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Gap Junctions: Cell-Cell Channels in Animals


* Corresponding Author: Venetian Institute of Molecular Medicine, University of Padova, via G. Orus 2, 35129 Padua, Italy. Email:

Gap junctions provide one of the most common forms of intercellular communication. The structures underlying these communicating cell junctions1 were soon resolved in membrane associated particles forming aggregates of six subunits.2 They are composed of membrane proteins that form a channel that is permeable to ions and small molecules, connecting the cytoplasm of acdjacent cells. Two unrelated protein families are involved in this function; connexins, which are found only in chordates, and pannexins, which are ubiquitous and present in both chordate and invertebrate genomes.3 In this chapter, structural and functional issues of gap junction channels are reviewed. Several types of pathologies associated to channel dysfunction, with an emphasis on deafness, are also examined.


Structure of Connexin Channels

Connexin-based gap junction channels are composed of two hemichannels, or connexons, each provided by one of two neighbouring cells. Two connexons join in the gap between the cells to form a gap junction channel that connects the cytoplasms of the two cells. A connexon is composed of six transmembrane proteins, called connexins (Cx), which form a multigene family. Connexons may be homomeric (composed of six identical connexin subunits) or heteromeric (composed of more than one species of connexins). The channel may be homotypic (if connexons are identical) or heterotypic (if the two connexons are different).4

At least 20 distinct human Cx isoforms have been cloned.5 Connexins are classified according to their molecular mass or grouped into α, β and γ subtypes, based on sequence similarities.6,7 They have highly conserved sequences and may have originated from a common ancestor. In the human genome, the majority of β connexin genes map to two gene clusters at either 1p34-p35 or 13q11-q12.8 Connexins have four α helical transmembrane domains (M1 to M4), intracellular N- and C-termini, two extracellular loops (E1 and E2), and a cytoplasmic loop (CL).9 E1 and E2 mediate the docking of the two hemichannels.7 The CL and the C-terminus constitute the least homologous regions across the connexin family, suggesting that many of the functional differences between connexins reside there.10

Electron cryomicroscopy of two-dimensional crystals11 permitted to derive a three-dimensional density map at 5.7 angstroms in-plane and 19.8 angstroms vertical resolution, and to identify the positions and tilt angles for the 24 alpha helices within each hemichannel. The four hydrophobic segments in connexin sequences were assigned to the alpha helices in the map based on biochemical and phylogenetic data. The final model specifies the coordinates of C-alpha atoms in the transmembrane domain.12 Figure 1 presents a model of the transmembrane segment of a single Cx26 connexon derived by the coordinates of the Cα model deposited in the Protein Data Bank (accession code 1TXH). The six connexin molecules are currently thought to be arranged so that M3 is the major pore-lining helix, together with M1.13,14 However, due to the low resolution of the density map, the estimate of the helices orientation around their principal axis is affected by a large error, possibly larger than 90°.

Figure 1. Atomic model of the transmembrane segment of a Cx26 connexon.

Figure 1

Atomic model of the transmembrane segment of a Cx26 connexon. A molecule of Lucifer Yellow, a fluorescent tracer commonly employed to assay cell-cell communication, is shown in the middle of the pore.

Oligomerization of connexins into hemichannels occurs in the ER–Golgi. Vesicles transport hemichannels to the plasma membrane shearing the general secretory pathway with other membrane proteins.15,16 Although each connexin exhibits a distinct tissue distribution, many cell types express more than one connexin isoform. The variations in the structural composition of gap junction channels allow for a greater versatility of their physiological properties, however not all connexins participate in the formation of hetero-connexons.17 Indeed, it has been suggested that an “assembly” signal allows connexin subunits to recognize each other, thus preventing the interaction of incompatible connexins.


