Activation of A1 reactive astrocytes in the medullary dorsal horn of rats participates in the chronification of trigeminal neuralgia. / å¤§é¼ å»¶é«“èƒŒè§’A1åž‹ååº”æ€§æ˜Ÿå½¢èƒ¶è´¨ç»†èƒžæ¿€æ´»å‚ä¸Žä¸‰å‰ç¥žç»ç—›æ ¢æ€§åŒ–.

OBJECTIVES: The activation of astrocytes is an important process in the formation of chronic pain. This study aims to observe the activation of A1 reactive astrocytes in the medullary dorsal horn in the rat model of trigeminal neuralgia, and to explore the mechanism of central sensitization caused by A1 reactive astrocyte. METHODS: The adult male rats were randomly divided into a sham group and a chronic constriction injury of infraorbital nerve (ION-CCI) group. The facial mechanical pain threshold and thermal withdrawal latency were measured before the operation and on the 1st, 3rd, 7th, 10th, and 14th day after the operation. After pain behavior observation, the expression of glial fibrillary acidic protein (GFAP) in the medullary dorsal horn was observed by immunohistochemistry and immunofluorescence colocalization of GFAP, complement 3 (C3)/S100A10, and 4', 6-diamidino-2-phenylindole (DAPI) was analyzed. Primary astrocytes were cultured and randomly divided into a naive group and a DHK group. The DHK group was treated with 1 mmol/L of astrocyte activation inhibitor dihydrokainic acid (DHK). Fura-2/AM was used to stain the astrocytes and the calcium wave of the 2 groups under the stimulation of high potassium was recorded and compared. The expression of C3 was detected by Western blotting. RESULTS: The facial mechanical pain threshold and thermal withdrawal latency of the ION-CCI group were significantly lower than those of the sham group (both P<0.05). There were a large number of GFAP positive astrocytes in the medullary dorsal horn of the ION-CCI group. The fluorescence intensity of GFAP in the ION-CCI group was higher than that in the sham group (P<0.05). GFAP and C3/S100A10 were co-expressed in astrocytes. Compared with the sham group, the fluorescence intensity of C3 and the protein expression of C3 in the ION-CCI group were increased (both P<0.05). The expression of C3 in ION-CCI group was significantly increased (P<0.05). Compared with the naive group, the C3 protein expression was significantly decreased in the DHK group (P<0.05). The intensity of calcium fluorescence was increased after high potassium stimulation in both groups. Furthermore, the peak and increase amplitude of calcium fluorescence in the naive group were much higher than those in the DHK group (both P<0.05). CONCLUSIONS: A1 reactive astrocytes in the medullary dorsal horn of trigeminal neuralgia model rats are increased significantly, which may participate in central sensitization of trigeminal neuralgia by impacting astrocyte calcium wave.

Zoom-in to molecular mechanisms underlying root growth and function under heterogeneous soil environment and abiotic stresses.

MAIN CONCLUSION: The review describes tissue-specific and non-cell autonomous molecular responses regulating the root system architecture and function in plants. Phenotypic plasticity of roots relies on specific molecular and tissue specific responses towards local and microscale heterogeneity in edaphic factors. Unlike gravitropism, hydrotropism in Arabidopsis is regulated by MIZU KUSSIE1 (MIZ1)-dependent asymmetric distribution of cytokinin and activation of Arabidopsis response regulators, ARR16 and ARR17 on the lower water potential side of the root leading to higher cell division and root bending. The cortex specific role of Abscisic acid (ABA)-activated SNF1-related protein kinase 2.2 (SnRK2.2) and MIZ1 in elongation zone is emerging for hydrotropic curvature. Halotropism involves clathrin-mediated internalization of PIN FORMED 2 (PIN2) proteins at the side facing higher salt concentration in the root tip, and ABA-activated SnRK2.6 mediated phosphorylation of cortical microtubule-associated protein Spiral2-like (SP2L) in the root transition zone, which results in anisotropic cell expansion and root bending away from higher salt. In hydropatterning, Indole-3-acetic acid 3 (IAA3) interacts with SUMOylated-ARF7 (Auxin response factor 7) and prevents expression of Lateral organ boundaries-domain 16 (LBD16) in air-side of the root, while on wet side of the root, IAA3 cannot repress the non-SUMOylated-ARF7 thereby leading to LBD16 expression and lateral root development. In root vasculature, ABA induces expression of microRNA165/microRNA166 in endodermis, which moves into the stele to target class III Homeodomain leucine zipper protein (HD-ZIP III) mRNA in non-cell autonomous manner. The bidirectional gradient of microRNA165/6 and HD-ZIP III mRNA regulates xylem patterning under stress. Understanding the tissue specific molecular mechanisms regulating the root responses under heterogeneous and stress environments will help in designing climate-resilient crops.

