Journal: PLoS ONE

Article Title: Predominant Functional Expression of Kv1.3 by Activated Microglia of the Hippocampus after S tatus epilepticus

doi: 10.1371/journal.pone.0006770

Figure Lengend Snippet: The leak conductances of the cells in A, B and C were 338, 862 and 332 pS, respectively. The graphs on the right represent the conductance, normalized to its maximum value, as a function of the membrane potential and its inhibition by α-dendrotoxin (A, n = 4, Dtx), agitoxin-2 (B, AgTx, n = 14 for 10 nM, n = 6 for 50 nM) and margatoxin (C, MgTX, n = 8 for 1 nM, n = 11 for 10 nM).

Article Snippet: Alpha-dendrotoxin, recombinant agitoxin-2 and recombinant margatoxin were purchased from Alomone labs (Jerusalem, Israel).

Techniques: Inhibition

Journal: Cells

Article Title: Paclitaxel Inhibits KCNQ Channels in Primary Sensory Neurons to Initiate the Development of Painful Peripheral Neuropathy

doi: 10.3390/cells11244067

Figure Lengend Snippet: Relationship between the paclitaxel- and Kv channel blocker-induced membrane depolarization of dorsal root ganglion (DRG) neurons. ( A ) Membrane potential changes induced by 100 nM α-dendrotoxin and 3 μM paclitaxel. r = −0.509, p = 0.102, Spearman rank order test. ( B ) Membrane potential changes induced by 1 μM phrixotoxin-2 and 3 μM paclitaxel. r = 0.066, p = 0.838, Spearman rank order test. ( C ) Membrane potential changes induced by 1 μM BDS-I (blood-depressing substance-I) and 3 μM paclitaxel. r = 0.038, p = 0.882, Spearman rank order test. ( D ) Membrane potential changes induced by 25 mM TEA and 3 μM paclitaxel. r = 0.673, p = 0.011, Spearman rank order test. ( E ) Representative traces showing XE-991- and paclitaxel-induced membrane depolarization and cell firing in the same cell. ( F ) Membrane potential changes induced by 10 μM XE-991 and 3 μM paclitaxel. r = 0.930, p < 0.0001, Spearman rank order test. Capsaicin-sensitive cells are indicated as red dots. Each filled dot in the plots represents one neuron. N in each panel indicates the number of DRG neurons patched. MP, membrane potential; Pacli, paclitaxel; Caps, capsaicin.

Article Snippet: Paclitaxel, XE-991 (Tocris, Bristol, UK), α-dendrotoxin (Alomone labs, Jerusalem, Israel), phrixotoxin-2 (Alomone labs, Jerusalem, Israel), BDS-II (Alomone labs, Jerusalem, Israel), KT10 (Abcam, Waltham, MA, USA), wortmannin (Tocris, Bristol, UK), BAPTA (Life Tech, Carlsbad, CA, USA), U73122 (Tocris, Bristol, UK), and st-Ht31 (Tocris, Bristol, UK) were added to either bath solution (Paclitaxel, XE-991, α-dendrotoxin, phrixotoxin-2, BDS-II) or pipette solution (KT10, wortmannin, BAPTA, U73122, st-Ht31).

Techniques:

Journal: PLoS ONE

Article Title: Electrical Remodeling of Preoptic GABAergic Neurons Involves the Kv1.5 Subunit

doi: 10.1371/journal.pone.0096643

Figure Lengend Snippet: A. Hongotoxin-1 (0.3 nM) -sensitive component of I DR . Representative currents from a GAD65-GFP (left) and a Kv4.2−/−;GAD65-GFP (right) neuron. B. DPO-1 (0.5 µM) - sensitive component of I DR . Representative currents from a GAD65-GFP (left) and a Kv4.2−/−;GAD65-GFP (right) neuron. C. Margatoxin (5 nM) - sensitive component of I DR . Representative currents from a GAD65-GFP (left) and a Kv4.2−/−;GAD65-GFP (right) neuron. A-C. The currents were elicited by the same voltage step protocol depicted in the inset. TTX (1 µM) was present in all extracellular solutions. D. Bar chart summarizing the current density of the hongotoxin-1 (0.3 nM, n = 13 and n = 16 neurons, respectively)-sensitive, DPO-1 (0.5 µM, n = 23 and n = 26 neurons, respectively)-sensitive, dendrotoxin-K (50 nM, n = 5 and n = 6 neurons, respectively)-sensitive, and margatoxin (5 nM, n = 7 and n = 6 neurons, respectively)-sensitive in GAD65-GFP and Kv4.2−/−;GAD65-GFP neurons. Bars represent averages + S.D. The current densities were calculated using the I DR amplitude at the end of the depolarizing step to 0 mV. ** represents significant differences between GAD65-GFP and Kv4.2−/−;GAD65-GFP groups P<0.01 (t-test).

