Pannexin 1: The Molecular Substrate of Astrocyte "Hemichannels"

The Dominick P. Purpura Department of Neuroscience, Albert Einstein College of Medicine, Bronx, New York 10461, USA.
The Journal of Neuroscience : The Official Journal of the Society for Neuroscience (Impact Factor: 6.34). 06/2009; 29(21):7092-7. DOI: 10.1523/JNEUROSCI.6062-08.2009
Source: PubMed


Purinergic signaling plays distinct and important roles in the CNS, including the transmission of calcium signals between astrocytes. Gap junction hemichannels are among the mechanisms proposed by which astrocytes might release ATP; however, whether the gap junction protein connexin43 (Cx43) forms these "hemichannels" remains controversial. Recently, a new group of proteins, the pannexins, have been shown to form nonselective, high-conductance plasmalemmal channels permeable to ATP, thereby offering an alternative for the hemichannel protein. Here, we provide strong evidence that, in cultured astrocytes, pannexin1 (Panx1) but not Cx43 forms hemichannels. Electrophysiological and fluorescence microscope recordings performed in wild-type and Cx43-null astrocytes did not reveal any differences in hemichannel activity, which was mostly eliminated by treating Cx43-null astrocytes with Panx1-short interfering RNA [Panx1-knockdown (Panx1-KD)]. Moreover, quantification of the amount of ATP released from wild-type, Cx43-null, and Panx1-KD astrocytes indicates that downregulation of Panx1, but not of Cx43, prevented ATP release from these cells.

