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. 2015 Sep;63(9):1646-59.
doi: 10.1002/glia.22834. Epub 2015 Apr 9.

Voltage-dependent K+ Currents Contribute to Heterogeneity of Olfactory Ensheathing Cells

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Free PMC article

Voltage-dependent K+ Currents Contribute to Heterogeneity of Olfactory Ensheathing Cells

Lorena Rela et al. Glia. .
Free PMC article

Abstract

The olfactory nerve is permissive for axon growth throughout life. This has been attributed in part to the olfactory ensheathing glial cells that encompass the olfactory sensory neuron fascicles. Olfactory ensheathing cells (OECs) also promote axon growth in vitro and when transplanted in vivo to sites of injury. The mechanisms involved remain largely unidentified owing in part to the limited knowledge of the physiological properties of ensheathing cells. Glial cells rely for many functions on the properties of the potassium channels expressed; however, those expressed in ensheathing cells are unknown. Here we show that OECs express voltage-dependent potassium currents compatible with inward rectifier (Kir ) and delayed rectifier (KDR ) channels. Together with gap junction coupling, these contribute to the heterogeneity of membrane properties observed in OECs. The relevance of K(+) currents expressed by ensheathing cells is discussed in relation to plasticity of the olfactory nerve.

Keywords: ensheathing glia; gap junctions; potassium conductance.

