Pannexin 1 (Panx1) represents a class of vertebrate membrane channels, bearing significant sequence homology with the invertebrate gap junction proteins, the innexins and more distant similarities in ...the membrane topologies and pharmacological sensitivities with gap junction proteins of the connexin family. In the nervous system, cooperation among pannexin channels, adenosine receptors, and K(ATP) channels modulating neuronal excitability via ATP and adenosine has been recognized, but little is known about the significance in vivo. However, the localization of Panx1 at postsynaptic sites in hippocampal neurons and astrocytes in close proximity together with the fundamental role of ATP and adenosine for CNS metabolism and cell signaling underscore the potential relevance of this channel to synaptic plasticity and higher brain functions. Here, we report increased excitability and potently enhanced early and persistent LTP responses in the CA1 region of acute slice preparations from adult Panx1(-/-) mice. Adenosine application and N-methyl-D-aspartate receptor (NMDAR)-blocking normalized this phenotype, suggesting that absence of Panx1 causes chronic extracellular ATP/adenosine depletion, thus facilitating postsynaptic NMDAR activation. Compensatory transcriptional up-regulation of metabotropic glutamate receptor 4 (grm4) accompanies these adaptive changes. The physiological modification, promoted by loss of Panx1, led to distinct behavioral alterations, enhancing anxiety and impairing object recognition and spatial learning in Panx1(-/-) mice. We conclude that ATP release through Panx1 channels plays a critical role in maintaining synaptic strength and plasticity in CA1 neurons of the adult hippocampus. This result provides the rationale for in-depth analysis of Panx1 function and adenosine based therapies in CNS disorders.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Abstract Glial cells play a crucial role in the pathomechanism of perinatal hypoxic-ischemic brain injury (HI) and are involved in the maintenance of a chronic state of inflammation that causes ...delayed neuronal damage. Activation of astrocytes is one factor prolonging brain damage and contributing to the formation of a glial scar that limits neuronal plasticity. In this context, the major astrocytic gap junction protein Connexin 43 (Cx43) has been ascribed various functions including regulation of astrocytic migration and proliferation. Here, we investigate glial responses like microglia/macrophages and astrocytic activation in a rat model of neonatal HI and characterize changes of these parameters upon transplantation of human umbilical cord blood cells (hUCB). As an alleviation of motor function in lesioned rats has previously been described in transplanted animals, we analyze the putative correlation between motor function and glial activation over time. The lesion-induced impairment of motor function, assessed by forelimb use bias, muscle strength and distal spasticity, was alleviated upon transplantation of hUCB short and long term. HI induced an acute inflammatory reaction with activation of microglia/macrophages and reactive astrogliosis associated with perilesional upregulation of Cx43 that slowly declined during the chronic post-ischemic phase. hUCB transplantation accelerated the regression of inflammatory events, narrowed the perilesional astrocytic wall and led to a downregulation of the investigated astrocytic proteins. Thus, in the immature brain, hUCB may indirectly reduce secondary cell death upon hypoxia-ischemia and facilitate post-ischemic plasticity through the attenuation of reactive gliosis. This article is part of a Special Issue entitled Electrical Synapses.
A large conductance (∼300 picosiemens) channel (LCC) of unknown molecular identity, activated by Ca2+ release from the sarcoplasmic reticulum, particularly when augmented by caffeine, has been ...described previously in isolated cardiac myocytes. A potential candidate for this channel is pannexin 1 (Panx1), which has been shown to form large ion channels when expressed in Xenopus oocytes and mammalian cells. Panx1 function is implicated in ATP-mediated auto-/paracrine signaling, and a crucial role in several cell death pathways has been suggested. Here, we demonstrate that after culturing for 4 days LCC activity is no longer detected in myocytes but can be rescued by adenoviral gene transfer of Panx1. Endogenous LCCs and those related to expression of Panx1 share key pharmacological properties previously used for identifying and characterizing Panx1 channels. These data demonstrate that Panx1 constitutes the LCC of cardiac myocytes. Sporadic openings of single Panx1 channels in the absence of Ca2+ release can trigger action potentials, suggesting that Panx1 channels potentially promote arrhythmogenic activities.
Transplantation of human umbilical cord blood (hucb) cells in a model of hypoxic-ischemic brain injury led to the amelioration of lesion-impaired neurological and motor functions. However, the ...mechanisms by which transplanted cells mediate functional recovery after brain injury are largely unknown. In this study, the effects of hucb cell transplantation were investigated in this experimental paradigm at the cellular and molecular level. As the pathological cascade in hypoxic-ischemic brain injury includes inflammation, reduced blood flow, and neuronal cell death, we analyzed the effects of peripherally administered hucb cells on these detrimental processes, investigating the expression of characteristic marker proteins. Application of hucb cells after perinatal hypoxic-ischemic brain injury correlated with an increased expression of the proteins Tie-2 and occludin, which are associated with angiogenesis. Lesion-induced apoptosis, determined by expression of cleaved caspase-3, decreased, whereas the number of vital neurons, identified by counting of NeuN-positive cells, increased. In addition, we observed an increase in the expression of neurotrophic and pro-angiogenic growth factors, namely BDNF and VEGF, in the lesioned brain upon hucb cell transplantation. The release of neurotrophic factors mediated by transplanted hucb cells might cause a lower number of neurons to undergo apoptosis and result in a higher number of living neurons. In parallel, the increase of VEGF might cause growth of blood vessels. Thus, hucb transplantation might contribute to functional recovery after brain injury mediated by systemic or local effects.