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Available from: Eliana Scemes
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    • "Cultured astrocytes from the offspring of LPS-exposed dams exhibited a 3-fold increase in Etd uptake compared with those under control conditions (Fig. 1A–E). Astrocytes express functional hemichannels formed by Cx43 (Contreras et al., 2002) and Panx1 channels (Iglesias et al., 2009). The possible role of Cx43 hemichannels in prenatal LPS exposure-induced Etd uptake was studied using mimetic peptides with sequences homologous to the first extracellular (Gap26) or intracellular (Gap 19 and TAT-L2) loop domains of Cx43 (Wang et al., 2013a). "
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    ABSTRACT: Several epidemiological studies indicate that children born from mothers exposed to infections during gestation, have an increased risk to develop neurological disorders, including schizophrenia, autism and cerebral palsy. Given that it is unknown if astrocytes and their crosstalk with neurons participate in the above mentioned brain pathologies, the aim of this work was to address if astroglial paracrine signaling mediated by Cx43 and Panx1 unopposed channels could be affected in the offspring of LPS-exposed dams during pregnancy. Ethidium uptake experiments showed that prenatal LPS-exposure increases the activity of astroglial Cx43 and Panx1 unopposed channels in the offspring. Induction of unopposed channel opening by prenatal LPS exposure depended on intracellular Ca(2+) levels, cytokine production and activation of p38 MAP kinase/iNOS pathway. Biochemical assays and Fura-2AM/DAF-FM time-lapse fluorescence images revealed that astrocytes from the offspring of LPS-exposed dams displayed increased spontaneous Ca(2+) dynamics and NO production, whereas iNOS levels and release of IL-1β/TNF-α were also increased. Interestingly, we found that prenatal LPS exposure enhanced the release of ATP through astroglial Cx43 and Panx1 unopposed channels in the offspring, resulting in an increased neuronal death mediated by the activation of neuronal P2X7 receptors and Panx1 channels. Altogether, this evidence suggests that astroglial Cx43 and Panx1 unopposed channel opening induced by prenatal LPS exposure depended on the inflammatory activation profile and the activation pattern of astrocytes. The understanding of the mechanism underlying astrocyte-neuron crosstalk could contribute to the development of new strategies to ameliorate the brain abnormalities induced in the offspring by prenatal inflammation. GLIA 2015. © 2015 Wiley Periodicals, Inc.
    Full-text · Article · Jun 2015 · Glia
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    • "Recently, pannexin-1 was co-immunoprecipitated (IP) with P2X 7 R as part of P2X 7 R complex (Pelegrin and Surprenant, 2006; Silverman et al., 2009). The P2X 7 R-triggered ATP release was demonstrated to mediate through pannexin-1 hemichannels in T-cells (Schenk et al., 2008) and in astrocytes (Iglesias et al., 2009). Thus, activation of P2X 7 R may trigger D-serine release through pannexin-1 hemichannel of the P2X 7 R-pannexin-1 complex. "
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    ABSTRACT: d-serine is a coagonist of N-methyl-d-aspartate (NMDA) subtype of glutamate receptor and plays a role in regulating activity-dependent synaptic plasticity. In this study, we examined the mechanism by which extracellular ATP triggers the release of d-serine from astrocytes and discovered a novel Ca2+-independent release mechanism mediated by P2X7 receptors (P2X7R). Using [3H] d-serine, which was loaded into astrocytes via the neutral amino acid transporter 2 (ASCT2), we observed that ATP and a potent P2X7R agonist, 2′(3′)-O-(4-benzoylbenzoyl)adenosine-5′-triphosphate (BzATP), stimulated [3H]D-serine release and that were abolished by P2X7R selective antagonists and by shRNAs, whereas enhanced by removal of intracellular or extracellular Ca2+. The P2X7R-mediated d-serine release was inhibited by pannexin-1 antagonists, such as carbenoxolone (CBX), probenecid (PBN), and 10Panx-1 peptide, and shRNAs, and stimulation of P2X7R induced P2X7R-pannexin-1 complex formation. Simply incubating astrocytes in Ca2+/Mg2+-free buffer also induced the complex formation, and that enhanced basal d-serine release through pannexin-1. The P2X7R-mediated d-serine release assayed in Ca2+/Mg2+-free buffer was enhanced as well, and that was inhibited by CBX. Treating astrocytes with general protein kinase C (PKC) inhibitors, such as chelerythrine, GF109203X, and staurosporine, but not Ca2+-dependent PKC inhibitor, Gö6976, inhibited the P2X7R-mediated d-serine release. Thus, we conclude that in astrocytes, P2X7R-pannexin-1 complex formation is crucial for P2X7R-mediated d-serine release through pannexin-1 hemichannel. The release is Ca2+-independent and regulates by a Ca2+-independent PKC. The activated P2X7R per se is also functioned as a permeation channel to release d-serine in part. This P2X7R-mediated d-serine release represents an important mechanism for activity-dependent neuron-glia interaction. GLIA 2015
    Full-text · Article · Jan 2015 · Glia
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    • "Channels formed by connexin (Cx) or pannexin proteins (connexon and pannexon, respectively) were shown to impact cellular properties and underlie various pathological processes by serving as conduits for ions and various autocrine and paracrine signaling molecules (Contreras et al., 2002; Bennett et al., 2003; Scemes et al., 2007; Iglesias et al., 2009; Figure 1A). Some of these channels can assemble into intercellular structures. "
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    ABSTRACT: Unapposed hemichannels (HCs) formed by hexamers of gap junction proteins are now known to be involved in various cellular processes under both physiological and pathological conditions. On the other hand, less is known regarding how differences in the molecular composition of HCs impact electrical synaptic transmission between neurons when they form intercellular heterotypic gap junctions (GJs). Here we review data indicating that molecular differences between apposed HCs at electrical synapses are generally associated with rectification of electrical transmission. Furthermore, this association has been observed at both innexin and connexin (Cx) based electrical synapses. We discuss the possible molecular mechanisms underlying electrical rectification, as well as the potential contribution of intracellular soluble factors to this phenomenon. We conclude that asymmetries in molecular composition and sensitivity to cellular factors of each contributing hemichannel can profoundly influence the transmission of electrical signals, endowing electrical synapses with more complex functional properties.
    Full-text · Article · Oct 2014 · Frontiers in Molecular Neuroscience
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