentration of 1 mM over a range of cytosolic Ca2+ concentrations. Low level of luminal Ca2+ (1 mM) significantly increased the affinity of the RYR2 channel for ATP but without substantial activation of the channel. Higher levels of luminal Ca2+ (8–53 mM) markedly amplified the effects of ATP on the RYR2 activity by selectively increasing the maximal RYR2 activation by ATP, without affecting the affinity of the channel to ATP. Near-diastolic cytosolic Ca2+ levels (<500 nM) greatly amplified the effects of luminal Ca2+. Fractional inhibition by cytosolic Mg2+ was not affected by luminal Ca2+. In models, the effects of luminal and cytosolic Ca2+ could be explained by modulation of the allosteric effect of ATP on the RYR2 channel. Our results suggest that luminal Ca2+ ions potentiate the RYR2 gating activity in the presence of ATP predominantly by binding to a luminal site with an apparent affinity in the millimolar range, over which local luminal Ca2+ likely varies in cardiac myocytes.

Synthesis and Insecticidal Activities of Novel Anthranilic Diamides Containing Acylthiourea and Acylurea

Two series of anthranilic diamides containing acylthiourea and acylurea linkers were designed and synthesized, with changed length and flexibility of the linkers to compare to known anthranilic diamide insecticides. In total, 26 novel compounds were synthesized, and all compounds were characterized by (1)H nuclear magnetic resonance and high-resolution mass spectrometry. Their insecticidal activities against oriental armyworm (Mythimna separata), mosquito larvae (Culex pipiens pallens), and diamondback moth (Plutella xylostella) were evaluated. The larvicidal activities against oriental armyworm indicated that the introduction of acylthiourea into some structures could retain their insecticidal activity; 8 of the 15 compounds (13a-13e, 14a-14e, and 15a-15e) exhibited 100% larvicidal activity at 10 mg/L. However, the introduction of acylurea decreased the insecticidal activity; only 3 of the 11 compounds (17a-17k) exhibited 100% larvicidal activity at 200 mg/L. The whole-cell patch-clamp technique indicated that compound 13b and chlorantraniliprole exhibited similar effects on the voltage-gated calcium channel. The calcium-imaging technique was also applied to investigate the effects of compounds 13b and 15a on the intracellular calcium ion concentration ([Ca(2+)](i)), which indicated that they released stored calcium ions from endoplasmic reticulum. Experimental results denoted that several new compounds are potential activators of the insect ryanodine receptor (RyR).

FKBP12.6 overexpression does not protect against remodelling after myocardial infarction

Reducing the open probability of the ryanodine receptor (RyR) has been proposed to have beneficial effects in heart failure. We investigated whether conditional FKBP12.6 overexpression at the time of myocardial infarction (MI) could improve cardiac remodelling and cell Ca(2+) handling. Wild-type (WT) mice and mice overexpressing FKBP12.6 (Tg) were studied on average 7.5 ± 0.2 weeks after MI and compared with sham-operated mice for in vivo, myocyte function and remodelling. At baseline, unloaded cell shortening in Tg was not different from WT. The [Ca(2+)](i) transient amplitude was similar, but sarcoplasmic reticulum (SR) Ca(2+) content was larger in Tg, suggesting reduced fractional release. Spontaneous spark frequency was similar despite the increased SR Ca(2+) content, consistent with a reduced RyR channel open probability in Tg. After MI, left ventricular dilatation and myocyte hypertrophy were present in both groups, but more pronounced in Tg. Cell shortening amplitude was unchanged with MI in WT, but increased with MI in Tg. The amplitude of the [Ca(2+)](i) transient was not affected by MI in either genotype, but time to peak was increased; this was most pronounced in Tg. The SR Ca(2+) content and Na(+)- Ca(2+) exchanger function were not affected by MI. Spontaneous spark frequency was increased significantly after MI in Tg, and larger than in WT (at 4 Hz, 2.6 ± 0.4 sparks (100 μm)(-1) s(-1) in Tg MI versus 1.6 ± 0.2 sparks (100 μm)(-1) s(-1) in WT MI; P < 0.05). We conclude that FKPB12.6 overexpression can effectively reduce RyR open probability with maintained cardiomyocyte contraction. However, this approach appears insufficient to prevent and reduce post-MI remodelling, indicating that additional pathways may need to be targeted.

