OPERATING INSTRUCTIONS AND
SYSTEM DESCRIPTION FOR THE
ELC-03XS
UNIVERSAL AMPLIFIER
for EXTRA & INTRACELLULAR RECORDING,
SINGLE CELL STIMULATION and
ELECTROPORATION
VERSION 2.2
npi 2011
npi electronic GmbH, Hauptstrasse 96, D-71732 Tamm, Germany
Phone +49 (0)7141-9730230; Fax: +49 (0)7141-9730240
support@npielectronic.com; http://www.npielectronic.com
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Table of Contents
1. Safety Regulations ..............................................................................................................3
2. Introduction......................................................................................................................... 4
3. ELC-03XS amplifier...........................................................................................................6
3.1. ELC-03XS Components.............................................................................................. 6
3.1. Optional Accessories...................................................................................................6
3.2. System Description...................................................................................................... 6
Operation modes of the amplifier................................................................................ 6
Input configuration:.....................................................................................................7
Computer control of the mode of operation................................................................ 7
Output configuration.................................................................................................... 7
Digital displays............................................................................................................ 7
Oscillation shut-off unit............................................................................................... 7
Penetration unit............................................................................................................ 7
3.3. Front Panel View of the ELC-03XS Amplifier........................................................... 8
3.4. Description of the Front Panel..................................................................................... 9
4. Description of the Rear Panel............................................................................................. 19
Grounding.................................................................................................................... 21
5. Setting up the ELC-03XS ................................................................................................... 22
6. Passive Cell Model ............................................................................................................. 23
6.1. Cell Model Description ............................................................................................... 23
6.2. Connections and Operation ......................................................................................... 24
7. Headstage............................................................................................................................ 26
7.1. Headstage Elements..................................................................................................... 26
7.2. Headstage Bias Current Adjustment............................................................................ 27
7.3. Headstage with x1 / x100 current range (option)........................................................ 28
8. Introduction into Experiments ............................................................................................ 29
8.1. Recordings with the Differential Headstage (optional)............................................... 29
8.2. Extracellular Voltage Measurement............................................................................30
8.3. Extracellular Stimulation and Electroporation............................................................31
Stimulation with Current............................................................................................. 31
Electroporation with Current....................................................................................... 31
Stimulation with Voltage............................................................................................. 31
Electroporation with Voltage....................................................................................... 31
8.4. Intracellular Recording................................................................................................ 32
Current Clamp Recording............................................................................................ 32
Voltage Clamp Recording........................................................................................... 32
9. Literature............................................................................................................................. 33
10. Technical Data............................................................................................................. 35
Index........................................................................................................................................37
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1. Safety Regulations
VERY IMPORTANT: Instruments and components supplied by npi electronic are NOT
intended for clinical use or medical purposes (e.g. for diagnosis or treatment of humans), or
for any other life-supporting system. npi electronic disclaims any warranties for such purpose.
Equipment supplied by npi electronic must be operated only by selected, trained and
adequately instructed personnel. For details please consult the GENERAL TERMS OF
DELIVERY AND CONDITIONS OF BUSINESS of npi electronic, D-71732 Tamm,
Germany.
GENERAL: This system is designed for use in scientific laboratories and must be operated
only by trained staff. General safety regulations for operating electrical devices should be
followed.
AC MAINS CONNECTION: While working with npi systems, always adhere to the
appropriate safety measures for handling electronic devices. Before using any device please
read manuals and instructions carefully.
The device is to be operated only at 115/230 Volt 60/50 Hz AC. Please check for appropriate
line voltage before connecting any system to mains.
Always use a three-wire line cord and a mains power-plug with a protection contact
connected to ground (protective earth).
Before opening the cabinet, unplug the instrument.
Unplug the instrument when replacing the fuse or changing line voltage. Replace fuse only
with an appropriate specified type.
STATIC ELECTRICITY: Electronic equipment is sensitive to static discharges. Some
devices such as sensor inputs are equipped with very sensitive FET amplifiers, which can be
damaged by electrostatic charge and must therefore be handled with care. Electrostatic
discharge can be avoided by touching a grounded metal surface when changing or adjusting
sensors. Always turn power off when adding or removing modules, connecting or
disconnecting sensors, headstages or other components from the instrument or 19” cabinet.
TEMPERATURE DRIFT / WARM-UP TIME: All analog electronic systems are sensitive to
temperature changes. Therefore, all electronic instruments containing analog circuits should
be used only in a warmed-up condition (i.e. after internal temperature has reached steady-state
values). In most cases a warm-up period of 20-30 minutes is sufficient.
HANDLING: Please protect the device from moisture, heat, radiation and corrosive
chemicals.
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2. Introduction
“Loose patch” recordings (or “loose seal” recordings [Roberts & Almers, 1992]) are used to
record from single excitable cells without damage, i.e. without a direct access to the cell
interior. The first recordings were made around 1960 from muscles cells by Alfred Strickholm
long time before “tight seal” recording was invented by Erwin Neher and Bert Sakmann
twenty years later: “A method has been developed permitting measurement of membrane
impedance and current, as a function of transmembrane potential, at small, electrically
isolated regions of the muscle cell surface without microelectrode impalement.”[Strickholm
1961].
