Properties of a nonjunctional current expressed from a rat connexin46 cDNA in Xenopus oocytes.

Ebihara, Lisa; Steiner, E.

doi:10.1085/jgp.102.1.59

Cited by 225 publications

(250 citation statements)

References 19 publications

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“…Hemichannels, unlike cell-cell channels, allow examination at the single channel level using techniques widely applied to other membrane channels and receptors and their study will likely routinely accompany studies of connexin structurefunction and regulation. For the most part, functional hemichannels have been observed using the Xenopus oocyte expression system (3,(14)(15)(16), but there are reports of functional hemichannels in mammalian cells transfected with connexins (17). Support for functional hemichannels in vivo comes from horizontal cells of the teleost retina, where GJs mediate the lateral inhibition of excitatory stimuli in the horizontal cell layer of the outer retina.…”

Section: Discussionmentioning

confidence: 99%

“…As expected for a large aqueous channel, the alkali cation permeability sequence was in order of their aqueous mobilities, but the permeabilities of the organic cations TMA and TEA were reduced more than expected, perhaps due to interactions with the channel wall. Anion permeabilities were low and their small contribution to E rev did not allow reliable discrimination among the anions Cl -, Br -, NO 3 -, and acetate -. Inward rectification and cation selectivity can be explained by the presence of fixed negative charges toward the extracellular end of the unapposed hemichannel (Trexler EB and Verselis VK, unpublished results).…”

Section: Conductance and Permeability Of Cx46 Hemichannels And Cell-cmentioning

confidence: 94%

“…Macroscopic currents obtained from Xenopus oocytes expressing rat connexin46 (Cx46) suggested that some connexins could function as hemichannels (3,4). We reported that recordings of single Cx46 hemichannels obtained by patching onto the surface membrane of Xenopus oocytes expressing Cx46 could be obtained with regularity (5).…”

Section: Introductionmentioning

confidence: 99%

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Connexin hemichannels and cell-cell channels: comparison of properties

Verselis

Trexler

Bukauskas

2000

Braz J Med Biol Res

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Connexin46 (Cx46) forms functional hemichannels in the absence of contact by an apposed hemichannel and we have used these hemichannels to study gating and permeation at the single channel level with high time resolution. Using both cell-attached and -excised patch configurations, we find that single Cx46 hemichannels exhibit some properties expected of half of a gap junction channel, as well as novel properties. Cx46 hemichannels have a large unitary conductance (~300 pS) and a relatively large pore as inferred from permeability to TEA. Both monovalent cations and anions can permeate, but cations are substantially more permeable. The open channel conductance shows marked inward rectification in symmetric salts. We find that the conductance and permeability properties of Cx46 cell-cell channels can be explained by the series addition of two hemichannels. These data suggest that the pore structures of unapposed hemichannels and cell-cell channels are conserved. Also like cell-cell channels, unapposed Cx46 hemichannels are closed by elevated levels of H + or Ca 2+ ions on the cytoplasmic face. Closure occurs in excised patches indicating that the actions of these agents do not require a soluble cytoplasmic factor. Fast (<0.5 ms) application of H + to either side of the open hemichannel causes an immediate small reduction in unitary conductance followed by complete closure with latencies that are dependent on H + concentration and side of application; sensitivity is much greater to H + on the cytoplasmic side. Closure by cytoplasmic H + does not require that the hemichannel be open. Thus, H + ions readily permeate Cx46 hemichannels, but at high enough concentration close them by acting at a cytoplasmic site(s) that causes a conformational change resulting in complete closure. Extracellular H + may permeate to act on the cytoplasmic site or act on a lower affinity extracellular site. Thus, the unapposed hemichannel is a valuable tool in addressing fundamental questions concerning the operation of gap junction channels that are difficult to answer by existing methods. The ability of Cx46, and perhaps other connexins, to form functional unapposed hemichannels that are opened by moderate depolarization may represent an unexplored role of connexins as mediators of transport across the plasma membrane.

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Section: Discussionmentioning

confidence: 99%

Section: Conductance and Permeability Of Cx46 Hemichannels And Cell-cmentioning

confidence: 94%

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Connexin hemichannels and cell-cell channels: comparison of properties

Verselis

Trexler

Bukauskas

2000

Braz J Med Biol Res

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“…Most connexin hemichannels are kept closed by normal extracellular Ca levels; hemichannel activity is enhanced when the Ca concentration drops below ∼0.5mM (Ebihara and Steiner, 1993;Ebihara, 1996;Beahm and Hall, 2002;Ebihara et al, 2003). Unfortunately, this can also enhance other permeability pathways in the plasma membrane, including P2X7 receptors.…”