Pannexins, a class of membrane channels, bear significant sequence homology with the invertebrate gap junction proteins, innexins, and more distant similarities in their membrane topologies and ...pharmacological sensitivities with the gap junction proteins, connexins. However, the functional role for the pannexin oligomers or pannexons, is different from connexin oligomers, the connexons. Many pannexin publications have used the term "hemichannels" to describe pannexin oligomers while others use the term "channels" instead. This has led to confusion within the literature about the function of pannexins that promotes the idea that pannexons serve as gap junction hemichannels and thus, have an assembly and functional state as gap junctional intercellular channels. Here, we present the case that unlike the connexin gap junction intercellular channels, so far, pannexin oligomers have repeatedly been shown to be channels that are functional in single membranes, but not as intercellular channels in appositional membranes. Hence, they should be referred to as channels and not hemichannels. Thus, we advocate that in the absence of firm evidence that pannexins form gap junctions, the use of the term "hemichannel" be discontinued within the pannexin literature.
In mammals, a single pannexin1 gene (Panx1) is widely expressed in the CNS including the inner and outer retinae, forming large-pore voltage-gated membrane channels, which are involved in calcium and ...ATP signaling. Previously, we discovered that zebrafish lack Panx1 expression in the inner retina, with drPanx1a exclusively expressed in horizontal cells of the outer retina. Here, we characterize a second drPanx1 protein, drPanx1b, generated by whole-genome duplications during teleost evolution. Homology searches strongly support the presence of pannexin sequences in cartilaginous fish and provide evidence that pannexins evolved when urochordata and chordata evolution split. Further, we confirm Panx1 ohnologs being solely present in teleosts. A hallmark of differential expression of drPanx1a and drPanx1b in various zebrafish brain areas is the non-overlapping protein localization of drPanx1a in the outer and drPanx1b in the inner fish retina. A functional comparison of the evolutionary distant fish and mouse Panx1s revealed both, preserved and unique properties. Preserved functions are the capability to form channels opening at resting potential, which are sensitive to known gap junction and hemichannel blockers, intracellular calcium, extracellular ATP and pH changes. However, drPanx1b is unique due to its highly complex glycosylation pattern and distinct electrophysiological gating kinetics. The existence of two Panx1 proteins in zebrafish displaying distinct tissue distribution, protein modification and electrophysiological properties, suggests that both proteins fulfill different functions in vivo.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Until recently, members of the connexin gene family were believed to comprise the sole molecular component forming gap junction channels in vertebrates. The recent discovery of the pannexin gene ...family has challenged this view, as these genes may encode for a putative second class of gap junction proteins in vertebrates. The expression of pannexin genes overlaps with those cellular networks known to exhibit a high degree of gap junctional coupling. We investigated the spatio‐temporal mRNA distribution of one member of this gene family, pannexin1 (Panx1), in the brain and retina of mice using quantitative real‐time polymerase chain reaction and a combination of in situ hybridization and immunohistochemistry for cellular resolution. Our results demonstrate a widespread expression of Panx1 in the brain, retina and other non‐neuronal tissues. In the cortex, cerebellum and eye, Panx1 is expressed at early embryonic time points and peaks around embryonic day 18 followed by a decline towards adulthood. Most notably, Panx1 is detectable in neurons of many brain nuclei, which are known to be coupled by gap junctions as well as in previously unrecognized areas. Abundant expression was found in the adult hippocampal and neocortical pyramidal cells and interneurons, neurons of the reticular thalamus, the inferior olive, magnocellular hypothalamic neurons, midbrain and brain stem motoneurons, Purkinje cells and the retina.
Brain damage around birth may cause lifelong neurodevelopmental deficits. We examined the therapeutic potential of human umbilical cord blood-derived mononuclear cells containing multipotent stem ...cells to facilitate motor recovery after cerebral hypoxic-ischemic damage in neonatal rats. Left carotid artery ligation followed by 8% O(2) inhalation for 80 min was performed on postnatal d 7, succeeded by intraperitoneal transplantation of human umbilical cord blood-derived mononuclear cells on postnatal d 8 in a sham-controlled design. Histologic and immunohistochemical analysis on postnatal d 21 revealed that neonates developed severe cerebral damage after the hypoxic-ischemic insult. These animals also suffered from contralateral spastic paresis, as evidenced by their locomotor behavior. After transplantation of human umbilical cord blood-derived mononuclear cells, spastic paresis was largely alleviated, resulting in a normal walking behavior. This "therapeutic" effect was accompanied by the fact that mononuclear cells had entered the brain and were incorporated around the lesion without obvious signs of transdifferentiation. This study demonstrates that intraperitoneal transplantation of human umbilical cord blood-derived mononuclear cells in a rat model of perinatal brain damage leads to both incorporation of these cells in the lesioned brain area and to an alleviation of the neurologic effects of cerebral palsy as assessed by footprint and walking pattern analysis.
Electrical synapses can undergo activity-dependent plasticity. The calcium/calmodulin-dependent kinase II (CaMKII) appears to play a critical role in this phenomenon, but the underlying mechanisms of ...how CaMKII affects the neuronal gap junction protein connexin36 (Cx36) are unknown. Here we demonstrate effective binding of ³⁵S-labeled CaMKII to 2 juxtamembrane cytoplasmic domains of Cx36 and in vitro phosphorylation of this protein by the kinase. Both domains reveal striking similarities with segments of the regulatory subunit of CaMKII, which include the pseudosubstrate and pseudotarget sites of the kinase. Similar to the NR2B subunit of the NMDA receptor both Cx36 binding sites exhibit phosphorylation-dependent interaction and autonomous activation of CaMKII. CaMKII and Cx36 were shown to be significantly colocalized in the inferior olive, a brainstem nucleus highly enriched in electrical synapses, indicating physical proximity of these proteins. In analogy to the current notion of NR2B interaction with CaMKII, we propose a model that provides a mechanistic framework for CaMKII and Cx36 interaction at electrical synapses.
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