The loose seal has a resistance of a few ten to a few hundred M, and it creates an electrically
isolated access to a single neuron. This isolated area can be used for precise recording,
stimulation or drug and dye application on the single cell level without damaging the cell
[Babour & Isope, 2000]. In contrast to tight seal recordings the same electrode can be reused
for recording from several cells, which is a great advantage.
Since its beginnings several attempts have been made to make such precise extracellular
methods accessible to various preparations. A nice overview can be found in the chapter by
Roberts & Almers [Roberts & Almers, 1992]. Over the years the method was extended to
cultured neurons and brain slice preparations, and also for in vivo recordings [Bureau et al,
2004]. The method is particularly well suited for long term recording with little damage to the
recorded neuron [Nunemaker et al, 2003]. It can be used both for somatic and axonal
recording [Khaliq & Raman 2005]. Even subcellular structures such as synaptic boutons are
accessible to loose patch recordings [Auger & Marty, 2000].
Another valuable application of this method is single cell stimulation. The high resistance
loose patch makes possible the application of 1-2 V stimuli to one cell only [Babour & Isope,
2000].
In the nineties of the last century the method of juxtacellular dye application (juxtasomal
filling) became popular [Pinault, 1996]. This staining method is based on repetitive current
pulse trains applied in the close vicinity of cell somata or dendrites and is meanwhile well
established in the field of slice and in vivo preparations [Klausberger, 2004]. Juxtacellular
filling together with extracellular measurements are today often summarized under the term
“juxtacellular recording”.
In parallel attempts were made towards transfection of single cells by electroporation using
patch pipettes. DNA or other large molecules were successfully inserted through a patch
pipette into living cells by using an optimized protocol (application of 10 V / 1 ms pulse
trains) [Rathenberg et al, 2003].
Far in excess of classical in vivo recording methods [Lalley et al, 1999] several new
approaches are used for monitoring neuronal activity under natural conditions, using new
techniques, e.g. the combination of two photon excitation and patch clamp in vivo [Helmchen
et al, 2001; Stosiek et al, 2003; Brecht et al, 2002]. Assays have been developed that allow to
monitor and manipulate single cells under in vivo conditions [Brecht et al, 2004]. Besides
sophisticated optics these techniques always require precise recording and stimulation
amplifiers, mostly based on the use of patch electrodes.
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Today three methods are used to record electrical in vivo or in vitro:
Recordings using patch (suction) electrodes from single neurons
Whole cell patch clamp technique (tight seal recording, intracellular)
Loose patch technique (loose seal recording, extracellular)
Intracellular recordings with sharp microelectrodes
Extracellular recordings with glass or metal electrodes
The amplifiers used for such recordings are specialized on the recording of the potentials or
currents generated by the neurons under investigation. If these recording methods are
combined with dye injection, electroporation, stimulation protocols etc. through the recording
electrode, serious constraints occur and several additional devices have to be added to the
experimental set-up.
The ELC series of amplifiers fills this gap. It allows intracellular, extracellular, voltage –
clamp or current clamp recordings both with sharp or patch electrodes as well as additional
protocols like electroporation or juxtasomal recordings. Even iontophoresis and
voltammetry/amperometry can be performed.
The ELC amplifier is the “Swiss Army Knife” of modern electrophysiology. It is easy to use,
versatile, and makes possible a lot of sophisticated experiments with only one instrument.
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3. ELC-03XS amplifier
3.1. ELC-03XS Components
The following items are shipped with the system:
ELC-03XS amplifier
GND (2.4 mm banana jack) connector and (optional) REF. connector for headstage
Spare fuse
Headstage
User manual
3.1. Optional Accessories
o Differential headstage
o Differential miniature headstage
o Cell model
o Pipette holder
3.2. System Description
The ELC-03XS was designed for intra- and extracellular recording, precise (single cell)
electrical stimulation as well as juxtasomal filling with patch electrodes. The system consists
of an amplifier in a rackmount cabinet and a small headstage with mounting plate or a holding
bar. It can be used in slices or in in vivo preparations using the optional headstage with a
differential input. It has separate capacity compensation controls for VC and CC mode, all
controls (Bridge balance, CC cap comp, offset, holding controls) are calibrated 10-turn
potentiometers.
The ELC-03XS is capable to record extracellulary DC or AC coupled, to stimulate with
current or voltage and to perform non-invasive juxtasomal filling of cells with dyes or DNA.
The amplifier can also be used to record and stimulate intracellulary in current clamp (CC)
with sharp or patch electrodes, or voltage (patch) clamp (VC) with patch electrodes. It is also
suitable for amperometric or voltammetric investigations with carbon fiber electrodes.
Operation modes of the amplifier
The operation modes of the amplifier are selected by a rotary switch with six positions:
The selected mode is indicated by LEDs above the MODE OF OPERATION switch.
EXT: All MODEs OF OPERATION can be selected by TTL signals connected to the
rear panel (see chapter 4).