Early Presynaptic and Postsynaptic Calcium Signaling Abnormalities Mask Underlying Synaptic Depression in Presymptomatic Alzheimer's Disease Mice

Alzheimer's disease (AD)-linked presenilin (PS) mutations result in pronounced endoplasmic reticulum calcium disruptions that occur before detectable histopathology and cognitive deficits. More subtly, these early AD-linked calcium alterations also reset neurophysiological homeostasis, such that calcium-dependent presynaptic and postsynaptic signaling appear functionally normal yet are actually operating under aberrant calcium signaling systems. In these 3xTg-AD mouse brains, upregulated ryanodine receptor (RyR) activity is associated with a shift toward synaptic depression, likely through a reduction in presynaptic vesicle stores and increased postsynaptic outward currents through small-conductance calcium-activated potassium SK2 channels. The deviant RyR-calcium involvement in the 3xTg-AD mice also compensates for an intrinsic predisposition for hippocampal long-term depression (LTD) and reduced long-term potentiation (LTP). In this study, we detail the impact of disrupted RyR-mediated calcium stores on synaptic transmission properties, LTD, and calcium-activated membrane channels of hippocampal CA1 pyramidal neurons in presymptomatic 3xTg-AD mice. Using electrophysiological recordings in young 3xTg-AD and nontransgenic (NonTg) hippocampal slices, we show that increased RyR-evoked calcium release in 3xTg-AD mice "normalizes" an altered synaptic transmission system operating under a shifted homeostatic state that is not present in NonTg mice. In the process, we uncover compensatory signaling mechanisms recruited early in the disease process that counterbalance the disrupted RyR-calcium dynamics, namely increases in presynaptic spontaneous vesicle release, altered probability of vesicle release, and upregulated postsynaptic SK channel activity. Because AD is increasingly recognized as a "synaptic disease," calcium-mediated signaling alterations may serve as a proximal trigger for the synaptic degradation driving the cognitive loss in AD.

How well do muscle biomechanics predict whole-animal locomotor performance? The role of Ca2+ handling

It is important to determine the enabling mechanisms that underlie locomotor performance to explain the evolutionary patterns and ecological success of animals. Our aim was to determine the extent to which calcium (Ca(2+)) handling dynamics modulate the contractile properties of isolated skeletal muscle, and whether the effects of changing Ca(2+) handling dynamics in skeletal muscle are paralleled by changes in whole-animal sprint and sustained swimming performance. Carp (Cyprinus carpio) increased swimming speed by concomitant increases in tail-beat amplitude and frequency. Reducing Ca(2+) release from the sarcoplasmic reticulum (SR) by blocking ryanodine receptors with dantrolene decreased isolated peak muscle force and was paralleled by a decrease in tail-beat frequency and whole-animal sprint performance. An increase in fatigue resistance following dantrolene treatment may reflect the reduced depletion of Ca(2+) stores in the SR associated with lower ryanodine receptor (RyR) activity. Blocking RyRs may be detrimental by reducing force production and beneficial by reducing SR Ca(2+) depletion so that there was no net effect on critical sustained swimming speed (U(crit)). In isolated muscle, there was no negative effect on force production of blocking Ca(2+) release via dihydropyridine receptors (DHPRs) with nifedipine. Nifedipine decreased fatigue resistance of isolated muscle, which was paralleled by decreases in tail-beat frequency and U(crit). However, sprint performance also decreased with DHPR inhibition, which may indicate a role in muscle contraction of the Ca(2+) released by DHPR into the myocyte. Inhibiting sarco(endo)plasmic reticulum Ca(2+)-ATPase (SERCA) activity with thapsigargin decreased fatigue resistance, suggesting that SERCA activity is important in avoiding Ca(2+) store depletion and fatigue. We have shown that different molecular mechanisms modulate the same muscle and whole-animal traits, which provides an explanatory model for the observed variations in locomotor performance within and between species.