Although gap junctions have been traditionally described as nonselective pores, various studies have revealed a high degree of selectivity among the different connexins.4 The unitary conductance (i.e., the conductance of a single gap junction channel) varies widely (25 pS for Cx45, 350 pS for Cx37), yet, the sequence of monovalent cation selectivity is the same (K>Na>Li>TEA) for different connexins.18 Gap junction channels are permeable to soluble molecules such as metabolites or second messengers (cAMP, Ca2+ and IP3).19-22

A classical method for evaluating connexin permeability is the intercellular transfer of membrane impermeant fluorescent molecules (e.g., Lucifer Yellow, see Fig. 1) delivered intracellularly to a single cell.23 By employing a wide array of molecular tracers, it has been demonstrated that the selective transfer of molecules through gap junction channels is not dictated by size alone but is affected also by other parameters such as charge or rigidity.18,24 Recently, this type of analysis has been extended to signalling molecules (Fig. 2).19,25,26

Figure 2. IP3 permeability measurement in pairs of transfected HeLa cells expressing hCx26.

Figure 2

IP3 permeability measurement in pairs of transfected HeLa cells expressing hCx26. A) Differential interference contrast (DIC) image merged with the fluorescence image of the same field illuminated at 500 nm showing a small cluster of HeLa cells, two of (more...)

The rate of permeation of metabolites through gap junctions has been shown to be conditioned by connexin composition. For instance, Cx32 is permeable to both cAMP and cGMP, whereas heteromeric connexons composed of Cx32 and Cx26 lose permeability to cAMP, but not to cGMP.27 IP3 has been shown, in transfected HeLa cells loaded with fura-2 to monitor [Ca2+]i, to permeate homotypic Cx32 gap junctions approximately four times more efficiently than Cx26 and 2.5 times more efficiently than Cx43 gap junctions.28 Indeed, it has been suggested that the most physiologically consequential difference between different connexin channels is their differing abilities to permit intercellular passage of molecules (such as second messengers) considerably larger than current carrying ions.29

Voltage Gating

Gap junction channels exhibit voltage gating, a property shared by several ion channels, whereby conductance is sensitive to voltage. Voltage gating depends primarily on transjunctional voltage Vj, i.e., the potential difference between the cytoplasm of the two adjacent cells (Fig. 3).30 Over the last two decades, the view has changed from one with gap junction channel having a single transjunctional voltage-sensitive (Vj-sensitive) gating mechanism to one with each hemichannel of a formed channel, as well as unapposed hemichannels, containing two, molecularly distinct gating mechanisms.30 In few connexin channels, changes in membrane potential (Vm) may activate an additional gate (Vm gate).31

Figure 3. Voltage dependence of gap junction conductance.

Figure 3

Voltage dependence of gap junction conductance. Human connexins 26 and 30 were expressed either through the bicistronic pIRES-EGFP expression vector, or as EYFP-tagged chimera in isolated HeLa cell pairs. A) top: voltage commands applied to one of two (more...)

The voltage sensitivity exhibited by gap junctions has the potential to uncouple communicating cells.32 In homotypic channels, the intercellular conductance, gj is an even function of Vj (i.e., it is symmetric about Vj = 0): the same reduction in gj, is obtained either by hyperpolarizing one cell of a connected pair, or by depolarizing the other cell (by the same amount). In some cases, gj does not decline to zero with increasing Vj, but reaches a plateau or residual conductance that varies depending on the Cx isoform, indicating that a wide range of voltage gating behaviours exists.30,33 Some channels exhibit fast Vj gating transitions (˜1 ms) to the residual state and slow Vj gate transitions (˜10 ms) to the fully closed state, which are mediated by molecularly distinct processes (see next section). It has recently been proposed that the passage of large molecules between cells, such as fluorescent tracer molecules and cAMP, may be controlled by transjunctional voltage differences that activate the connexin voltage gate while having little effect on the electrical coupling arising from the passage of small electrolytes.26

Analysis of connexin sequences failed to reveal a region similar to ion-channels' S4, thereby implying that the molecular mechanisms which mediate voltage-induced closure of gap junction channels must be different.9 A proline in the second transmembrane domain, which is conserved among all members of the connexin family, may play a central role in a conformational change that links the voltage sensor and the voltage gate of intercellular channels.34 However, published reports suggest that both sensorial and gating elements of the fast gating mechanism are formed by transmembrane and cytoplasmic components of connexins among which the N terminus is most essential and which determines gating polarity.30 Recent data are consistent with a gating model in which the voltage sensor is positioned in the N-terminus and its inward movement initiates Vj-gating.35