A norepinephrine-dependent glial calcium wave travels in the spinal cord upon acoustovestibular stimuli.

Although calcium waves have been widely observed in glial cells, their occurrence in vivo during behavior remains less understood. Here, we investigated the recruitment of glial cells in the hindbrain and spinal cord after acousto-vestibular (AV) stimuli triggering escape responses using in vivo population calcium imaging in larval zebrafish. We observed that gap-junction-coupled spinal glial network exhibits large and homogenous calcium increases that rose in the rostral spinal cord and propagated bi-directionally toward the spinal cord and toward the hindbrain. Spinal glial calcium waves were driven by the recruitment of neurons and in particular, of noradrenergic signaling acting through α-adrenergic receptors. Noradrenergic neurons of the medulla-oblongata (NE-MO) were revealed in the vicinity of where the calcium wave started. NE-MO were recruited upon AV stimulation and sent dense axonal projections in the rostro-lateral spinal cord, suggesting these cells could trigger the glial wave to propagate down the spinal cord. Altogether, our results revealed that a simple AV stimulation is sufficient to recruit noradrenergic neurons in the brainstem that trigger in the rostral spinal cord two massive glial calcium waves, one traveling caudally in the spinal cord and another rostrally into the hindbrain.

Increasing SERCA function promotes initiation of calcium sparks and breakup of calcium waves.

KEY POINTS: Increasing sarcoplasmic/endoplasmic reticulum calcium ATPase (SERCA) pump activity enhances sarcoplasmic reticulum calcium (Ca) load, which increases both ryanodine receptor opening and driving force of Ca release flux. Both of these effects promote Ca spark formation and wave propagation. However, increasing SERCA activity also accelerates local cytosolic Ca decay as the wave front travels to the next cluster, which limits wave propagation. As a result, increasing SERCA pump activity has a biphasic effect on the propensity of arrhythmogenic Ca waves, but a monotonic effect to increase Ca spark frequency and amplitude. ABSTRACT: Waves of sarcoplasmic reticulum (SR) calcium (Ca) release can cause arrhythmogenic afterdepolarizations in cardiac myocytes. Ca waves propagate when Ca sparks at one Ca release unit (CRU) recruit new Ca sparks in neighbouring CRUs. Under normal conditions, Ca sparks are too small to recruit neighbouring Ca sparks where Ca sensitivity is also low. However, under pathological conditions such as a Ca overload or ryanodine receptor (RyR) sensitization, Ca sparks can be larger and propagate more readily as macro-sparks or full Ca waves. Increasing SERCA pump activity promotes SR Ca load, which promotes RyR opening and increases driving force of the Ca release flux from SR to cytosol, promoting Ca waves. However, high sarcoplasmic/endoplasmic reticulum calcium ATPase (SERCA) activity can also decrease local cytosolic [Ca] as it approaches the next CRU, thereby reducing wave appearance and propagation. In this study, we use a physiologically detailed model of subcellular Ca cycling and experiments in phospholamban-knockout mice, to show how Ca waves are initiated and propagate and how different conditions contribute to the generation and propagation of Ca waves. We show that reducing diffusive coupling between Ca sparks by increasing SERCA activity prevents Ca waves by reducing [Ca] at the next CRU, as do Ca buffers, low intra-SR Ca diffusion and distance between CRUs. Increasing SR Ca uptake rate has a biphasic effect on Ca wave propagation; initially it enhances Ca spark probability and amplitude and CRU coupling, thereby promoting arrhythmogenic Ca wave propagation, but at higher levels SR Ca uptake can abort those arrhythmogenic Ca waves.

Rapid systemic signaling during abiotic and biotic stresses: is the ROS wave master of all trades?

