Heterogeneity and Remodeling of Ion Currents in Cultured Right Atrial Fibroblasts From Patients With Sinus Rhythm or Atrial Fibrillation

Figures

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  FIGURE 1. Overview of different ion current patterns in human atrial fibroblasts recorded using a ramp pulse protocol in whole-cell mode. (A) Exemplary traces of different current types (inward rectification, outward rectification, no rectification) with the ramp protocol used (top trace). (B) Distribution of current types between cells from different patients with SR or AF. Note that percentages may add up to >100% because some cells show both inward and outward rectification; the number of cells assessed for each patient is indicated below the x-axis. (C) Distribution of current types between cells from SR and AF patients (error bar = standard deviation between patients). (D) Pooled current/voltage (IV) curves of all cells from patients with SR (black) or AF (red): Both IV curves have a reversal potential of approximately –20 mV. No statistically significant differences in current density between cells from SR and AF patients were observed.

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  FIGURE 2. Currents profiles with fast kinetics identifiable with a step-pulse protocol (whole-cell mode). (A–C): Representative recording of three different current profiles measured during the first 30 ms after depolarization: rapidly activating and inactivating transient outward current (fast current, (A), more slowly activating and inactivating outward current [slow current, (B)] and inward current observed at holding potentials ranging from –20 to +60 mV (C). 1 s-long voltage clamp pulses were used to depolarize cells from –80 to +60 mV in steps of 20 mV; the holding potential used between pulses was –80 mV (top schematic). (D–F) Representative IV curves of peak current. In panel (F), the difference current IV curve was obtained by subtracting from the original IV curve the one of the fast current (D). (G) Distribution of current profiles within cells from three patients each with SR and AF. Note that percentages may add up to >100% because some cells presented two current activation patterns. (H) Comparison of the percentage of cells exhibiting fast and slow outward, and inward current in all cells analyzed from patients with SR and AF. No statistically significant differences in the…

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  FIGURE 3. Fast transient outward current measured with a step-pulse protocol in cells from tissue donors with SR and AF. (A) Pulse protocol and a representative current recording. Inset: Enlargement of early (first 10–15 ms) current dynamics, as indicated in A (stimulation artifact truncated in the enlarged image). Depolarizing steps activate a linearly increasing, transient current of <10 ms in duration. (B–D) IV curves constructed from peak current values (P), from values 10 ms after P (P′), and from values at the end of each voltage step (S, steady state), respectively, for fibroblasts from SR and AF donors. Please note different scales of Y-axes. *Indicates significant difference.

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  FIGURE 4. Large Ca2+-dependent conductance activated in the positive potential (outward current) range in human atrial fibroblasts. (A) Representative current trace measured with the ramp pulse protocol (whole-cell recording, ruptured patch mode). The inset shows the final 200 ms of the trace (outward current) at positive potentials; the current displays clear single channel openings. (B) Voltage-dependent current amplitude at different intracellular Ca2+ concentrations (from 0 to 0.1 mM), and after returning to nominally Ca2+-free solution (washout). Recordings obtained in inside-out configuration (I.O.). (C) Channel activity in an inside-out patch transiently superfused with paxilline-containing solution. Note that the paxilline-induced inhibition of channel activity was partially reversed during wash-out. (D) Single channel activity in cell-attached patches without (control) and with 3 μmol L− 1 paxilline in the pipette solution. (E) Single channel events during representative current traces at various holding potentials (cell-attached patch mode) in a cell from an SR and AF patient. (F) Percentage of cells in relation to all cells studied per patient (see total numbers below the…

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  FIGURE 5. Effects of BKCa channel blocker paxilline on proliferation rate and cell shape. (A) Proliferation assay in the presence (dashed lines) and absence (continuous lines) of the BKCa channel blocker paxilline (10 μmol L− 1) after 1 and 3 days in matched samples from five individual SR patients (triplicate measurements for each point). (B): Mean values for results from the proliferation assay. When normalized to the absorbance value of day 1, the mean increase in absorbance of cells incubated with paxilline was lower (1.6 ± 0.077) than for control conditions (1.8 ± 0.077). (C,D) Representative images of fibroblasts cultured for 2 days with and without paxilline. Scale bar = 20 μm. (E) Comparison of cell-shape (circularity) of cells incubated with DMSO (control) vs. 10 μmol L− 1 paxilline in DMSO.