Article Snippet: DPO-1 and TTX were from Tocris (Ellisville, MO), dendrotoxin-K, hongotoxin-1, margatoxin, were from Alomone Labs (Jerusalem, Israel), while the other substances were purchased from Sigma.

Techniques:

Journal: PLoS ONE

Article Title: Action Potential Modulation in CA1 Pyramidal Neuron Axons Facilitates OLM Interneuron Activation in Recurrent Inhibitory Microcircuits of Rat Hippocampus

doi: 10.1371/journal.pone.0113124

Figure Lengend Snippet: (A) Top, K + current evoked in outside-out patches from the axon (434 µm) of CA1 pyramidal neuron by 200-ms pulses to 50 mV in the absence (Black trace, I Control ) and presence (Gray trace, I 100 nM α-DTX ) of 100 nM α-DTX. Middle, the blue trace represents the α-DTX-sensitive current component obtained by digital subtraction. Bottom, bar plot of the average time constant of inactivation (n = 10). (B) Summary bar graph showing the effect of 100 nM α-DTX in axonal patches (left, P<0.001, n = 13) and somatic patches (right, P>0.1, n = 6). Bars indicate mean ± SEM; circles denote individual experiments. Data points for the same experimental conditions are connected by lines. (C) Recovery of α-DTX-sensitive K + channel from inactivation. Stimulation protocol and example traces of recovery from inactivation, as probed with a double-pulse protocol (100-ms prepulse to –120 mV, 150-ms conditioning pulses and 30-ms test pulses to +50 mV, separated by a recovery interval of variable duration at –120 mV). The holding potential before and after the pulse protocol was –90 mV. Axonal patch is 388 µm from the soma. (D) Summary plot of the amplitude of the peak current evoked by the test pulse, divided by that evoked by the conditioning pulse, plotted against interpulse interval. Data points represent means from 5 patches. Red curves represent double-exponential fit to the data points. (E, F) Gating properties of α-DTX-sensitive axonal K + channels. Stimulation protocol and traces of activation and steady-state inactivation of α-DTX-sensitive K + channels in CA1 pyramidal neuron axons. (E) To probe steady-state activation, a 100-ms prepulse to –120 mV was followed by a 200-ms test pulse to various potentials (–120 to 50 mV). (F) To test steady-state inactivation, a 5-s prepulse to various potentials (–120 to 0 mV) was followed by a 200-ms test pulse to 50 mV. Axonal patch is 423 µm from the soma. (G) Activation (blue circles) and steady-state inactivation (black squares) curves. Conductance values were normalized to the maximal value. Data points represent means from 8 patches for activation curve and 6 patches for steady-state inactivation. Blue curves represent Boltzmann functions fit to the data points. For the activation curve (blue line), the midpoint potential was –12 mV and the sloe factor 26.4 mV. For the inactivation curve (black line), the midpoint potential was –64 mV and the slope factor 15.2 mV. (H) Length constant of the CA1 pyramidal neuron axons. Plot of axonal to somatic voltage deflection during hyperpolarizing current pulses (1 s, –30 pA) applied at the soma, plotted against distance. Each data point represents a simultaneous axon–soma recording. Red line represents a fit of data points with an exponential function, resulting in a mean length constant of 712 µm. Note that the long length constant results in particularly efficient propagation of subthreshold membrane potential changes from the soma to the axon. (Inset) voltage changes recorded in the soma (black) and the axon (blue) during hyperpolarizing current pulses applied at the soma. Axonal recording site is 705 µm from the soma. Traces shown represent average of 20 (A, black trace), 22 (A, gray trace), 2 (C), 3 (E), and 2 (F) single sweeps. Transient inward Na + currents at the beginning of the pulse are truncated. Error bars, SEM.