CC: CURRENT CLAMP MODE: used to inject predefined current signals
CCx10: CURRENT CLAMP MODE : for stimulation and electroporation
OFF: CC Mode with all input signals turned off
VC: VOLTAGE CLAMP mode: potential commands are applied to the electrode
VCx10: High range VC mode, for stimulation and electroporation
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In addition, using a toggle switch a bridge balance circuit can be activated, to compensate for
the electrode artifact (BRIDGE mode, only in CC mode). An ELECTRODE RESISTANCE
test mode can be activated with a push button. The electrode resistance is measured directly in
M and displayed on the POTENTIAL display.
Input configuration:
The amplifier has two inputs, both for VC and CC mode. The signal applied to the analog
input BNCs is converted either into a voltage command signal (x1 or x10 scaling) for the VC
or VCx10 voltage clamp modes, or to a current in the CC, CCx10 and BRIDGE mode.
Besides this, a signal generated from the 10-turn HOLD potentiometer can be transferred into
a pulse using the GATE TTL input BNC. This control can be also used as HOLDING
potentiometer if the switch in the GATE BNC is turned off.
Computer control of the mode of operation
In the EXT position of the MODE SELECT switch all MODEs OF OPERATION can be
selected by TTL signals connected to the rear panel (see chapter 4).
Output configuration
The ELC-03XS amplifier has two output BNCs for POTENTIAL and one output BNC for the
CURRENT signal. The POTENTIAL OUTPUT x1 is a pure DC output that monitors the
electrode potential directly from the headstage. The signal at the POTENTIAL OUTPUT can
be high and low-pass filtered and amplified.
The current output signal is amplified and can be filtered by a low-pass filter.
Digital displays
All ELC amplifiers are equipped with two digital displays, one for CURRENT (nA) and one
for POTENTIAL (mV) or ELECTRODE RESISTANCE (M). The mode of operation is
indicated by LEDs located close to the digital displays.
Oscillation shut-off unit
All ELC amplifiers are equipped with an oscillation shut-off unit to protect the preparation. If
it is active all outputs of the amplifier were disconnected if the system begins to oscillate.
Penetration unit
All ELC amplifiers are also equipped with a penetration unit (BUZZ) to facilitate the
penetration of the cell membrane or to clean clogged electrodes.
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3.3. Front Panel View of the ELC-03XS Amplifier
Figure 1: ELC-03XS front panel view
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3.4. Description of the Front Panel
In the following description of the front panel elements each element has a number (in bold)
that is related to that in Figure 1. The number is followed by the name (in uppercase letters)
written on the front panel and the type of the element (in lowercase letters). Then, a short
description of the element is given. Some elements are grouped in functional units (e.g.
PENETRATION / BUZZ unit) and are described as units regardless of the order of numbers.
(1) HEADSTAGE connector
Connector for the headstage with optional differential input. REF of the headstage
must be connected to ground (single-ended measurement) or to a reference electrode
in the bath (differential measurement), see also Figure 11.
PENETRATION / BUZZ unit
The PENETRATION / BUZZ unit consist of (2) mode switch, (3) REMOTE
connector, (4) MANUAL push button, (5) FREQUENCY potentiometer,
(47) AMPLITUDE potentiometer and (48) DURATION potentiometer.
(2) mode switch
5-position switch to set the mode of PENETRATION
CC: The BUZZ circuit is activated (duration set by 48). To facilitate the penetration
of the cell membrane the BUZZ circuit is provided which is based on
oscillations caused by overcompensating the capacitance compensation system.
The overcompensation of capacitance compensation yields to very powerful
high-frequency (approx. 2 kHz) oscillations (see Figure 2).
PULSE: A pulse train is applied to the electrode. The pulses are positive and rectangular.
The AMPLITUDE is set by 47, the DURATION of the train is set by 48 and the
FREQUENCY of pulses within the train is set by 5.
+I
max
: The maximum positive current is applied to the electrode. The DURATION is
set by 48.
-I
max
: The maximum negative current is applied to the electrode. The DURATION is
set by 48.
+I
max
or -I
max
can also be used to clean the tip of the electrode by passing large amounts of
positive or negative current. The maximum current is dependent on the setting of the
current range, see also (42).
OFF: The PENETRATION / BUZZ unit is disabled. When in OFF position the
preparation is protected from unintentional use of the unit.
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(3) REMOTE connector
BNC connector to attach a remote switch in active-low configuration for activating
the PENETRATION / BUZZ unit.
(4) MANUAL push button
Push button for activating the PENETRATION / BUZZ unit manually. The
DURATION is set by 48 and the mode by 2.
Note: The duration is dependent on the setting of the DURATION potentiometer, but
independent from how long the button is pushed or a REMOTE switch is pressed.
-12000
-8000
-4000
0
4000
8000
12000
time
potential (mV)
start stop
Figure 2: BUZZ function of the ELC-03XS (CC Mode)
(5) FREQUENCY potentiometer
Potentiometer to set the FREQUENCY of pulses within a train in PULSE mode
(see also 2). Range: 100 Hz to 2 kHz.
(47) AMPLITUDE potentiometer
Potentiometer for setting the AMPLITUDE of pulses in PULSE mode (see also 2).
(48) DURATION potentiometer
Control to set the duration of the BUZZ (potentiometer, clockwise, range: ~1 ms to
~100 ms). It is effective in all modes; REMOTE controlled and when pushing the
MANUAL button.
