Gap junction channel gating

AbstractOver the last two decades, the view of gap junction (GJ) channel gating has changed from one with GJs having a single transjunctional voltage-sensitive (Vj-sensitive) gating mechanism to one with each hemichannel of a formed GJ channel, as well as unapposed hemichannels, containing two, molecularly distinct gating mechanisms. These mechanisms are termed fast gating and slow or ‘loop’ gating. It appears that the fast gating mechanism is solely sensitive to Vj and induces fast gating transitions between the open state and a particular substate, termed the residual conductance state. 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 and GJ channel opening during de novo channel formation. A distinguishing feature of the slow gate is that the gating transitions appear to be slow, consisting of a series of transient substates en route to opening and closing. Published reports suggest that both sensorial and gating elements of the fast gating mechanism are formed by transmembrane and cytoplamic components of connexins among which the N terminus is most essential and which determines gating polarity. We propose that the gating element of the slow gating mechanism is located closer to the central region of the channel pore and serves as a ‘common’ gate linked to several sensing elements that are responsive to different factors and located in different regions of the channel

Heterotypic gap junction channels as voltage-sensitive valves for intercellular signaling

Gap junction (GJ) channels assembled from connexin (Cx) proteins provide a structural basis for direct electrical and metabolic cell–cell communication. By combining fluorescence imaging and dual whole-cell voltage clamp methods, we demonstrate that in response to transjunctional voltage (Vj) Cx43/Cx45 heterotypic GJs exhibit both Vj-gating and dye transfer asymmetries. The later is affected by ionophoresis of charged fluorescent dyes and voltage-dependent gating. We demonstrate that small differences in resting (holding) potentials of communicating cells can fully block (at relative negativity on Cx45 side) or enhance (at relative positivity on Cx45 side) dye transfer. Similarly, series of high frequency Vj pulses resembling bursts of action potentials (APs) can fully block or increase the transjunctional flux (Jj) of dye depending on whether pulses are generated in the cell expressing Cx43 or Cx45, respectively. Asymmetry of Jj-Vj dependence is enhanced or reduced when ionophoresis and Vj-gating act synergistically or antagonistically, whereas single channel permeability (Pγ) remains unaffected. This modulation of intercellular signaling by Vj can play a crucial role in many aspects of intercellular communication in the adult, in embryonic development, and in tissue regeneration

A Stochastic Four-State Model of Contingent Gating of Gap Junction Channels Containing Two “Fast” Gates Sensitive to Transjunctional Voltage

Connexins, a family of membrane proteins, form gap junction (GJ) channels that provide a direct pathway for electrical and metabolic signaling between cells. We developed a stochastic four-state model describing gating properties of homotypic and heterotypic GJ channels each composed of two hemichannels (connexons). GJ channel contain two “fast” gates (one per hemichannel) oriented opposite in respect to applied transjunctional voltage (Vj). The model uses a formal scheme of peace-linear aggregate and accounts for voltage distribution inside the pore of the channel depending on the state, unitary conductances and gating properties of each hemichannel. We assume that each hemichannel can be in the open state with conductance γh,o and in the residual state with conductance γh,res, and that both γh,o and γh,res rectifies. Gates can exhibit the same or different gating polarities. Gating of each hemichannel is determined by the fraction of Vj that falls across the hemichannel, and takes into account contingent gating when gating of one hemichannel depends on the state of apposed hemichannel. At the single-channel level, the model revealed the relationship between unitary conductances of hemichannels and GJ channels and how this relationship is affected by γh,o and γh,res rectification. Simulation of junctions containing up to several thousands of homotypic or heterotypic GJs has been used to reproduce experimentally measured macroscopic junctional current and Vj-dependent gating of GJs formed from different connexin isoforms. Vj-gating was simulated by imitating several frequently used experimental protocols: 1), consecutive Vj steps rising in amplitude, 2), slowly rising Vj ramps, and 3), series of Vj steps of high frequency. The model was used to predict Vj-gating of heterotypic GJs from characteristics of corresponding homotypic channels. The model allowed us to identify the parameters of Vj-gates under which small changes in the difference of holding potentials between cells forming heterotypic junctions effectively modulates cell-to-cell signaling from bidirectional to unidirectional. The proposed model can also be used to simulate gating properties of unapposed hemichannels

Application of Stochastic Automata Networks for Creation of Continuous Time Markov Chain Models of Voltage Gating of Gap Junction Channels

The primary goal of this work was to study advantages of numerical methods used for the creation of continuous time Markov chain models (CTMC) of voltage gating of gap junction (GJ) channels composed of connexin protein. This task was accomplished by describing gating of GJs using the formalism of the stochastic automata networks (SANs), which allowed for very efficient building and storing of infinitesimal generator of the CTMC that allowed to produce matrices of the models containing a distinct block structure. All of that allowed us to develop efficient numerical methods for a steady-state solution of CTMC models. This allowed us to accelerate CPU time, which is necessary to solve CTMC models, ∼20 times

Application of Stochastic Automata Networks for Creation of Continuous Time Markov Chain Models of Voltage Gating of Gap Junction Channels

The primary goal of this work was to study advantages of numerical methods used for the creation of continuous time Markov chain models (CTMC) of voltage gating of gap junction (GJ) channels composed of connexin protein. This task was accomplished by describing gating of GJs using the formalism of the stochastic automata networks (SANs), which allowed for very efficient building and storing of infinitesimal generator of the CTMC that allowed to produce matrices of the models containing a distinct block structure. All of that allowed us to develop efficient numerical methods for a steady-state solution of CTMC models. This allowed us to accelerate CPU time, which is necessary to solve CTMC models, ∼20 times