Synthesis and Insecticidal Activities of Novel Phthalic Acid Diamides

AbstractIn order to discover novel insecticides with the new action mode on ryanodine receptor (RyR), a series of novel phthalic acid diamide derivatives were designed and synthesized. All compounds were characterized by 1H NMR spectra and HRMS. The preliminary results of biological activity assessment indicated that some title compounds exhibited excellent insecticidal activities against Mythimna separata, Spodoptera exigua, and Plutella xylostella. The title compound 3‐nitro‐N‐cyclopropyl‐N′‐[2‐methyl‐4‐(perfluoropropan‐2‐yl)phenyl]phthalamide (4a) was more efficient against diamondback moths than the control (chlorantraniliprole). The effects of some title compounds on intracellular calcium of neurons from the Spodoptera exigua proved that the title compounds were RyR activators.

Semaphorin3A facilitates axonal transport through a local calcium signaling and tetrodotoxin-sensitive voltage-gated sodium channels

Semaphorin3A (Sema3A), a secreted factor that navigates axons and dendrites of developing neurons, facilitates axonal transport. However, little is known about the mechanism underlying Sema3A-induced facilitation and its functional implications. Here we show that Sema3A induces facilitation of axonal transport via local calcium signaling in growth cone. The facilitation of axonal transport was blocked by inhibitors of voltage-gated sodium channels (tetrodotoxin, TTX), L-type voltage-gated calcium channel, and ryanodine receptor (RyR). Sema3A evoked intracellular Ca2+ elevation in growth cone by local application of Sema3A to growth cone. Sema3A also activated RyR in growth cone as well as cell body. Notably, TTX suppressed Sema3A-induced RyR activation in cell body but not in growth cone. Our results identify a novel mechanism of Sema3A-induced axonal transport, and further suggest that Sema3A-induced local calcium signaling in growth cone is propagated to cell body in a TTX-sensitive manner.

PCB-95 Promotes Dendritic Growth via Ryanodine Receptor–Dependent Mechanisms

Background: Aroclor 1254 (A1254) interferes with normal dendritic growth and plasticity in the developing rodent brain, but the mechanism(s) mediating this effect have yet to be established. Non-dioxin-like (NDL) polychlorinated biphenyls (PCBs) enhance the activity of ryanodine receptor (RyR) calcium ion (Ca2+) channels, which play a central role in regulating the spatiotemporal dynamics of intracellular Ca2+ signaling. Ca2+ signaling is a predominant factor in shaping dendritic arbors, but whether PCB potentiation of RyR activity influences dendritic growth is not known.Objective: We determined whether RyR activity is required for PCB effects on dendritic growth.Methods and Results: Golgi analysis of hippocampi from weanling rats confirmed that developmental exposure via the maternal diet to NDL PCB-95 (2,2´,3,5´6-pentachlorobiphenyl), a potent RyR potentiator, phenocopies the dendrite-promoting effects of A1254. Dendritic growth in dissociated cultures of primary hippocampal neurons and in hippocampal slice cultures is similarly enhanced by PCB-95 but not by PCB-66 (2,3,4´,4-tetrachlorobiphenyl), a congener with negligible effects on RyR activity. The dendrite-promoting effects of PCB-95 are evident at concentrations as low as 2 pM and are inhibited by either pharmacologic blockade or siRNA knockdown of RyRs.Conclusions: Our findings demonstrate that environmentally relevant levels of NDL PCBs modulate neuronal connectivity via RyR-dependent effects on dendritic arborization. In addition, these findings identify RyR channel dysregulation as a novel mechanism contributing to dysmorphic dendritogenesis associated with heritable and environmentally triggered neurodevelopmental disorders.

PCB-95 Modulates the Calcium-Dependent Signaling Pathway Responsible for Activity-Dependent Dendritic Growth