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(6) CURRENT OUTPUT connector
BNC connector providing the current output signal; scaling is set by CURRENT
OUTPUT SENSITIVITY (V/nA) switch 46, filter is set by CURRENT OUTPUT
FILTER LOWPASS (Hz) switch 7.
(7) CURRENT OUTPUT FILTER LOWPASS (Hz) switch
16-position rotary switch to select the corner frequency of the LOWPASS
FILTER (range: 20 Hz to 20 kHz) for the CURRENT OUTPUT connector (6).
(8) CURRENT FROM HEADSTAGE (0.1V/nA) connector
BNC connector providing a voltage proportional to the current at the electrode
with a fixed scaling of 100 mV / nA. This current signal is not filtered.
CURRENT set unit
The CURRENT set unit consist of (9) CURRENT nA potentiometer, (10)
+/0/- switch and (11) GATE / HOLD switch.
(9) CURRENT nA potentiometer
Ten-turn potentiometer for generating a holding current (switch 11 in HOLD
position) or a gated stimulus (switch 11 in GATE position) in CC / BRIDGE or
CCx10 mode, range: 10 nA (100 = 1 nA) or 100 nA (100 = 10 nA),
respectively. Polarity is set by the switch 10.
(10) +/0/- switch
Switch for setting the polarity of the holding current or gated current stimulus,
respectively (see also 9). In zero position the holding current or gated current
stimulus is disabled.
(11) GATE / HOLD switch
Switch that determines the function of the CURRENT nA potentiometer 9 (see
also 9).
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(12) BRIDGE BALANCE M potentiometer
Potentiometer for balancing the BRIDGE circuit; 10 M / turn, range: 0 to 100
M. The BRIDGE must be balanced correctly before using the series resistance
compensation in VC mode, because this measure is taken as value for the series
resistance compensation (see also 43 and 44).
(13) STIMULUS INPUT switch
Switch for grounding BNC connector 14. If 14 is not in use, please switch the INPUT
off by setting this switch to OFF. This prevents noise pick-up through an open input.
(14) STIMULUS INPUT 1 nA/V connector
BNC connector for the current stimulus in CC / BR or in CCx10 mode; scaling 1 nA /
V or 10 nA / V, respectively.
(15) GATE TTL connector
BNC connector for gating the potential step in VC / VCx10 mode, or the current step
in CC / CCx10 mode.
VC modes: As long as the voltage linked to this BNC is HIGH, i.e. +5 V, a voltage stimulus
with the amplitude set by the POTENTIAL (mV) potentiometer 21 is generated
by the amplifier. Switch 19 has to be in GATE position.
CC modes: As long as the voltage linked to this BNC is HIGH, i.e. +5 V, a current stimulus
with the amplitude set by the CURRENT (nA) potentiometer 9 is generated by
the amplifier. Switch 11 has to be in GATE position.
(16) COMMAND INPUT switch
Switch for grounding BNC connector 17. If 17 is not in use, please switch the INPUT
off by setting this switch to OFF. This prevents noise pick-up through an open input.
(17) COMMAND INPUT 10 connector
BNC connector for the command potential in VC mode or VCx10 mode; scaling:
10 mV or 1 mV, respectively.
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(18) SERIES RESISTANCE COMP. (%) potentiometer
Potentiometer for setting the compensation for the SERIES RESISTANCE (VC
mode) in percent of the resistance value that is set at the BRIDGE BALANCE
potentiometer 12. Therefore, it is important to set the BRIDGE BALANCE
correctly before using the series resistance compensation (see also 40 and 41).
Very Important: When using the SERIES RESISTANCE compensation do not use the
potential values recorded from the POTENTIAL OUTPUT (mV) BNC connector (27) or the
value shown at the POTENTIAL display (39) as a measure for the membrane potential, since
these values include also the additional voltage generated by SERIES RESISTANCE COMP.
for compensation of the voltage drop at the SERIES RESISTANCE.
For instance, when for calculating I/V curves do not use the potential values recorded from
the POTENTIAL OUTPUT (mV) BNC connector (27). Use the COMMAND potential
instead.
COMMAND set unit
The COMMAND set unit consist of (19) GATE / HOLD switch, (20) +/0/switch and (21) POTENTIAL (mV) potentiometer
(19) GATE / HOLD switch
Switch that determines the function of the POTENTIAL (mV) potentiometer 21
(see also 21).
(20) +/0/- switch
Switch for setting the polarity of the holding potential or gated potential stimulus in
VC mode, respectively (see also 21). In zero position the holding potential or gated
potential stimulus is disabled.
(21) POTENTIAL (mV) potentiometer
Ten-turn potentiometer for generating a holding potential (switch 19 in HOLD
position) or a gated stimulus (switch 19 in GATE position) in VC or VCx10
mode; range: 1 V (100 = 100 mV) or 10 V (100 = 1 V), respectively. Polarity
is set by the switch 20.
OSCILLATION SHUT OFF unit
The OSCILLATION SHUT OFF unit consist of (22) THRESHOLD potentiometer,
(23) DISABLED / RESET switch and of (35) OSCILLATION SHUT OFF LED.