Rapidly communicating the perception of an abiotic stress event, wounding or pathogen infection, from its initial site of occurrence to the entire plant, i.e. rapid systemic signaling, is essential for successful plant acclimation and defense. Recent studies highlighted an important role for several rapid whole-plant systemic signals in mediating plant acclimation and defense during different abiotic and biotic stresses. These include calcium, reactive oxygen species (ROS), hydraulic and electric waves. Although the role of some of these signals in inducing and coordinating whole-plant systemic responses was demonstrated, many questions related to their mode of action, routes of propagation and integration remain unanswered. In addition, it is unclear how these signals convey specificity to the systemic response, and how are they integrated under conditions of stress combination. Here we highlight many of these questions, as well as provide a proposed model for systemic signal integration, focusing on the ROS wave.

Kulikovia alborostrata and Kulikovia fulva comb. nov. (Nemertea: Heteronemertea) are Sister Species with Prezygotic Isolating Barriers.

The heteronemertean Kulikovia alborostrata (Takakura, 1898) was originally described as Lineus alborostratus based on material from Misaki, Japan. Although this species was regarded as consisting of two color variants, purple and brown-yellow, the identity of these variants has never been examined based on topotypes. In this study, we performed a multi-locus phylogeny reconstruction, species delimitation analyses, and cross-fertilization experiments to examine the species status of Takakura's original taxon concept consisting of these color variants. Our results suggest that the purple type is identical to Lineus alborostratus Takakura, 1898 auct. (currently Kulikovia alborostrata), whereas the brown-yellow type is conspecific with Lineus fulvus Iwata, 1954, originally established from Hokkaido. These two species appear to have a sister-taxon relationship and are reproductively isolated from each other by prezygotic mechanisms involving gamete incompatibility, minimally separated with 2.8% (16S rRNA) and 14.4% (COI) uncorrected p-distances. We propose that the purple type be considered as representing the true identity of the nominal species Lineus alborostratus (currently assigned to the genus Kulikovia) to maintain the common usage of the name. Although Takakura's type material is not extant, we consider that neotypification is unnecessary in this case because no taxonomic/nomenclatural confusion persists. We also propose to transfer Lineus fulvus to yield Kulikovia fulva comb. nov.

Dynamics and mechanisms of intracellular calcium waves elicited by tandem bubble-induced jetting flow.

One of the earliest events in cellular mechanotransduction is often an increase in intracellular calcium concentration associated with intracellular calcium waves (ICWs) in various physiologic or pathophysiologic processes. Although cavitation-induced calcium responses are believed to be important for modulating downstream bioeffects such as cell injury and mechanotransduction in ultrasound therapy, the fundamental mechanisms of these responses have not been elucidated. In this study, we investigated mechanistically the ICWs elicited in single HeLa cells by the tandem bubble-induced jetting flow in a microfluidic system. We identified two distinct (fast and slow) types of ICWs at varying degrees of flow shear stress-induced membrane deformation, as determined by different bubble standoff distances. We showed that ICWs were initiated by an extracellular calcium influx across the cell membrane nearest to the jetting flow, either primarily through poration sites for fast ICWs or opening of mechanosensitive ion channels for slow ICWs, which then propagated in the cytosol via a reaction-diffusion process from the endoplasmic reticulum. The speed of ICW (CICW ) was found to correlate strongly with the severity of cell injury, with CICW in the range of 33 µm/s to 93 µm/s for fast ICWs and 1.4 µm/s to 12 µm/s for slow ICWs. Finally, we demonstrated that micrometer-sized beads attached to the cell membrane integrin could trigger ICWs under mild cavitation conditions without collateral injury. The relation between the characteristics of ICW and cell injury, and potential strategies to mitigate cavitation-induced injury while evoking an intracellular calcium response, may be particularly useful for exploiting ultrasound-stimulated mechanotransduction applications in the future.

Insemination or phosphatidic acid induces an outwardly spiraling disk of elevated Ca2+ to produce the Ca2+ wave during Xenopus laevis fertilization.