Section: Dependence On Low Camentioning

confidence: 99%

Connexin channel permeability to cytoplasmic molecules

Harris¹

2007

Progress in Biophysics and Molecular Biology

414

346

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Connexin channels are known to be permeable to a variety of cytoplasmic molecules. The first observation of second messenger junctional permeability, made ∼30 years ago, sparked broad interest in gap junction channels as mediators of intercellular molecular signaling. Since then, much has been learned about the diversity of connexin channels with regard to isoform diversity, tissue and developmental distribution, modes of channel regulation, assembly and expression, biochemical modification and permeability, all of which appear to be dynamically regulated. This information has expanded the potential roles of connexin channels in development, physiology and disease, and made their elucidation much more complex -30 years ago such an orchestra of junctional dynamics was unanticipated. Only recently, however, have investigators been able to directly address, in this more complex framework, the key issue: What specific biological molecules, second messengers and others, are able to permeate the various types of connexin channels, and how well? An important related issue, given the ever-growing list of connexin-related pathologies, is how these permeabilities are altered by disease-causing connexin mutations. Together, many studies show that a variety of cytoplasmic molecules can permeate the different types of connexin channels. A few studies reveal differences in permeation by different molecules through a particular type of connexin channel, and differences in permeation by a particular molecule through different types of connexin channels. This article describes and evaluates the various methods used to obtain these data, presents an annotated compilation of the results, and discusses the findings in the context of what can be inferred about mechanism of selectivity and potential relevance to signaling. The data strongly suggest that highly specific interactions take place between connexin pores and specific biological molecular permeants, and that those interactions determine which cytoplasmic molecules can permeate and how well. At this time, the nature of those interactions is unclear. One hopes that with more detailed permeability and structural information, the specific molecular mechanisms of the selectivity can be elucidated.

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“…Thus, there was no change in the current flowing into the extracellular space, and the hemichannels were closed before opening to form a cell-cell channel. When expression of Cx46 in Xenopus oocytes was found to be fatal and to cause a large non-specific increase in membrane conductance in low Ca 2+ conditions, it seemed to be a rule that proved the exception: opening of (non-junctional) hemichannels is bad for cells, and something unusual about Cx46 expressed in oocytes allows hemichannel opening unless relatively high Ca 2+ concentrations are added to prevent it [8,9]. …”

mentioning

confidence: 99%

New roles for astrocytes: Gap junction hemichannels have something to communicate

Bennett

Contreras

Bukauskas

et al. 2003

Trends in Neurosciences

376

279

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Gap junctions are clusters of aqueous channels that connect the cytoplasm of adjoining cells. Each cell contributes a hemichannel, or connexon, to each cell-cell channel. The cell-cell channels are permeable to relatively large molecules, and it was thought that opening of hemichannels to the extracellular space would kill cells through loss of metabolites, collapse of ionic gradients and influx of Ca 2+ . Recent findings indicate that specific non-junctional hemichannels do open under both physiological and pathological conditions, and that opening is functional or deleterious depending on the situation. Most of these studies utilized cells in tissue culture that expressed a specific gap junction protein, connexin 43. Several such examples are reviewed here, with a particular focus on astrocytes.Gap junctions mediate intercellular communication by providing ultrastructural cytoplasmic continuity, and they are integral to formation of the functional syncytium of astrocytes. Each of the joined cells contributes a hemichannel or connexon to each cell-cell channel ( Figure 1; Box 1). Each hemichannel comprises a hexamer of connexins arranged around a central pore, and the cell-cell channels are gated by several stimuli, including transjunctional voltage, low pH and various pharmacological agents. Connexins are encoded by a gene family with at least 20 members in mammals [1].Gap-junction pores are nominally 1.0-1.5 nm in diameter and are permeable to molecules of 1 kDa [2]. Junctions formed from connexin 43 (Cx43), the predominant connexin in astrocytes, are permeable to Lucifer Yellow (443 Da, −2 charge) and propidium (420 Da, +2 charge). Other connexins appear to be more charge selective: Cx32 is more permeable to anions and Cx45 is more permeable to cations [2]. Single-channel conductance varies widely, from ~15 pS for Cx36 [3] and 110 pS for Cx43 [4] to ~375 pS for Cx37 [5]. The maximum size of permeant species also varies. Such diversity in selectivity and conductance are likely to account for the expansion of the connexin family. Why shouldn't hemichannels be open?Given the permeability and conductance of gap junctions, and the expectation that an open hemichannel would have twice the conductance and permeability of a cell-cell channel (in terms of ion or molecular flux, rather than selectivity), it was thought unlikely that hemichannels in the non-junctional membrane would open [6]. Direct evidence for this was provided by measurements during formation of the first channel between a newly apposed pair of cells. Each cell was held at a different potential, and then when the first channel NIH-PA Author ManuscriptNIH-PA Author Manuscript NIH-PA Author Manuscript opened, increased current was seen in the cell held at a more positive potential and decreased (more negative) current was seen in the other cell, but the sum of the currents clamping the two cells remained constant [7]. Thus, there was no change in the current flowing into the extracellular space, and the hemichannels were closed before openin...

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Properties of a nonjunctional current expressed from a rat connexin46 cDNA in Xenopus oocytes.

Cited by 225 publications

References 19 publications

Connexin hemichannels and cell-cell channels: comparison of properties

Connexin hemichannels and cell-cell channels: comparison of properties

Connexin channel permeability to cytoplasmic molecules

New roles for astrocytes: Gap junction hemichannels have something to communicate

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