Article Snippet: For paired recordings, K-gluconate and EGTA concentrations were modified to 135 mM and 0.1 mM, respectively. α-Dendrotoxin (α-DTX) was purchased from Alomone labs. For experiments with α-DTX, 0.1% bovine serum albumin (BSA) was added to the extracellular solutions to prevent peptide adsorption.

Techniques: Activation Assay

Journal: PLoS ONE

Article Title: Action Potential Modulation in CA1 Pyramidal Neuron Axons Facilitates OLM Interneuron Activation in Recurrent Inhibitory Microcircuits of Rat Hippocampus

doi: 10.1371/journal.pone.0113124

Figure Lengend Snippet: (A) Block of Kv1 channels enhances transmission. Top, unitary EPSCs at CA1 PN–O-LM IN synapses evoked by single presynaptic APs in control conditions and in the presence of 100 nM α-DTX. Traces represent averages from 10 single sweeps. Bottom, summary bar graph of the effects of α-DTX on EPSC peak amplitude. Note that α-DTX markedly increased synaptic efficacy. (B) Block of Kv1 channels reduces facilitation of transmission. Top, EPSCs at CA1 PN–O-LM IN synapses evoked by trains of five presynaptic APs in control conditions (left) and in the presence of 100 nM α-DTX (right). Bottom, facilitation ratio (EPSC n /EPSC 1 ), plotted against stimulus number, in control conditions (squares) and in the presence of α-DTX (circles). Somatic holding potential –60 mV. (C) Block of Kv1 channels abolishes static analog modulation of transmission. Top, unitary EPSCs at CA1 PN–O-LM IN synapses evoked by single presynaptic APs in control conditions and in the presence of 100 nM α-DTX. Presynaptic membrane potential were held at –60 mV (left; same recording as in (A)), and –50 mV (right). Bottom, summary bar graph of the effects of α-DTX on EPSC peak amplitude (EPSC DTX /EPSC Control ) for two different presynaptic holding potential (black, –60 mV; red, –50 mV). Note that α-DTX occluded the effects of changing membrane potential in the presynaptic neuron. (D) Block of Kv1 channels broadens the axonal AP. Top, axonal AP traces in control conditions and in the presence of 100 nM α-DTX. Bottom, summary bar graph showing the effects of α-DTX on half-duration of somatic and axonal AP. Axonal recording site is 264 µm from the soma. Note that α-DTX selectively increased axonal AP duration. (E) Block of Kv1 channels reduces activity-dependent AP broadening. Top, superposition of 1 st , 5th, and 50 th axonal AP in control conditions (left) and in the presence of 100 nM α-DTX (right). Bottom, plot of axonal AP half-width against stimulus number in control conditions (squares) and in the presence of 100 nM α-DTX (circles). Somatic holding potential –60 mV. Data from 7 recordings at distances of 200 to 500 µm. Axonal recording site is 264 µm from the soma. (F) Block of Kv1 channels reduces static AP broadening. Top, superposition of axonal APs in control conditions and in the presence of 100 nM α-DTX at –60 mV (left; same recording as in (D)), and –50 mV (right). Bottom, summary bar graph of the effects of α-DTX on AP broadening at –60 mV (black) and –50 mV (red). Note that the depolarization-induced AP broadening reduced the effect of α-DTX. Axonal recording site is 264 µm from the soma. Bars indicate mean ± SEM. Open circles represent data from individual experiments. Data from the same experiment or for the same experimental conditions were connected by lines. *0.01≤P<0.05.

Article Snippet: For paired recordings, K-gluconate and EGTA concentrations were modified to 135 mM and 0.1 mM, respectively. α-Dendrotoxin (α-DTX) was purchased from Alomone labs. For experiments with α-DTX, 0.1% bovine serum albumin (BSA) was added to the extracellular solutions to prevent peptide adsorption.

Techniques: Blocking Assay, Transmission Assay, Activity Assay