Chemical Gating

Transjunctional/transmembrane voltages are neither the sole nor the most important parameters capable of influencing the state of gap junction channels. Permeability can be altered also by specific changes in cytosolic ion composition, as well as by post-translational modifications. These alternative mechanisms are termed slow or ‘loop’ gating. A distinguishing feature of the slow gate is that the gating transitions consist of a series of transient substates en route to opening and closing.30 The slow gating mechanism is also sensitive to Vj, but there is evidence that this gate may mediate gating by transmembrane voltage, Vm, intracellular Ca2+ and pH, chemical uncouplers (Fig. 4) and gap junction channel opening during de novo channel formation.30 At the single channel level, the chemical/slow gate closes the channels slowly and completely, whereas the fast Vj gate closes the channels rapidly and incompletely.36 Chemical agents that reduce coupling usually do not leave a residual conductance and their effect is readily reversible.

Figure 4. Example of chemical gating.

Figure 4

Example of chemical gating. A) Confocal section through the organ of Corti of the guinea pig, labelled with an antibody against connexin 26 (Cx26); scale bar, 30 μm. B) Schematics of dual patch clamp recordings. C) effect of gap junction inhibitors, (more...)

Chemical gating of gap junction channels is a complex phenomenon that may involve intra- and intermolecular interactions among connexin domains and a cytosolic molecule (possibly calmodulin) that may function as channel plug.37 Some evidence suggests that low pHi affects gating via an increase in [Ca2+]i; in turn, Ca2+ is likely to induce gating by activation of CaM, which may act directly as a gating particle. The effective concentrations of both Ca2+ and H+ vary depending on cell type, type of connexin expressed and procedure employed to increase their cytosolic concentrations; however, pHi as high as 7.2 and [Ca2+]i as low as 150 nM or lower have been reported to be effective in some cells. At least three molecular models of channel gating have been proposed, but all of them are mostly based on circumstantial evidence.31

Regulation of Cell-to-Cell Communication

The number and distribution of gap junction channels is generally relatively stable under physiological conditions. However, the flux of connexins into and out of gap junctions is frequently reported to be highly dynamic.16 New channels are added to the outside of the junctional plaque while older channels are internalized from the centre to be degraded.16,38 The regulation of gap junction trafficking, assembly/disassembly and degradation is likely to be critical in the control of intercellular communication and phosphorylation has been implicated in the regulation of the connexin “lifecycle” at several stages.39 The C-terminal region of connexins appears to be the primary region that becomes phosphorylated on serine or threonine residues.39,40 Cx26 is the only connexin that has been reported not to be phosphorylated.41 This may be due to the fact that it is the shortest connexin and only has a few C-terminal tail amino acids (a.a.) that could interact with cytoplasmic signalling elements. Clearly, connexin phosphorylation is not required for the formation of all gap junction channels, as recombinant homomeric-homotypic channels formed by wild-type Cx26 are functional.42

Intercellular Calcium Waves

Connexin channels are permeable to some of the more important secondary messengers involved in cell signalling, such as cAMP,20 and IP3,21 the first and the principal inositol phosphate that is formed from inositol lipid hydrolysis due to G protein-linked receptor stimulation of phospholipase C.43 IP3 molecules diffuse rapidly throughout the cell,44 interact with specific receptors (IP3R) present in the endoplasmic reticulum and Ca2+ is liberated, raising its concentration ([Ca2+]i) in the cytosol.45,46 In some cells, these [Ca2+]i signals are targeted to control processes in limited cytoplasmic domains, but in other systems long-range signalling involves intercellular Ca2+ waves.47 Two pathways have been implicated so far in intercellular Ca2+ signalling (Fig. 5): (i) the extracellular diffusion of molecules48,49 such as adenosine nucleotides, glutamate and other neurotransmitters which, by interacting with receptors on nearby cells alter their intracellular calcium levels50 and (ii) the diffusion of cytoplasmic messenger molecules through gap junction.51

Figure 5. Mechanisms underlying the propagation of intercellular Ca2+ waves.