During Xenopus fertilization, the initial intracellular calcium ((Ca2+)i) release at the sperm-egg binding site (hot spot) has not been described without the use of inhibitors, nor related to underlying ER structure. Without inhibitors, we now report that sperm induce an initial hot spot after sperm addition to Xenopus eggs that was ~25 µm. This area is consistent with the size of ER patches and clusters of IP3 receptors that have enhanced activity. Furthermore, we find a new mechanism for the fertilization (Ca2+)i wave; instead of outward diffusion of inositol 1,4,5-trisphosphate (IP3), we find that the wave was generated by an outward, clockwise rotation of a ~63 µm disk of elevated (Ca2+)i moving very rapidly at ~65 µm/s. We also suggest a new mechanism for the acceleration of the fertilization (Ca2+)i wave as the disk accelerated and was joined by other rotating disks (some rotating counterclockwise) at a time when the speed of the (Ca2+)i wave increases. To examine the role of phosphatidic acid (PA) in the release of (Ca2+)i during Xenopus fertilization, we find that two inhibitors of PA production delayed the appearance of fertilization hot spots by ~9-12 min but did not reduce the size of hot spots and actually accelerated the later (Ca2+)i wave. Surprisingly, global addition of PA to Xenopus eggs induced localized hot spots at a time and size that was similar to those induced after sperm addition. In contrast, sperm induce a rapid (Ca2+)i wave (~4 µm/s) within ~30 s after hot spot appearance, whereas hot spots induced by PA required an ~32 min to induce a very slow (~1 µm/s) (Ca2+)i wave with a lower peak of (Ca2+)i. Thus, PA may not be required for the initial release of (Ca2+)i at the sperm-egg binding site, but mimics sperm by inducing a similarly sized localized (Ca2+)i release. As compared with sperm, PA may induce a weak, slow (Ca2+)i wave by slowly increasing IP3 receptor clustering. Addition of PA to Xenopus oocytes, or Ca2+ ionophore to either Xenopus oocytes or eggs, did not induce hot spots but a global (Ca2+)i wave that rapidly moved at ~12 µm/s.

Gap junction-dependent coordination of intercellular calcium signalling in the developing appendicularian tunicate Oikopleura dioica.

We characterized spontaneous Ca2+ signals in Oikopleura dioica embryos from pre-fertilization to gastrula stages following injection of GCaMP6 mRNA into unfertilized eggs. The unfertilized egg exhibited regular, transient elevations in intracellular Ca2+ concentration with an average duration of 4-6 s and an average frequency of about 1 every 2.5 min. Fertilization was accompanied by a longer Ca2+ transient that lasted several minutes. Thereafter, regular Ca2+ transients were reinstated that spread within seconds among blastomeres and gradually increased in duration (by about 50%) and decreased in frequency (by about 20%) by gastrulation. Peak amplitudes also exhibited a dynamic, with a transitory drop occurring at about the 4-cell stage and a subsequent rise. Each peak was preceded by about 15 s by a smaller and shorter Ca2+ increase (about 5% of the main peak amplitude, average duration 3 s), which we term the "minipeak". By gastrulation, Ca2+ transients exhibited a stereotyped initiation site on either side of the 32-64-cell embryo, likely in the nascent muscle precursor cells, and spread thereafter symmetrically in a stereotyped spatial pattern that engaged blastomeres giving rise to all the major tissue lineages. The rapid spread of the transients relative to the intertransient interval created a coordinated wave that, on a coarse time scale, could be considered an approximate synchronization. Treatment with the divalent cations Ni2+ or Cd2+ gradually diminished peak amplitudes, had only moderate effects on wave frequency, but markedly disrupted wave synchronization and normal development. The T-type Ca2+ channel blocker mibefradil similarly disrupted normal development, and eliminated the minipeaks, but did not affect wave synchronization. To assess the role of gap junctions in calcium wave spread and coordination, we first characterized the expression of two Oikopleura connexins, Od-CxA and Od-CxB, both of which are expressed during pre-gastrulation and gastrula stages, and then co-injected double-stranded inhibitory RNAs together with CGaMP6 to suppress connexin expression. Connexin mRNA knockdown led to a gradual increase in Ca2+ transient peak width, a decrease of interpeak interval and a marked disruption of wave synchronization. As seen with divalent cations and mibefradil, this desynchronization was accompanied by a disruption of normal development.

A calcium-mediated actin redistribution at egg activation in Drosophila.