Background: Non-dioxin-like (NDL) polychlorinated biphenyls (PCBs) promote dendritic growth in hippocampal neurons via ryanodine receptor (RyR)-dependent mechanisms; however, downstream signaling events that link enhanced RyR activity to dendritic growth are unknown. Activity-dependent dendritic growth, which is a critical determinant of neuronal connectivity in the developing brain, is mediated by calcium ion (Ca2+)-dependent activation of Ca2+/calmodulin kinase-I (CaMKI), which triggers cAMP response element binding protein (CREB)-dependent Wnt2 transcription. RyRs regulate the spatiotemporal dynamics of intracellular Ca2+ signals, but whether RyRs promote dendritic growth via modulation of this signaling pathway is not known.Objective: We tested the hypothesis that the CaMKI–CREB–Wnt2 signaling pathway couples NDL PCB-enhanced RyR activity to dendritic arborization.Methods and Results: Ca2+ imaging of dissociated cultures of primary rat hippocampal neurons indicated that PCB-95 (2,2´,3,5´6-pentachlorobiphenyl; a potent RyR potentiator), enhanced synchronized Ca2+ oscillations in somata and dendrites that were blocked by ryanodine. As determined by Western blotting and quantitative polymerase chain reaction, PCB-95 also activated CREB and up-regulated Wnt2. Blocking CaMKK, CaMKIα/γ, MEK/ERK, CREB, or Wnt2 prevented PCB-95–induced dendritic growth. Antagonism of γ-aminobutyric acid (GABA) receptors with bicuculline (BIC) phenocopied the dendrite-promoting effects of PCB-95, and pharmacological antagonism or siRNA knockdown of RyR blocked BIC-induced dendritic growth in dissociated and slice cultures of hippocampal neurons.Conclusions: RyR activity contributes to dynamic remodeling of dendritic architecture in response to NDL PCBs via CaMKI–CREB–Wnt2 signaling in rats. Our findings identify PCBs as candidate environmental risk factors for neurodevelopmental disorders, especially in children with heritable deficits in calcium signaling associated with autism.

Selenoprotein N in skeletal muscle: from diseases to function

Selenoprotein N (SelN) deficiency causes several inherited neuromuscular disorders collectively termed SEPN1-related myopathies, characterized by early onset, generalized muscle atrophy, and muscle weakness affecting especially axial muscles and leading to spine rigidity, severe scoliosis, and respiratory insufficiency. SelN is ubiquitously expressed and is located in the membrane of the endoplasmic reticulum; however, its function remains elusive. The predominant expression of SelN in human fetal tissues and the embryonic muscle phenotype reported in mutant zebrafish suggest that it is involved in myogenesis. In mice, SelN is also mostly expressed during embryogenesis and especially in the myotome, but no defect was detected in muscle development and growth in the Sepn1 knock-out mouse model. By contrast, we recently demonstrated that SelN is essential for muscle regeneration and satellite cell maintenance in mice and humans, hence opening new avenues regarding the pathomechanism(s) leading to SEPN1-related myopathies. At the cellular level, recent data suggested that SelN participates in oxidative and calcium homeostasis, with a potential role in the regulation of the ryanodine receptor activity. Despite the recent and exciting progress regarding the physiological function(s) of SelN in muscle tissue, the pathogenesis leading to SEPN1-related myopathies remains largely unknown, with several unsolved questions, and no treatment available. In this review, we introduce SelN, its properties and expression pattern in zebrafish, mice, and humans, and we discuss its potential roles in muscle tissue and the ensuing clues for the development of therapeutic options.

Dantrolene ameliorates cognitive decline and neuropathology in Alzheimer triple transgenic mice

Disruption of intracellular calcium homeostasis via abnormal and excessive activation of ryanodine receptors plays an important role in the neuropathology of Alzheimer's disease. We investigated the therapeutic effect of dantrolene, a ryanodine receptor antagonist, on cognitive dysfunction and neuropathology in the triple transgenic Alzheimer mouse model (3xTg-AD). 3xTg-AD mice were treated with dantrolene from 2 to 13 months of age. Learning and memory were measured with the Morris Water Maze at 6, 10, and 13 months of age. Amyloid and tau neuropathology and biomarkers for synaptic dysfunction and neurodegeneration were examined in the brain using immunoblotting or immunohistochemistry. Dantrolene treatment for 11 months significantly reduced both memory deficits and amyloid plaque load in the hippocampus in 13-month-old 3xTg-AD mice. Dantrolene treatment, however, had no effect on phosphorylated tau, phosphorylated or total GSK-3β, synaptic markers, or mitochondrial or cytosolic cytochrome C. Our results suggest that dantrolene significantly improves cognition in a murine model of Alzheimer's disease and is associated with a reduction in amyloid plaque burden, forming the basis for a novel therapeutic approach for Alzheimer's disease.