Figure 5

Mechanisms underlying the propagation of intercellular Ca2+ waves. Waves can be mediated by the intracellular transfer of signalling molecules (red arrows), such as IP3, trough gap junction channels (in green). The extracellular pathway involves the diffusion (more...)

Models of Ca2+ waves that involve the production of IP3 are based on the diffusion of IP3 from a stimulated cell through gap junction channels into neighboring cells where it elicits Ca2+ release from intracellular stores.52,53 However, more complex schemes have been proposed to account for the long-range periodic Ca2+ waves in the liver, where a messenger diffusing through gap junction channels appears to be regenerated in each participating cell.54

Intercellular propagation of Ca2+ waves has been described in a wide variety of cell types and is considered a mechanism by which cell activity is coordinated.55 In the central nervous system, such waves occur among and between neurons and glial cells both under normal and pathological conditions.56 In summary, direct propagation of intercellular Ca2+ waves from the cytosol of one cell to an adjacent cell requires the presence of gap junction channels, which allow signaling molecules, such as IP3,21,57 to be transferred across coupled cells. The alternative, but not mutually exclusive, pathway for communication of the Ca2+ signals involves the diffusion of signaling molecules, such as ATP, through the extracellular space, activating P2 receptors in neighboring cells that may or may not be in contact.48,49

Connexin-Related Pathologies

Recent evidence has shown that mutations within several connexins give rise to various hereditary diseases in humans such as Charcot–Marie–Tooth disease and erythrokeratodermia variabilis. In addition to these disorders, connexin mutations are also involved in deafness, cataracts, and numerous skin diseases. The association of connexin mutations with an increasing number of human pathologies, provides an unequivocal demonstration that gap junctional communication is crucial for diverse physiological processes.

Skin Diseases

In human skin, intercellular communication is mediated by all known β-connexin genes. Autosomal dominant mutations in connexins genes have been linked with several skin disorders which involve an increased thickness of the skin outer layers. This would indicate a critical role for connexins in maintaining the balance between proliferation and differentiation of the epidermis.58,59GJB3 was the first connexin gene reported to cause the autosomal dominant skin disorder erythrokeratodermia variabilis (EKV) and several mutations have been identified so far in its protein product, Cx31: G12R, G12D, C86S, R42P and F137L.60 Dominant mutations in Cx26 have also been described in syndromic deafness associated with skin disease. These disorders include keratitis-ichthyosis-deafness syndrome (KID), palmoplantar keratoderma with deafness (PPK), Vohwinkel syndrome (VS) and hystrix-like ichthyosis-deafness syndrome (HID).61,62 Dominant mutations in GJB2 affect a.a residues positioned in the highly conserved first extracellular domain (G59A and D66H), or at the boundary of this domain with the first (DelE42) or second (R75W) transmembrane domain. The exact role of Cx26 in skin disease, however, can not be attributed to loss of Cx26 channel activity alone due to the fact that many Cx26 mutations result in nonsyndromic deafness (see below). This would indicate that mutant forms of Cx26 associated with skin disorders must impact additional genes in a manner that upsets tissue homeostasis and results in disease.

Different missense mutations in Cx30, G11R, V37E and A88V, affecting conserved a.a. residues positioned in the N-terminal end and in the second transmembrane domains, have been identified as cause of ectodermal dysplasia (HED)63 or Clouston syndrome. These mutations impair trafficking of the protein to the cell membrane and lead to cytoplasmic accumulation when transiently expressed in cultured keratinocytes, thus resulting in a complete loss of gap junction function.64