Egg activation is the essential process in which mature oocytes gain the competency to proceed into embryonic development. Many events of egg activation are conserved, including an initial rise of intracellular calcium. In some species, such as echinoderms and mammals, changes in the actin cytoskeleton occur around the time of fertilization and egg activation. However, the interplay between calcium and actin during egg activation remains unclear. Here, we use imaging, genetics, pharmacological treatment, and physical manipulation to elucidate the relationship between calcium and actin in living Drosophila eggs. We show that, before egg activation, actin is smoothly distributed between ridges in the cortex of the dehydrated mature oocytes. At the onset of egg activation, we observe actin spreading out as the egg swells though the intake of fluid. We show that a relaxed actin cytoskeleton is required for the intracellular rise of calcium to initiate and propagate. Once the swelling is complete and the calcium wave is traversing the egg, it leads to a reorganization of actin in a wavelike manner. After the calcium wave, the actin cytoskeleton has an even distribution of foci at the cortex. Together, our data show that calcium resets the actin cytoskeleton at egg activation, a model that we propose to be likely conserved in other species.

Regulation of Trpm activation and calcium wave initiation during Drosophila egg activation.

The transition from a developmentally arrested mature oocyte to a developing embryo requires a series of highly conserved events, collectively known as egg activation. All of these events are preceded by a ubiquitous rise of intracellular calcium, which results from influx of external calcium and/or calcium release from internal storage. In Drosophila, this calcium rise initiates from the pole(s) of the oocyte by influx of external calcium in response to mechanical triggers. It is thought to trigger calcium responsive kinases and/or phosphatases, which in turn alter the oocyte phospho-proteome to initiate downstream events. Recent studies revealed that external calcium enters the activating Drosophila oocyte through Trpm channels, a feature conserved in mouse. The local entry of calcium raises the question of whether Trpm channels are found locally at the poles of the oocyte or are localized around the oocyte periphery, but activated only at the poles. Here, we show that Trpm is distributed all around the oocyte. This requires that it thus be specially regulated at the poles to allow calcium wave initiation. We show that neither egg shape nor local pressure is sufficient to explain this local activation of Trpm channels.

Transient Elevation of Glucose Increases Arrhythmia Susceptibility in Non-Diabetic Rat Trabeculae With Non-Uniform Contraction.

BACKGROUND: In non-diabetic patients with acute coronary syndrome, stress hyperglycemia occasionally occurs and is related to their mortality. Whether transient elevation of glucose affects arrhythmia susceptibility in non-diabetic hearts with non-uniform contraction was examined.Methods and Results:Force, intracellular Ca2+([Ca2+]i), and membrane potential were measured in trabeculae from rat hearts. Non-uniform contraction was produced by a jet of paralyzing solution. Ca2+waves and arrhythmias were induced by electrical stimulation (2.0 mmol/L [Ca2+]o). The activity of Ca2+/calmodulin-dependent protein kinaseII (CaMKII) was measured. An elevation of glucose from 150 to 400 mg/dL increased the velocity of Ca2+waves and the number of spontaneous action potentials triggered by electrical stimulation. Besides, the elevation of glucose increased the CaMKII activity. In the presence of 1 µmol/L KN-93, the elevation of glucose did not increase the velocity of Ca2+waves and the number of triggered action potentials. In addition, in the presence of 1 µmol/L autocamtide-2 related inhibitory peptide or 50 µmol/L diazo-5-oxonorleucine, the elevation of glucose did not increase the number of triggered action potentials. Furthermore, the elevation of glucose by adding L-glucose did not increase their number. CONCLUSIONS: In non-diabetic hearts with non-uniform contraction, transient elevation of glucose increases the velocity of Ca2+waves by activating CaMKII,probably through glycosylation with O-linked ß-N-acetylglucosamine, thereby increasing arrhythmia susceptibility.

Stochastic initiation and termination of calcium-mediated triggered activity in cardiac myocytes.

Cardiac myocytes normally initiate action potentials in response to a current stimulus that depolarizes the membrane above an excitation threshold. Aberrant excitation can also occur due to spontaneous calcium (Ca2+) release (SCR) from intracellular stores after the end of a preceding action potential. SCR drives the Na+/Ca2+ exchange current inducing a "delayed afterdepolarization" that can in turn trigger an action potential if the excitation threshold is reached. This "triggered activity" is known to cause arrhythmias, but how it is initiated and terminated is not understood. Using computer simulations of a ventricular myocyte model, we show that initiation and termination are inherently random events. We determine the probability of those events from statistical measurements of the number of beats before initiation and before termination, respectively, which follow geometric distributions. Moreover, we elucidate the origin of randomness by a statistical analysis of SCR events, which do not follow a Poisson process observed in other eukaryotic cells. Due to synchronization of Ca2+ releases during the action potential upstroke, waiting times of SCR events after the upstroke are narrowly distributed, whereas SCR amplitudes follow a broad normal distribution with a width determined by fluctuations in the number of independent Ca2+ wave foci. This distribution enables us to compute the probabilities of initiation and termination of bursts of triggered activity that are maintained by a positive feedback between the action potential upstroke and SCR. Our results establish a theoretical framework for interpreting complex and varied manifestations of triggered activity relevant to cardiac arrhythmias.