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(22) THRESHOLD potentiometer
Control to set the activation THRESHOLD of the OSCILLATION SHUT-OFF
circuit (potentiometer, linear clockwise, range: 0-1200 mV).
(23) DISABLED/RESET switch
Switch to DISABLE the OSCILLATION SHUT-OFF unit or RESET the circuit.
RESET is done if one wants to reset the circuit after previous activation. After
resetting the OSCILLATION SHUT-OFF unit is active again.
(35) OSCILLATION SHUT-OFF LED
Indicates whether the OSCILLATION SHUT-OFF circuit is active (LED: red) or
not (LED: green).
Note: If the OSCILLATION SHUT-OFF unit is active and the SHUT-OFF was triggered, the
output of the amplifier to the headstage is disabled. The amplifier is automatically switched to
CC-OFF mode and only potential measurement works.
POTENTIAL OUTPUT FILTER
The POTENTIAL OUTPUT FILTER consist of (24) HIGHPASS (Hz)
and of (26) LOWPASS (Hz), and is used mainly for filtering
extracellular signals.
(24) HIGHPASS (Hz) switch
16-position rotary switch to select the corner frequency of the HIGHPASS filter
(range: DC to 3 kHz) for the POTENTIAL OUTPUT (mV) connector 27. In DC
position the HIGHPASS is disabled.
(26) LOWPASS (Hz) switch
16-position rotary switch to select the corner frequency of the LOWPASS filter
(range: 20 Hz to 20 kHz) for the POTENTIAL OUTPUT (mV) connector 27.
(25) POTENTIAL OUTPUT FROM HEADSTAGE (V) connector
BNC connector providing the POTENTIAL at the electrode from the headstage.
This POTENTIAL OUTPUT is not filtered and fixed scaled (x1 V).
(27) POTENTIAL OUTPUT (mV) connector
BNC connector providing the POTENTIAL at the electrode. The scaling is set by
POTENTIAL OUTPUT GAIN switch 34. This POTENTIAL OUTPUT is filtered by
(26) LOWPASS and (24) HIGHPASS FILTER.
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CAPACITY COMPENSATION (VC) unit
The CAPACITY COMPENSATION (VC) unit consist of (28) TIME
CONSTANT (-FAST) potentiometer and of (32) AMPLITUDE (C-FAST)
potentiometer. It compensates for the electrode capacity and functions only in
VC mode. Tuning is done by application of voltage pulses to the electrode and
trying to get the resulting current pulses as fast as possible using potentiometers
28 and 32.
(28) TIME CONSTANT (-FAST) potentiometer
Potentiometer for setting -FAST in the electrode capacity compensation circuit.
(32) AMPLITUDE (C-FAST) potentiometer
Potentiometer for setting C-FAST in the electrode capacity compensation circuit.
Note: CAPACITY COMPENSATION works only for the electrode capacity, not for the
capacity of the cell membrane. Therefore, capacitive transients are always present when
square shaped pulses are applied in VC mode.
(29) GROUND connector
Banana jack providing system GROUND. System GROUND is not connected to the
chassis or to protective earth, respectively.
(30) POWER switch
Push button to switch the amplifier ON (pushed) or OFF (released).
(31) AUDIO volume potentiometer
Potentiometer for setting the volume of the AUDIO monitor for POTENTIAL.
Turning the potentiometer clockwise increases the volume.
(33) PIPETTE HOLD POTENTIAL (VC)
Ten-turn potentiometer setting the current through the pipette to zero when
approaching a cell in VC mode, i.e. the pipette is clamped to its OFFSET potential
resulting in zero current flow through the pipette. In VCx10 mode the pipette hold
potential does not change. Range is 100 mV and zero position is 5, i.e. the same as the
OFFSET potentiometer 50. This is very convenient since this adjustment can be done starting
with the same value as in 50.
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(34) POTENTIAL OUTPUT GAIN switch
7-position switch for selecting the amplification for the signal at (27)
POTENTIAL OUTPUT (mV); range: x10 to x1000.
(36) ELECTRODE RESISTANCE TEST push button
Push button for activating the ELECTRODE RESISTANCE TEST. The
ELECTRODE RESISTANCE is determined by application of 1 nA square pulses to
the electrode and is shown in M at display 39. Function is indicated by the M
LED 37. The ELECTRODE RESISTANCE TEST works in OFF mode as well.
Hints: This function can also be used to adjust (51) CAPACITY COMP. (CC). The
POTENTIAL OUTPUT FILTER LOWPASS (26) should be set to 20 kHz and the
HIGHPASS (24) to DC.
With an amplifier with switchable headstage (ELC-SWI) the electrode resistance test works
also in enhanced (x100) current mode.
Note: If the ELC-03XS is equipped with the optional SEAL TEST circuit, the push button is
replaced by a three position switch. In the upper position the ELECTRODE RESISTANCE
TEST is carried out and in the lower position the SEAL resistance test is accomplished. The
value of the SEAL resistance is shown at the CURRENT display (45).
(37) M LED
LED indicating that the unit of the POTENTIAL/RESISTANCE display (39) is M.