Peripheral Neuropathy and Congenital Cataract

The first disease associated to mutations in connexin genes was a form of Charcot-Marie-Tooth disease (CMT). Dominant and recessive X-linked mutations in the Cx32 sequence result in progressive degeneration of peripheral nerve and are characterized by distal muscle weakness and atrophy.65 Cx32 is present in Schmidt–Lanterman incisures and nodes of Ranvier of myelinating Schwann cells,66 providing continuity between the Schwann cell body and the cytoplasmic collar of the myelin sheath adjacent to the axon. It has been suggested that the S26L mutation of Cx32, implicated in X-linked CMT, alters the permeability of the gap junction to cyclic nucleotides.67 At least another connexin, possibly Cx31, participates in forming gap junctions in these cells.68 Cx31 was reported to be expressed in mouse auditory and sciatic nerves in a pattern similar to that of mouse Cx32.69

In the ocular lens, gap junctional communication is a key component of homeostatic mechanisms preventing cataract formation.70 Altered biochemical coupling, resulting from inappropriate mixing of Cx46 and Cx50, perturbed cellular homeostasis but not lens growth, suggesting that unique biochemical modes of gap junctional communication influence lens clarity and lens growth, and that biochemical coupling is modulated by the connexin composition of the gap junction channels.71,72


In developed countries deafness has an important genetic origin and at least 60% of the cases are inherited. In particular, the early onset forms of hearing impairment in developed countries are almost exclusively genetic in origin and not due to undiagnosed infections. Thirty-seven genes responsible for isolated hearing impairment in humans are known to date. Mutations in one of them, underlying the DFNB1 form of deafness, have been found to be responsible for about half of all cases of human deafness in countries surrounding the Mediterranean Sea. DFNB1 is thus almost as frequent as cystic fibrosis. DFNB1 can be caused by mutations in the GJB2 gene (121011), which encodes the gap junction protein Cx26. However the complex DFNB1 locus (13q11-q12) has been shown to contain another gap junction gene GJB6 (604418, which maps to 13q12), and encodes Cx30.73 Specifically, DFNB1 can also be due to a deletion of 342 Kb involving GJB6 (Ballana E, Ventayol M, Rabionet R, Gasparini P, Estivill X. Connexins and deafness Homepage. World wide web URL: and which has been related to a dominant type of deafness in an Italian family.74,75

In the cochlea, gap junctions are found in two networks of cells: the epithelial network of supporting cells and the connective tissue network of fibrocytes and cells of the stria vascularis.76,77 Cx26 is the most prominent connexin expressed in the inner ear, although it is also expressed in various other organs, including the proximal tubules of the kidney, the liver and the rat placenta.8 Cx26 colocalizes with Cx30 between supporting cells, in the spiral limbus, the spiral ligament and the stria vascularis, where they may form heteromeric connexons.78,79 Cx30 is found also in adult mouse brain and skin,80 and its a.a. sequence shares 77% identity with that of Cx26.81

More than 50 distinct recessive mutations of GJB2 have been described, including nonsense, missense, splicing, frame-shift mutations and in-frame deletions.82 A large French dominant family affected by prelingual deafness showed linkage to chromosome 13q12 (DFNA3), the same region to which DFNB1 had been mapped.83 An interaction between GJB2 mutations and a mitochondrial mutation appears to be the cause of hearing impairment in some heterozygote patients.84

In addition to Cx26 and Cx30, several other connexins are known to be expressed in the inner ear. Cx31 (GJB3, 1p32-p36) localize to the inner ear connective tissue. It is expressed at P12 and reaches the adult pattern at P60.85 Moreover, measurable mRNA levels have been detected in the stria vascularis for Cx43, Cx37, Cx30.2 and Cx46,86 although is not clear if these connexins are expressed in the cochlea. Cx43 is expressed in the connective tissues only during development. However, from P8, Cx43 is almost exclusively expressed in the bone of the otic capsule.87

Not surprisingly, other connexin genes are also involved in deafness: GJB1 (Cx32), which is also responsible for X-linked Charcot-Marie-Tooth disease type I;88 GJB3 (Cx31), involved in deafness89 or a skin disease60 (erythrokeratodermia variabilis) depending on the location of the mutation; GJA1 (Cx43), involved in recessive deafness.90