Cyclic Nucleotides Cause Divergent Ryanodine Receptor Modulation in Pulmonary Arterial Myocytes from Immature Chronic Hypoxic Sheep

Calcium sparks are due to ryanodine receptor (RyR) activation and are intimate to local and global calcium signals in pulmonary arterial (PA) myocytes. RyRs are heavily regulated and cyclic nucleotides, including cAMP and cGMP, can increase RyR activity and thereby augment Ca2+ sparks and global Ca2+ waves. One possibility is that enhanced RyR activity contributes to altered pulmonary arterial reactivity responses found in sheep born at high altitude. Yet, the influence of chronic hypoxia (CH) on cyclic nucleotide regulation of RyRs is unknown. In these studies, we tested the hypothesis that cAMP and cGMP increase Ca2+ sparks and waves in PA myocytes from hypoxic sheep. Spark and wave activity was determined by visual analysis of line‐scan confocal recordings of Fluo‐4 or Fluo‐8 loaded PA from term‐fetal, ~ 10 day old, or adult sheep that lived at 3,200 meters for &lt;100 days. Similar percentages of fetal, newborn, and adult myocytes had sparks and waves before cyclic nucleotide treatment, although newborns had more sparks per recording. cAMP and cGMP increased the number of adult cells with sparks and waves, yet, they respectively decreased the number of newborn and fetal myocytes with sparks. Spark activity was increased in adult myocytes by cGMP. The data suggest that cyclic nucleotides cause divergent regulation of RyRs from PA myocytes of immature CH sheep. NSF MRI 092355 (SMW), NIH P01HD031226, R01HD003807 (LDL)

Chlorantraniliprole Susceptibility in Leptinotarsa decemlineata in the North Xinjiang Uygur Autonomous Region in China

The Colorado potato beetle (Leptinotarsa decemlineata (Say)) in the north Xinjiang Uygur autonomous region has evolved resistance to various types of insecticides. Chlorantraniliprole is a novel anthranilic diamide insecticide that binds and activates ryanodine receptors. It exhibited excellent efficacy against L. decemlineata in several field trails in Europe. In the present paper, the susceptibility of L. decemlineata fourth-instar larvae derived from six field populations and L. decemlineata adults derived from three field populations to chlorantraniliprole was determined by a topical application. The fourth-instar larvae were substantially more susceptible to chlorantraniliprole than adults, although the range of susceptibility was far greater among the fourth-instar larvae. Regarding stomach toxicities, adult beetles were less susceptible to chlorantraniliprole than larvae. Chlorantraniliprole was most toxic to second-instar larvae, followed by third- and fourth-instar larvae. These data suggested that the appropriate timing for chlorantraniliprole spraying is the early larval stage. Moreover, the synergistic activities of chlorantraniliprole in combination with triphenyl phosphate, diethyl maleate, or piperonyl butoxide against fourth-instar larvae from two field populations and adults from one field population were tested. Piperonyl butoxide had synergistic effects with chlorantraniliprole against fourth-instar larvae but not against adult beetles. Conversely, triphenyl phosphate and diethyl maleate exerted little synergistic effects. It appears that there is a potential risk of resistance against chlorantraniliprole resulting from cytochrome P450 monooxygenase activity.

Can the Sodium-Calcium Exchanger Initiate or Suppress Calcium Sparks in Cardiac Myocytes?

Positive feedback of Calcium (Ca)-induced Ca release is the mechanism of Ca spark formation in cardiac myocytes. To initiate this process, a certain amount of Ca in the cleft space is necessary. When the membrane potential becomes higher during excitation-contraction coupling, Ca can enter through both Ca current (ICaL) and sodium-calcium exchanger (NCX) and may activate ryanodine receptors to initiate a Ca spark. On the other hand, at the resting membrane potential (Vm ∼–80 mV), NCX removes Ca from the cell (forward mode). If Ca released from the sarcoplasmic reticulum is quickly removed via forward mode NCX before Ca-induced Ca release starts, the Ca release becomes nonspark Ca leak. This would also be influenced by the cleft/noncleft distribution of NCX, which is unknown. Using a physiologically detailed mathematical model of subcellular Ca cycling, we analyze how NCX strength and distribution alter Ca spark formation. During excitation-contraction coupling, most Ca sparks are induced by ICaL with very few due to NCX current. At the resting membrane potential if most NCX is localized to the cleft, spontaneous Ca sparks are significantly reduced.