(38) mV LED
LED indicating that the unit of the POTENTIAL/RESISTANCE display (39) is mV.
(39) POTENTIAL / RESISTANCE display
Display for the potential at the electrode in XXXX mV (1999 mV max.) or the
electrode resistance in XXX M (999 M max.). The unit is indicated by 37 or
38.
SERIES RESISTANCE COMPENSATION unit
The SERIES RESISTANCE COMPENSATION unit consist of (18) SERIES
RESISTANCE COMP. (%) potentiometer, (40) RS COMP. switch and (41) RS
COMP. LED
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(18) SERIES RESISTANCE COMP. (%) potentiometer
Potentiometer for setting the compensation for the SERIES RESISTANCE in
percent of the value that is set at the BRIDGE BALANCE potentiometer 12.
Therefore, it is important to set the BRIDGE BALANCE correctly before using the
series resistance compensation to avoid oscillations (see also 40 and 41).
(40) RS COMP. switch
Switch for activating the SERIES RESISTANCE COMPENSATION circuit. The
active circuit is indicated by 41.
(41) RS COMP. LED
LED indicating that the SERIES RESISTANCE COMPENSATION circuit is active. The
amount of SERIES RESISTANCE COMPENSATION is set by 18.
Very Important: When using the SERIES RESISTANCE compensation do not use the
potential values recorded from the POTENTIAL OUTPUT (mV) BNC connector (27) or the
value shown at the POTENTIAL display (39) as a measure for the membrane potential, since
these values include also the additional voltage generated by SERIES RESISTANCE COMP.
for compensation of the voltage drop at the SERIES RESISTANCE.
For instance, when for calculating I/V curves do not use the potential values recorded from
the POTENTIAL OUTPUT (mV) BNC connector (27). Use the COMMAND potential
instead.
(42) MODE OF OPERATION switch and LEDs
Switch for selecting the MODE OF OPERATION
VC x10: the amplifier operates in Voltage (Patch) Clamp mode. The COMMAND
POTENTIAL is enhanced by a factor of ten.
VC: the amplifier operates in Voltage (Patch) Clamp mode
Important: VC modes do not function properly with sharp microelectrodes, i.e. electrodes
with more than 10 M resistance.
OFF: all inputs of the amplifier are switched OFF, and the amplifier is set to CC mode.
R
EL
test and potential measurement work
CC: the amplifier operates in Current Clamp mode
CC x10: the amplifier operates in Current Clamp mode. The STIMULUS amplitude is
enhanced by a factor of ten.
EXT: the amplifier is set to CC mode. RS, REL test, BR, x10 or VC / CC modes can be
selected by application of a TTL HIGH (+5 V) signal to the respective BNC at the
rear panel (see also chapter 4).
The MODE OF OPERATION that is currently activated, is indicated by the respective LED
above the switch.
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BRIDGE unit
The BRIDGE unit consist of (12) BRIDGE BALANCE M potentiometer, (43)
BRIDGE MODE LED and (44) BRIDGE MODE switch.
(12) BRIDGE BALANCE M potentiometer
Potentiometer for balancing the BRIDGE circuit; 10 M / turn, range: 0 to 100
M. The BRIDGE must be balanced correctly before using the series resistance
compensation in VC mode, because this measure is taken as value for the series
resistance compensation (see also 43 and 44).
(43) BRIDGE MODE LED
LED indicating that the BRIDGE circuit is active.
(44) BRIDGE MODE switch
Switch for activating the BRIDGE circuit. The active circuit is indicated by 43.
(45) CURRENT (nA) display
Display for the current at the electrode in XX.XX nA, i.e. 10.00 is 10 nA
(19.99 nA max.)
Note: If the ELC-03XS is equipped with the optional SEAL TEST circuit, the value of the
SEAL resistance is also shown at the CURRENT display in XX.XX G (19.99 G max.).
(46) CURRENT OUTPUT SENSITIVITY (V/nA) switch
7-position switch for selecting the amplification of the current output signal in
V/nA; range 0.1 V/nA to 10 V/nA.
(49) BIAS (CC) potentiometer
Ten-turn potentiometer for BIAS current cancellation of the headstage; (ten-turn
potentiometer, symmetrical, i.e. 0 pA = 5 on the dial), range: ±100 pA.
(50) OFFSET potentiometer
Ten-turn potentiometer for OFFSET cancellation of the electrode; (ten-turn
potentiometer, symmetrical, i.e. 0 mV = 5 on the dial), range: ±100 mV.
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(51) CAPACITY COMP. (CC) potentiometer
Ten-turn potentiometer for the capacity compensation of the electrode in CC mode
(ten turn potentiometer, clockwise, range: 0-30 pF).
4. Description of the Rear Panel
Figure 3: ELC-03XS rear panel view
MONITORING OUTPUT connectors
(1) FILTER CURRENT connector
BNC connector providing a voltage monitoring the position of the CURRENT FILTER
switch (-7 V to +8 V, 1V/STEP).
(2) CURRENT SENSITIVITY connector
BNC connector providing a voltage monitoring the position of the CURRENT OUTPUT
SENSITIVITY switch (+1 V to +7 V, 1V/STEP).