Targeted Ablation of Inner Ear Connexins

Gene targeting of connexins in mice has provided new insights into connexin function and the significance of connexin diversity.65 Complete removal of the Cx26 gene results in neonatal lethality, thereby preventing analysis of its function in hearing.91 This limitation was overcome through the generation of a cochlear specific knockout of Cx26 using the Cre–loxP system,92 successfully deleting Cx26 in the epithelial gap junction network without effecting Cx26 expression in other organs. Animals with this deletion are a model of recessive deafness and displayed normal patterns of cochlear development, but showed an increase in postnatal cell death within the cochlea along with significant hearing loss. The initiation of cell death was found to occur in supporting cells proximal to the inner hair cells, and coincided with the onset of audition. It was hypothesized that loss of Cx26 prevented recycling of K+ after sound stimulation, and that elevated K+ in the extracellular perilymph inhibited uptake of the neurotransmitter glutamate, which accumulated and resulted in cell death.92 While the evidence concerning Cx26 is compelling, this hypothesis becomes more complex when considering the activity of Cx30 in the inner ear. Lautermann et al93 have shown that Cx26 and Cx30 normally colocalize within the cochlea, and tissue-specific deletion of the Cx26 gene did not alter expression patterns of Cx30.92 This observation complicates the role of gap junctional dysfunction in the onset of deafness due to the fact that Cx30 passes K+ in an efficient manner94 but could not prevent hearing loss in the absence of Cx26. Thus, the presence of a single type of connexin with similar ionic selectivity was unable to rescue the phenotype observed in these mice.

Similar to the tissue-specific loss of Cx26, deletion of Cx30 in mice resulted in hearing loss, but did not alter the development of the inner ear.95 Although development was normal, Cx30 knockouts lacked the endocochlear potential, but maintained the [K+] of the endoplymph. In addition, Cx30 knockouts also presented increased apoptosis within the cochlear sensory epithelium. Deletion of Cx30 did not alter the cochlear expression of Cx26, again raising the question of why the continued expression of Cx26 was not able to compensate for the loss of Cx30.95

One possible explanation for these phenotypes is that gap junctions may have other roles in addition to recycling K+. This idea is supported by data from functional studies showing that, while the K+ conductances of Cx26 and Cx30 are similar, the channels display significant differences regarding the permeability to fluorescent dyes that are similar to cyclic nucleotides and second messengers in both size and charge. These findings indicate that specific loss of either Cx26 or Cx30 within cochlear epithelial cells would not simply reduce the intercellular passage of K+, but would also significantly alter the availability of larger solutes that could be exchanged between the coupled cells.59

Connexin Permeability Defects and Genetic Deafness

Connexin permeability defects may produce a deafness phenotype by interfering with potassium spatial buffering by cochlear supporting cells,96 which most probably requires a coordinated activity. The identification in DFNB1 patients of a recessive Cx26 mutation, V84L, that did not appreciably affect basic channel properties, has recently provided some insight into the molecular mechanisms that underlie the disease. Using state-of-the-art techniques we demonstrated that V84L channels exhibit an impaired permeability to IP3. The apparent permeability coefficient of V84L for IP3 is only 8% of that of wild-type Cx26. Thus mutated channels must have a structural modification that reduces the passage of IP3, without compromising that of other ions. Our report25 is the first direct demonstration that a connexin mutation results selectively in a defective transfer of a second-messenger molecule. Modification of gap-junction permeability has been suggested, on the basis of indirect evidence, to be at the basis of some pathological connexin mutation, e.g., the S26L mutation of Cx32,67 but no direct proof that this may be the case had yet been offered. In the organ of Corti, gap junction blockade impairs the spreading of Ca2+ waves (Fig. 6) and the formation of a functional syncytium. Wave propagation necessitates also a regenerative mechanism mediated by P2Y receptors97 and this may represent a fundamental mechanism by which cochlear supporting cells coordinate their responses to sound.

Figure 6. IP3 injection elicits intercellular Ca2+ waves in supporting cells of the organ of Corti.

