Biophysical properties of mouse connexin30 gap junction channels studied in transfected human HeLa cells.
1. Human HeLa cells expressing mouse connexin30 (Cx30) were used to study the electrical properties of Cx30 gap junction channels. Experiments were performed on cell pairs with the dual voltage-clamp method. 2. The gap junction conductance (gj) at steady state showed a bell-shaped dependence on junctional voltage (Vj; Boltzmann fit: Vj,0 = 27 mV, gj,min = 0.15, z = 4). The instantaneous gj decreased slightly with increasing Vj. 3. The gap junction currents (Ij) declined with time following a single exponential. The time constants of Ij inactivation (taui) decreased with increasing Vj. 4. Single channels exhibited a main state, a residual state and a closed state. The conductances gammaj,main and gammaj,residual were 179 and 48 pS, respectively (pipette solution, potassium aspartate; temperature, 36-37 degrees C; extrapolated to Vj = 0 mV). 5. The conductances gammaj,residual and gammaj,main showed a slight Vj dependence and were sensitive to temperature (Q10 values of 1.28 and 1.16, respectively). 6. Current transitions between open states (i.e. main state, substates, residual state) were fast (< 2 ms), while those between an open state and the closed state were slow (12 ms). 7. The open channel probability (Po) at steady state decreased from 1 to 0 with increasing Vj (Boltzmann fit: Vj,0 = 37 mV; z = 3). 8. Histograms of channel open times implied the presence of a single main state; histograms of channel closed times suggested the existence of two closed states (i.e. residual states). 9. We conclude that Cx30 channels are controlled by two types of gates, a fast one responsible for Vj gating involving transitions between open states (i.e. residual state, main state), and a slow one correlated with chemical gating involving transitions between the closed state and an open state.
Valiunas
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V
Department of Physiology, University of Bern, Buhlplatz 5, CH-3012 Bern, Switzerland.
Manthey
D
D
Vogel
R
R
Willecke
K
K
Weingart
R
R
D000818:Animals; D015152:Blotting, Northern; D000075302:Connexin 30; D017630:Connexins / Q000235:genetics / Q000502:physiology*; D004594:Electrophysiology; D017629:Gap Junctions / Q000502:physiology*; D006367:HeLa Cells; D006801:Humans; D051379:Mice; D009419:Nerve Tissue Proteins / Q000235:genetics / Q000502:physiology*; D014162:Transfection
D000075302:Connexin 30; D017630:Connexins; C491020:GJB6 protein, human; C100629:Gjb6 protein, mouse; D009419:Nerve Tissue Proteins