(3) LP FILTER POTENTIAL connector
BNC connector providing a voltage monitoring the position of the POTENTIAL
LOWPASS FILTER switch (-8 V to +7 V, 1V/STEP).
(4) HP FILTER POTENTIAL connector
BNC connector providing a voltage monitoring the position of the POTENTIAL
HIGHPASS FILTER switch (-8 V to +7 V, 1V/STEP).
(5) POTENTIAL SENSITIVITY connector
BNC connector providing a voltage monitoring the position of the POTENTIAL OUTPUT
GAIN switch (+1 V to +7 V, 1V/STEP).
RANGE connector
(6) HEADSTAGE connector
BNC connector for remote control of a switchable headstage (ELC-SWI, optional). A TTL
HI (+5 V) signal switches the feedback resistance in the switchable headstage from 1 M
to 10 M or from 1 M to 100 M. To which resistor is indicated by a box to the left.
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MODE SELECT connectors (see also chapter 3.4)
All MODEs OF OPERATION can be selected by TTL signals connected to the rear panel
(see below), if the MODE OF OPERATION switch (42, Figure 1) is in EXT position. This
is very convenient when switching often between electroporation and recording, because
this can be done automatically by the data acquisition system using TTL signals.
(7) RS connector
BNC connector for remote control of the series resistance compensation circuit. A TTL HI
(+5 V) signal is connected here to activate the series resistance compensation circuit
remotely.
(8) REL connector
BNC connector for remote control of the electrode resistance test. A TTL HI (+5 V) signal
can be connected here to select the electrode resistance test remotely.
(9) BR connector
BNC connector for remote control of the bridge mode. A TTL HI (+5 V) signal can be
connected here to select the bridge mode remotely.
(10) OFF connector
BNC connector to switch the ELC-03XS in OFF mode remotely with a TTL HI (+5 V)
signal.
(11) x10 MODE connector
BNC connector to switch the ELC-03XS to x10 mode of operation remotely (TTL HI
signal). Dependent on the signal level at 12, this is CCx10 (signal at 12 = LOW) or VCx10
(signal at 12 = HI).
(12) VC / CC connector
BNC connector for remote control of the VC / CC mode of operation. A TTL signal can be
connected here to select the mode of operation remotely (HI = VC, LO = CC).
(13) GROUND connector
Banana plug providing internal ground (see below).
(14) CHASSIS connector
Banana plug providing mains ground (see below).
(15) FUSE holder
Holder for the line fuse. For changing the fuse rotate the holder counter-clockwise using a
screw driver.
(16) LINE SELECT switch
Switch for selecting the line voltage. Switch to the right for 230 V AC, to the left for 115 V
AC. The selected voltage is indicated on the switch.
Caution: Before turning on the instrument, make sure that the correct line voltage is selected.
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(17) Mains connector
Plug socket for the mains power-plug.
Important: Check line voltage before connecting the ELC amplifier to power. Always use a
three-wire line cord and a mains power-plug with a protection contact connected to ground.
Disconnect mains power-plug when replacing the fuse or changing line voltage. Replace fuse
only by appropriate specified type (one spare fuse is supplied). Before opening the cabinet
unplug the instrument.
SEAL TEST SIGNALS connectors (optional)
The SEAL resistance is determined similar to the REL test.
±10 mV square pulses with 15 Hz are applied to the pipette and the resulting current is
measured. The resistance is calculated according to Ohm’s law and is indicated on the
CURRENT DISPLAY. The maximum SEAL resistance that can be displayed is 19.99 G.
The value of the SEAL resistance is also monitored at 20 (see below).
(18) COMMAND MONITOR (TTL) connector
BNC connector providing a TTL (+5V) signal synchronous to the ±10 mV test pulses.
(19) SEAL TEST INPUT (TTL) connector
Starts seal test remotely (see also 36, Figure 1).
(20) OUTPUT SEAL 0.1V / G connector
BNC connector monitoring the value of the SEAL resistance; scaling 100 mV / G.
Grounding
ELC instruments have two ground systems:
1. the internal ground (called internal GROUND) represents the zero level for the
recording electronics and is connected to the recording chamber and the BNC
input/output sockets
2. mains ground (CHASSIS) is connected to the 19” cabinet and through the power cable
to the protection contact of the power outlet.
Both grounds are provided at the rear panel:
GROUND (black socket): internal system ground
CHASSIS (green/yellow socket): mains ground, 19” cabinet
All ELC systems have a high quality toroid transformer in order to minimize stray fields. In
spite of this, noise problems could occur if other mains-operated instruments are used in the
same setup. The internal system ground (GROUND sockets) should be connected to only one
point on the measuring ground. Multiple grounding should be avoided, i.e. all ground points
should originate from a central point to avoid ground loops.
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5. Setting up the ELC-03XS
The following steps should help you set up the ELC-03XS correctly. Always adhere to the
appropriate safety measures (see chapter 1).
After unpacking, the ELC-03XS is attached to the setup by assembling the electrical
connections. It is assumed that first a cell model will be attached.
Electrical connections
Turn POWER off.
Plug the power cord of the instrument into a grounded outlet.
Connect the headstage to the HEADSTAGE connector (#1, Figure 1) at the ELC-
03XS.