Characterization of calcium transients during early embryogenesis in ascidians Ciona robusta (Ciona intestinalis type A) and Ciona savignyi.

The calcium ion (Ca2+) is an important second messenger, and a rapid increase in Ca2+ level (Ca2+ transient) is involved in various aspects of embryogenesis. Although Ca2+ transients play an important role in early developmental stages, little is known about their dynamics throughout embryogenesis. Here, Ca2+ transients were characterized by visualizing Ca2+ dynamics in developing chordate embryos using a fluorescent protein-based Ca2+ indicator, GCaMP6s in combination with finely tuned microscopy. Ca2+ transients were detected in precursors of muscle cells in the late gastrula stage. In the neurula stage, repetitive Ca2+ transients were observed in left and right neurogenic cells, including visceral ganglion (VG) precursors, and the duration of Ca2+ transients was 39±4s. In the early tailbud stage, Ca2+ transients were observed in differentiating precursors of nerve cord neurons. A small population of VG precursors showed rhythmical Ca2+ transients with a duration of 22±4s, suggesting a central pattern generator (CPG) origin. At the mid tailbud stage, Ca2+transients were observed in a wide area of epidermal cells and named CTECs. The number and frequency of CTECs increased drastically in late tailbud stages, and the timing of the increase coincided with that of the relaxation of the tail bending. The experiment using Ca2+ chelator showed that the CTECs were largely depending on the extracellular Ca2+. The waveform analysis of Ca2+ transients revealed different features according to duration and frequency. The comprehensive characterization of Ca2+ transients during early ascidian embryogenesis will help our understanding of the role of Ca2+ signaling in chordate embryogenesis.

Spontaneous calcium waves in the developing enteric nervous system.

The enteric nervous system (ENS) is an extensive network of neurons in the gut wall that arises from neural crest-derived cells. Like other populations of neural crest cells, it is known that enteric neural crest-derived cells (ENCCs) influence the behaviour of each other and therefore must communicate. However, little is known about how ENCCs communicate with each other. In this study, we used Ca2+ imaging to examine communication between ENCCs in the embryonic gut, using mice where ENCCs express a genetically-encoded calcium indicator. Spontaneous propagating calcium waves were observed between neighbouring ENCCs, through both neuronal and non-neuronal ENCCs. Pharmacological experiments showed wave propagation was not mediated by gap junctions, but by purinergic signalling via P2 receptors. The expression of several P2X and P2Y receptors was confirmed using RT-PCR. Furthermore, inhibition of P2 receptors altered the morphology of the ENCC network, without affecting neuronal differentiation or ENCC proliferation. It is well established that purines participate in synaptic transmission in the mature ENS. Our results describe, for the first time, purinergic signalling between ENCCs during pre-natal development, which plays roles in the propagation of Ca2+ waves between ENCCs and in ENCC network formation. One previous study has shown that calcium signalling plays a role in sympathetic ganglia formation; our results suggest that calcium waves are likely to be important for enteric ganglia development.

Adenosine triphosphate acts as a paracrine signaling molecule to reduce the motility of T cells.

Organization of immune responses requires exchange of information between cells. This is achieved through either direct cell-cell contacts and establishment of temporary synapses or the release of soluble factors, such as cytokines and chemokines. Here we show a novel form of cell-to-cell communication based on adenosine triphosphate (ATP). ATP released by stimulated T cells induces P2X4/P2X7-mediated calcium waves in the neighboring lymphocytes. Our data obtained in lymph node slices suggest that, during T-cell priming, ATP acts as a paracrine messenger to reduce the motility of lymphocytes and that this may be relevant to allow optimal tissue scanning by T cells.

Peeking at a plant through the holes in the wall - exploring the roles of plasmodesmata.

