Pclamp 10.0 software package

Manufactured by Molecular Devices

PCLAMP 10.0 is a software package designed for data acquisition and analysis of electrophysiological experiments. It provides a comprehensive suite of tools for recording, analyzing, and visualizing electrical signals from cells and tissues.

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Lab products found in correlation

Brainphys medium, by STEMCELL (1 mentions) Multiclamp 700b amplifier, by Molecular Devices (1 mentions) Bx61 microscope, by Olympus (1 mentions) Clampfit 10, by Molecular Devices (1 mentions) Digidata 1440a digitizer, by Molecular Devices (1 mentions)

4 protocols using pclamp 10.0 software package

Membrane Current Measurement Protocols

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Macroscopic membrane
current was recorded 48–72 h after injection as described previously.^{8 (link)} The tested compounds were applied at pH 5.5 at
various concentrations when the inward current reaches maximum. The
compounds were applied for 2 min, and residual membrane current was
compared with the membrane current before the application of compounds.
Membrane currents were analyzed with pCLAMP 10.0 software package
(Axon Instruments, Sunnyvale, CA).

Rey-Carrizo M., Barniol-Xicota M., Ma C., Frigolé-Vivas M., Torres E., Naesens L., Llabrés S., Juárez-Jiménez J., Luque F.J., DeGrado W.F., Lamb R.A., Pinto L.H, & Vázquez S. (2014). Easily Accessible Polycyclic Amines that Inhibit the Wild-Type and Amantadine-Resistant Mutants of the M2 Channel of Influenza A Virus. Journal of Medicinal Chemistry, 57(13), 5738-5747.

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Macroscopic Membrane Current Measurement

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Macroscopic membrane current was recorded 24–72 hours after injection as described previously.^{31 (link)} Oocytes were perfused at room temperature in Barth’s solution containing (in mM) 88 NaCl, 1 KCl, 2.4 NaHCO₃, 0.3 NaNO₃, 0.71 CaCl₂, 0.82 MgCl₂, and 15 HEPES for pH 8.5 or 15 MES for pH 5.5 at a rate of 2 mL/min. The tested compounds were dissolved in DMSO and applied (100 μM) at pH 5.5 when the inward current reaches maximum. The compounds were applied for 2 min, and residual membrane current was compared with the membrane current before the application of compounds. The compounds were typically applied for 2 min, and residual membrane current was compared with the membrane current before the application of compounds. Membrane currents were recorded at −20 mV and analyzed with pCLAMP 10.0 software package (Axon Instruments, Sunnyvale, CA).

Barniol-Xicota M., Gazzarrini S., Torres E., Hu Y., Wang J., Naesens L., Moroni A, & Vázquez S. (2017). Slow but steady wins the race: dissimilarities among new dual inhibitors of the wild-type and the V27A mutant M2 channels of influenza A virus. Journal of medicinal chemistry, 60(9), 3727-3738.

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Whole-cell recordings of hiPSC neurons

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For whole-cell recordings, hiPSC-derived neurons from one iPSC line (line 3) were visualized using an upright Olympus BX61 microscope equipped with a 40x objective and differential interference contrast optics. Neurons were constantly perfused with BrainPhys® medium (STEMCELL Technologies, Catalog #05790) preheated to 30-31°C. Patch electrodes were filled with internal solutions containing 130 mM K⁺gluconate, 6 mM KCl, 4 mM NaCl, 10mM Na⁺HEPES, 0.2 mM K⁺EGTA; 0.3mM GTP, 2mM Mg²⁺ATP, 0.2 mM cAMP, 10mM D-glucose. The pH and osmolarity of the internal solution were adjusted to resemble physiological conditions (pH 7.3, 290–300 mOsmol). Current- and voltage-clamp recordings were carried out using a Multiclamp 700B amplifier (Molecular Devices), digitized with Digidata 1440A digitizer and fed to pClamp 10.0 software package (Molecular Devices). For spontaneous EPSC recordings, neurons were held at chloride reversal potential of −75 mV. Data processing and analysis were performed using ClampFit 10.0 (Molecular Devices) and Prism software. CD49f⁺ astrocytes for co-cultures were from three iPSC lines. p-values to compare neurons alone to neurons with astrocytes or neurons with A0 astrocytes to neurons with A1 astrocytes were calculated using a two-tailed, unpaired t-test.

Barbar L., Jain T., Zimmer M., Kruglikov I., Sadick J., Wang M., Kalpana K., Rose I.V., Burstein S.R., Rusielewicz T., Nijsure M., Guttenplan K.A., di Domenico A., Croft G., Zhang B., Nobuta H., Hébert J.M., Liddelow S.A, & Fossati V. (2020). CD49f is a novel marker of functional and reactive human iPSC-derived astrocytes. Neuron, 107(3), 436-453.e12.

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Electrophysiological Characterization of DRG Neurons

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The current clamp recording was performed to record AP 2 to 6 h after DRG neurons plating using EPC‐10 amplifier and Pulse software (HEKA Instruments). The intracellular solution contained (in mm) 140 K‐gluconate, 10 NaCl, 5 EGTA, 1 CaCl₂, 10 HEPES, 2 Mg‐ATP, 0.2 Na‐ATP (pH 7.3 adjusted with NaOH, 300 mOsm). The extracellular solution contained (in mm) 129 NaCl, 5 KCl, 2 CaCl₂, 1 MgCl₂, 25 HEPES, 30 Glucose (pH 7.4 adjusted with KOH, 310 mOsm). DRG neurons were examined for evoked AP with a series of 1‐s current injections from 0 to 580 pA in 20 pA increments. The following values were measured in this study: RMP, AP threshold, rheobase current, evoked AP numbers, AP amplitude, AP peak, AP half‐width, and AHP amplitude. RMP was measured 2 min after a stable recording was obtained. The rheobase current was defined as the minimum current sufficient to evoke an AP in 20 ms. AP threshold was defined as the first point on the rising phase of an AP where depolarization was greater than 50 mV ms⁻¹.^[⁵³^] The AHP amplitude was measured between the maximum hyperpolarization and the final plateau voltage.^[⁶^] The DRG neurons that had stable membrane potentials more negative than −40 mV were included for further analysis.^[⁶^] The data were analyzed by the pCLAMP 10.0 software package (Molecular Devices). All experiments were performed at room temperature.

Sun Z., Waybright J.M., Beldar S., Chen L., Foley C.A., Norris‐Drouin J.L., Lyu T., Dong A., Min J., Wang Y., James L.I, & Wang Y. (2022). Cdyl Deficiency Brakes Neuronal Excitability and Nociception through Promoting Kcnb1 Transcription in Peripheral Sensory Neurons. Advanced Science, 9(10), 2104317.

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