Figures

Fig. 1
Fig. 1
Diversity of OEC membrane current profiles. A1: Whole cell currents (left) of OECs subjected to a series of voltage steps (−140 to 80 mV, Δ20 mV). Examples of cells with high (top), intermediate (middle) and low (bottom) Ri values. I/V curves are shown at the right. Inset: expanded scale (tick marks: 1 nA, 50 mV). A2: Derivatives of the I/V curves (dI/dVm) shown in A1. The top graph indicates maxima (Gmaxi, Gmaxo) and minima (Gmin) of interest. The vertical line separates the range of voltages eliciting inward (left) or outward (right) currents. B: 3D representation of Gmaxi, Gmin and Gmaxo. Each symbol represents a different OEC and 2D projections are in gray symbols. Vr is color coded (n=56). C: Gmaxi, Gmin and Gmaxo for the same OEC sample shown in B. Values corresponding to the same OEC are connected with lines. # p<0.05, compared to Gmin (Friedman test, n=56 cells).
Fig. 2
Fig. 2
Ba2+ blocks a hyperpolarization-activated inward current of OECs. A1: Whole cell currents (left) of an OEC subjected to a series of voltage steps (−140 to 80 mV, Δ20 mV) in control (top) and with 100 µM Ba2+ (bottom). I/V curves are shown at the right. A2: Conductance as a function of Vm for the curves shown in A1. The bottom graph shows the control conductance (gray) overimposed to the conductance in the presence of Ba2+ (black). B: Ba2+-sensitive component for the OEC shown in A1, with the I/V curve at the right. Insets: expanded scale (scale bars: 0.5 nA, 200 ms; tick marks: 0.25 nA, 50 mV). C: Average I/V curve of the Ba2+-sensitive current, normalized to the maximum inward current in control conditions (left, n=8) and average percent inhibition of the total current as a function of Vm (right, n=8).
Fig. 3
Fig. 3
TEA blocks depolarization-activated outward currents of OECs. A1: Representative whole cell currents (left) of an OEC subjected to a series of voltage steps (−140 to 80 mV, Δ20 mV) in control (top) and with 20 mM TEA (bottom). I/V curves are shown at the right. A2: Derivatives of the I/V curves shown in A1. The bottom graph shows the control derivative (gray) overimposed to the derivative in the presence of TEA (black). B: TEA-sensitive component of the OEC shown in A1, with the I/V curve at the right. Inset: expanded scale for better appreciation of outward rectification (scale bars: 0.2 nA, 200 ms; tick marks: 0.25 nA, 50 mV). C: Average I/V curve of the TEA-sensitive current, normalized to the maximum outward current in control conditions (left, n=10) and average percent inhibition of the total current as a function of Vm (right, n=10).
Fig. 4
Fig. 4
MFA blocks ohmic currents of OECs. A1: Representative whole cell currents (left) of an OEC subjected to a series of voltage steps (−140 to 80 mV, Δ20 mV) in control (top) and in the presence of 100 µM MFA (bottom). I/V curves are shown at the right. A2: Derivatives of the I/V curves shown in A1. The bottom graph shows the control derivative (gray) overimposed to the derivative in the presence of MFA (black). B: MFA-sensitive component of the OEC shown in A1, with the I/V curve at the right. C: Average I/V curve of the TEA-sensitive current, normalized to the maximum outward current in control conditions (left, n=12) and average percent inhibition of the total current as a function of Vm (right, n=12).
Fig. 5
Fig. 5
Effects of channel blockers on conductance and linearity of the I/V curve. The graphs show Gmaxi, Gmin and Gmaxo (left) or Vr (right) before (white symbols) and after (black symbols) treatment with 100 µM Ba2+ (A), 20 mM TEA (B) or 100 µM MFA (C). Each symbol corresponds to a different OEC. Statistical comparisons: Bonferroni posttests in a 2-way repeated measures-ANOVA with Treatment and Conductance of interest (COI) as factors. A: * p<0.05, Ba2+ vs. control; # p<0.05, ## p<0.001, Gmax vs. Gmin; (effect of COI: p<0.001; interaction: p<0.001, n=8). B: * p<0.01, TEA vs. control; # p<0.05, ## p<0.001, Gmax vs. Gmin; (effect of COI: p<0.001; interaction: p<0.01, n=10). C: * p<0.05, ** p<0.001, MFA vs. control; # p<0.05, ## p<0.001, Gmax vs. Gmin; (effect of Treatment: p<0.01; effect of COI: p<0.001; interaction: p<0.01 n=12). D: Histogram of OEC Ri values. Black line: preferred model of a comparison of non-linear regression models (Gaussian versus sum of two Gaussian functions, p<0.001, R2=0.782, n=63)
Fig. 6
Fig. 6
The degree of inhibition by channel blockers depended on the initial conductance of OECs. A: Same graph of G vs. Vm shown in Figure 1A2 indicating how changes in conductance were measured for each voltage range of interest. B: Change in conductance (ΔG) produced by each blocker vs. initial conductance (G) at the same voltage range for a subsample of OECs treated with 100 µM Ba2+ (B1), 20 mM TEA (B2) or 100 µM MFA (B3). Solid lines: linear regression analyses; broken lines: 95% confidence interval.
Fig. 7
Fig. 7
Kir, KDR and GJ channels contributed to variability of biophysical properties of OECs. A1: Same data as in Figure S5A (left) showing how distance to the mean was measured. A2: Change in distance to the 3D mean value produced by each channel blocker. * p<0.05, ** p<0.01, Wilcoxon test, comparing to a hypothetical value of zero. B: Change in conductance (ΔG) produced by each blocker vs. Gmin for a subsample of OECs treated with 100 µM Ba2+ (B1) or 20 mM TEA (B2). Solid lines: linear regression analyses; broken lines: 95% confidence interval.
Fig. 8
Fig. 8
GJ channels are regionally expressed in the ONL. A1: Representative image under DIC illumination showing the layers of the OB and the location of the recording pipette (top). The bottom drawing illustrates the procedure to measure distance from the recorded OECs to the inner ONL border. A2: Sensitivity to each blocker versus location of OECs. Each symbol represents a cell; solid lines: linear regression analyses; dotted lines: 95% confidence interval. B1: representative OB slice showing a lucifer yellow-filled OEC (green), immunoreactivity for Cx43 (red) and nuclear stain (blue). Dotted line: inner ONL border. Rectangular selection: representative region of interest (ROI) used for quantification. B2: Average plot profile representing the intensity of Cx43 immunoreactivity as a function of distance from the inner ONL border. Dotted lines: SEM (n=4).

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