Plasmodesmata (PD) are membrane-lined pores that connect neighbouring plant cells and allow molecular exchange via the symplast. Past studies have revealed the basic structure of PD, some of the transport mechanisms for molecules through PD, and a variety of physiological processes in which they function. Recently, with the help of newly developed technologies, several exciting new features of PD have been revealed. New PD structures were observed during early formation of PD and between phloem sieve elements and phloem pole pericycle cells in roots. Both observations challenge our current understanding of PD structure and function. Research into novel physiological responses, which are regulated by PD, indicates that we have not yet fully explored the potential contribution of PD to overall plant function. In this Viewpoint article, we summarize some of the recent advances in understanding the structure and function of PD and propose the challenges ahead for the community.

Towards an integrated understanding of cardiac arrhythmogenesis - Growing roles of experimental pathology.

Cardiac arrhythmias have long been regarded as derangement of electrical impulse initiation and conduction within the heart. However, underlying mechanisms for arrhythmogenesis are not fully understood solely from the electrophysiological viewpoint. This review article discusses pathogenesis of arrhythmias from non-electrical aspects, which were elucidated by spatiotemporal imaging of functional molecules in combination with morphological analysis of living heart tissues. Intracellular Ca2+ ([Ca2+ ]i ) overload, caused by myocardial injury, provokes Ca2+ waves that could lead to abnormal excitations, i.e., triggered arrhythmias. Depressed Ca2+ release from the sarcoplasmic reticulum, caused by ischemia, heart failure, or T-tubular remodeling, results in spatiotemporally inhomogeneous [Ca2+ ]i dynamics that could disturb impulse conduction, leading to reentrant tachyarrhythmias. Impairment of the gap junction-mediated intercellular communications, which provokes derangement of impulse propagation of the myocardium, also leads to reentrant arrhythmias. Interpositions of non-cardiomyocytes, especially fibroblasts, in the myocardium could also contribute to arrhythmogenesis via heterocellular gap-junctional coupling with cardiomyocytes. Furthermore, alterations in myocardial histology, e.g., density and arrangements of myocytes in association with gap-junctional distributions, could constitute important pathologic bases of atrial fibrillation. Integration of these molecular, functional, and morphological features of the myocardium, unveiled by experimental pathological approaches, would pave a new way for understanding pathogenesis of cardiac arrhythmias.

Unifying principles of calcium wave propagation - Insights from a three-dimensional model for atrial myocytes.

Atrial myocytes in a number of species lack transverse tubules. As a consequence the intracellular calcium signals occurring during each heartbeat exhibit complex spatio-temporal dynamics. These calcium patterns arise from saltatory calcium waves that propagate via successive rounds of diffusion and calcium-induced calcium release. The many parameters that impinge on calcium-induced calcium release and calcium signal propagation make it difficult to know a priori whether calcium waves will successfully travel, or be extinguished. In this study, we describe in detail a mathematical model of calcium signalling that allows the effect of such parameters to be independently assessed. A key aspect of the model is to follow the triggering and evolution of calcium signals within a realistic three-dimensional cellular volume of an atrial myocyte, but with low computational costs. This is achieved by solving the linear transport equation for calcium analytically between calcium release events and by expressing the onset of calcium liberation as a threshold process. The model makes non-intuitive predictions about calcium signal propagation. For example, our modelling illustrates that the boundary of a cell produces a wave-guiding effect that enables calcium ions to propagate further and for longer, and can subtly alter the pattern of calcium wave movement. The high spatial resolution of the modelling framework allows the study of any arrangement of calcium release sites. We demonstrate that even small variations in randomly positioned release sites cause highly heterogeneous cellular responses. This article is part of a Special Issue entitled: 13th European Symposium on Calcium.

TRPV4 is endogenously expressed in vertebrate spermatozoa and regulates intracellular calcium in human sperm.

Transient Receptor Potential Vanilloid sub-type 4 (TRPV4) is a non-selective cationic channel involved in regulation of temperature, osmolality and different ligand-dependent Ca(2+)-influx. Recently, we have demonstrated that TRPV4 is conserved in all vertebrates. Now we demonstrate that TRPV4 is endogenously expressed in all vertebrate sperm cells ranging from fish to mammals. In human sperm, TRPV4 is present as N-glycosylated protein and its activation induces Ca(2+)-influx. Its expression and localization differs in swim-up and swim-down cells suggesting that TRPV4 is an important determining factor for sperm motility. We demonstrate that pharmacological activation or inhibition of TRPV4 regulates Ca(2+)-wave propagation from head to tail. Such findings may have wide application in male fertility-infertility, contraception and conservation of endangered species as well.