Figure 6

IP3 injection elicits intercellular Ca2+ waves in supporting cells of the organ of Corti. A) Cell 1 was contacted by a patch pipette (outlined) loaded with 500 μM IP3. B) Fura-2 ratio changes, δR, from the corresponding regions of interest (more...)


Database search has led to the identification of a family of proteins, the pannexins, which share some structural features with the gap junction forming proteins of invertebrates and vertebrates.98 The pannexin genes PANX1, PANX2 and PANX3, encoding putative gap junction proteins homologous to invertebrate innexins, constitute a new family of mammalian proteins. In the brain, the pannexins may represent a novel class of electrical synapses.

Phylogenetic analysis revealed that pannexins are highly conserved in worms, molluscs, insects and mammals, pointing to their important function. Both innexins and pannexins are predicted to have four transmembrane regions, two extracellular loops, one intracellular loop and intracellular N and C termini. Both the human and mouse genomes contain three pannexin-encoding genes. Mammalian pannexins PANX1 and PANX3 are closely related, with PANX2 more distant. The human and mouse PANX1 mRNAs are ubiquitously, although disproportionately, expressed in normal tissues. Human PANX2 is a brain-specific gene; its mouse orthologue, Panx2, is also expressed in certain cell types in developing brain. In silico evaluation of Panx3 expression predicts gene expression in osteoblasts and synovial fibroblasts.99

On expression in Xenopus oocytes, pannexin 1 (Px1), but not Px2 forms functional hemichannels. Coinjection of both pannexin RNAs results in hemichannels with functional properties that are different from those formed by Px1 only. In paired oocytes, Px1, alone and in combination with Px2, induces the formation of intercellular channels. The functional characteristics of homomeric Px1 versus heteromeric Px1/Px2 channels and the different expression patterns of Px1 and Px2 in the brain indicate that pannexins form cell type-specific gap junctions with distinct properties that may subserve different functions.98 There is evidence for the interaction of Px1 with Px2 and that the pharmacological sensitivity of heteromeric Px1/Px2 is similar to that of homomeric Px1 channels. In contrast to most connexins, both Px1 and Px1/Px2 hemichannels were not gated by external Ca2+. In addition, they exhibited a remarkable sensitivity to blockade by carbenoxolone (with an IC50 of approximately 5 μM), whereas flufenamic acid exerted only a modest inhibitory effect, thus indicating that gap junction blockers are able to selectively modulate pannexin and connexin channels. 100 In the light of these findings, it is now necessary to consider pannexins as an alternative to connexins in vertebrate intercellular communication.3

Intercellular calcium wave propagation initiated by mechanical stress is a phenomenon found in nearly all cell types. However, the conduit for ATP has remained elusive and both a vesicular and a channel mediated release have been considered. Px1 channels are of large conductance and have been shown to be (a) permeant for ATP, and (b) mechanosensitive, suggesting that pannexins are candidates for the release of ATP to the extracellular space upon mechanical stress.101


Supported by grants from the Telethon Foundation, (Project n. GGP02043), Ministero dell'Università e Ricerca Scientifica (MIUR, FIRB n. RBAU01Z2Z8, PRIN-COFIN 2002067312_002) to F.M., Centro di Eccellenza (coordinator, Tullio Pozzan) and the Italian Health Ministry. I thank all present and past graduate students and post-doctoral fellows, directly or indirectly involved in work on the connexins, and in particular: Martina Beltramello, Valeria Piazza, Catalin D. Ciubotaru, Mario M. Bortolozzi, Victor H. Hernandez, Andrea Lelli, Sergio Pantano and Stefano Bastianello (Venetian Institute of Molecular Medicine, Padova, Italy). I also thank: Feliksas F. Bukauskas, Vytas Verselis, Michael V. L. Bennett (Department of Neuroscience, Albert Einstein College of Medicine, Bronx, New York), Paola D'Andrea (University of Trieste, Italy) and, last but not least, Roberto Bruzzone (Institut Pasteur, Paris, France) for helpful discussions.


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