EB cells plated on gelatin-coated glass coverslips were placed in the experimental chamber (23°C), and superfused with Tyrode solution of the following composition (mM): 140 NaCl, 5.4 KCl, 2 CaCl2, 1 MgCl2, 5 HEPES, 10 glucose (pH 7.4). Membrane currents or action potentials (AP) from single cells or small clusters of cells were recorded using a computer equipped with pCLAMP 8, a Digidata 1322A series interface and Axopatch 1C amplifier (Molecular Devices). Only cells expressing mCherry fluorescence driven by the MHCα promoter were used for recording. The perforated patch clamp technique was employed. Borosilicate glass pipettes (Sutter Instrument) were filled with (mM) 130 aspartic acid, 146 KOH, 10 NaCl, 2 CaCl2, 5 EGTA, 10 HEPES, 2 Mg-ATP, 100 μg/ml amphotericin (pH 7.2). After forming a gigaseal, progress in electrical access was evaluated by monitoring capacitance currents induced by 20 ms pulses from -35 mV to -40 mV. AP and If were recorded when series resistance was reduced to 40–50 MΩ and 20–30 MΩ respectively. If was induced by voltage steps ranging from -35 to -125 mV with duration decrementing with more negative pulses, followed by a 5 s long pulse to -85 mV to measure tail current and 0.5 s deactivating pulse to -5 mV. Holding potential was -35 mV.
Digidata 1322a series interface
Manufactured by Molecular Devices
The Digidata 1322A Series interface is a data acquisition system designed for electrophysiology applications. It provides high-speed, high-resolution analog-to-digital conversion for recording and analysis of electrical signals from various biological preparations.
Automatically generated - may contain errors
3 protocols using digidata 1322a series interface
1
Perforated Patch-Clamp Recording of Cardiac Myocytes
2
Electrophysiological Recordings of Striatal MSNs
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Brain slices were transferred to a submerged recording chamber and
continuously perfused with regular artificial cerebrospinal fluid (aCSF)
bubbled with 95% O2 and 5% CO2 (pH
7.4). The flow rate was kept at 1.5 mL/min, and bath temperature was
maintained at 30°C–32°C by an inline solution heater
and temperature controller (TC-344B, Warner Instruments, Hamden, CT, USA).
Whole-cell patch-clamp recordings were performed using Axopatch 200B and
Multiclamp 700B amplifiers. Somatic recording from visually identified MSNs
were performed with pipettes (resistance of 3–5 MΩ) filled
with internal solution containing (in mM): 145 K-gluconate, 2
MgCl2, 0.1 EGTA, 2 Na2ATP, and 10 HEPES (pH 7.2
with KOH; 290 mOsm). Access resistance (Ra) was monitored throughout the
recording and was typically <25 MΩ. Data acquisition and
stimulation were performed with a Digidata 1322A Series interface and pClamp
9 software (Molecular Device). Data were filtered at 2 kHz, digitized at 20
kHz, and were analyzed offline with pClamp 10 software. To measure MSN
intrinsic firing 20 μM of NBQX, 100 μM of DL-AP5, and 20
μM of bicuculline were added to regular aCSF in order to prevent
glutamatergic and GABAergic synaptic transmissions, respectively.
continuously perfused with regular artificial cerebrospinal fluid (aCSF)
bubbled with 95% O2 and 5% CO2 (pH
7.4). The flow rate was kept at 1.5 mL/min, and bath temperature was
maintained at 30°C–32°C by an inline solution heater
and temperature controller (TC-344B, Warner Instruments, Hamden, CT, USA).
Whole-cell patch-clamp recordings were performed using Axopatch 200B and
Multiclamp 700B amplifiers. Somatic recording from visually identified MSNs
were performed with pipettes (resistance of 3–5 MΩ) filled
with internal solution containing (in mM): 145 K-gluconate, 2
MgCl2, 0.1 EGTA, 2 Na2ATP, and 10 HEPES (pH 7.2
with KOH; 290 mOsm). Access resistance (Ra) was monitored throughout the
recording and was typically <25 MΩ. Data acquisition and
stimulation were performed with a Digidata 1322A Series interface and pClamp
9 software (Molecular Device). Data were filtered at 2 kHz, digitized at 20
kHz, and were analyzed offline with pClamp 10 software. To measure MSN
intrinsic firing 20 μM of NBQX, 100 μM of DL-AP5, and 20
μM of bicuculline were added to regular aCSF in order to prevent
glutamatergic and GABAergic synaptic transmissions, respectively.
3
Electrophysiological Recordings of Striatal MSNs
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