Connect a cell model (see chapter 6). Connect a digital/analog timing unit or a
stimulation device to STIMULUS INPUT or to GATE TTL if you intend to use a
gated stimulus.
Connect a store oscilloscope or a data acquisition system to the POTENTIAL
OUTPUT and to the CURRENT OUTPUT triggered from the stimulation device. Set
the desired gain at the POTENTIAL OUTPUT GAIN switch (#34, Figure 1) and the
CURRENT OUTPUT SENSITIVITY switch (#46, Figure 1).
Before using the ELC-03XS always make the basic settings to avoid oscillations.
Basic settings
Turn all controls to low values (less than 1) and the OFFSET and BIAS controls in the
range of 5 (zero position, see chapter 3.4).
Set the MODE OF OPERATION switch (#42, Figure 1) to CC.
Turn POWER switch on.
Now the ELC-03XS is ready for an initial check with the cell model.
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6. Passive Cell Model
The cell model is designed to be used to check the function of the instrument either
1. just after unpacking to see whether the instrument has been damaged during transport or
2. to train personnel in using the instrument or
3. in case of trouble to check which part of the setup does not work correctly e.g. to find out
whether the amplifier is broken or if something is wrong with the electrodes or holders
etc.
Figure 4: ELC-MOD passive cell model
The passive cell model consists only of passive elements, i.e. resistors that simulate the
resistance of the cell membrane and the electrodes, and capacitances that simulate the
capacitance of the cell membrane. A switch allows simulation of two different cell types: a
cell with 50 M and 22 pF (CELL 1, represents an astrocyte like cell) or a cell with 200 M
membrane resistance and 100 pF membrane capacitance (CELL 2, represents an neuron like
cell), or. Electrode immersed into the bath or SEAL formation can be mimicked as well. The
headstage of the amplifier can be connected to one of two different types of electrodes (see
below).
6.1. Cell Model Description
Figure 5: ELC-MOD passive cell model
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1, 3: connectors for the headstage, 1: electrode resistance: 50 M, 3: electrode resistance:
10 M
2: GND ground connector, to be connected to GND jack of the headstage
4: CELL: switch for cell membrane representing a membrane of either 50 M and 22 pF
(CELL 1) or 200 M and 100 pF (CELL 2).
5: In GROUND (lower) position the electrodes are connected to ground via a 1 k
resistor. In SEAL (upper) position are connected to a 1 G resistor simulating the
formation of a GIGASEAL with a patch electrode.
Figure 6: Schematic diagram of the passive cell model
6.2. Connections and Operation
Checking the configuration
Turn POWER switch of the amplifier off.
a) For simulation of an experiment using a suction electrode
Connect the BNC jack labeled 10M of the cell model to the BNC connector P
EL
of the
headstage.
b) For simulation of an experiment using a sharp electrode
Connect the BNC jack labeled 50M of the cell model to the BNC connector P
EL
at the
headstage.
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For a) and b)
Connect GND of the cell model to GND of the headstage.
Important: When using a differential headstage (optional) the REF connector at the
headstage must not be left open. It must be connected to ground.
Simulation of electrode in the bath
Set switch #4, Figure 5 to the lower position.
Set switch #5, Figure 5 to GROUND position. The 1 k resistor simulates the resistance
of the bath solution. This can be used to train cancellation of offsets, using the bridge
balance and using the capacity compensation.
Simulation of SEAL formation
Set switch #4, Figure 5 to the lower position.
Set switch #5, Figure 5 to SEAL position. The 1 G resistor simulates the SEAL
resistance when forming a GIGASEAL in patch clamp experiments.
Simulation of intracellular recording
Intracellular recordings can be mimicked with one of two cells with different properties. Use
the 50 M electrode connector (#1, Figure 5) for an experiment with sharp electrodes or the
10 M electrode connector (#3, Figure 5) for simulating an experiment with patch electrodes.
Switch the CELL membrane switch (see #4, Figure 5) to the desired position (CELL 1 or
CELL 2).
Turn all controls at the amplifier to low values (less than 1) and the OFFSET in the range
of 5 (zero position) and the OSCILLATION SHUTOFF in the DISABLED position.
Turn POWER switch of the amplifier on.
Now you can adjust the amplifier (see below) and apply test pulses to the cell model. The
upper position the CELL membrane switch (CELL 1) simulates a cell with a resistance of
50 M and a capacitance of 22 pF. In the lower position (CELL 2) a cell membrane with
200 M and 100 pF is simulated.
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7. Headstage
The ELC-03XS comes with a headstage for connecting suction electrodes for loose-patch
clamp or whole cell recordings and / or stimulation or electroporation, respectively or sharp
electrodes for intracellular or extracellular recordings. The use of metal electrodes is possible
as well.
A differential (miniature) headstage (see Optional accessories in chapter 3.1) for
measurements in vivo is also available (see also chapter 8.1 and contact npi for details).
7.1. Headstage Elements
Figure 7: ELC-03XS headstage
P
EL
BNC connector for the electrode holder
REF Connector for the reference electrode (differential headstage only)
GND Ground connector
TYPE Type of amplifier, BA Bridge amplifier, ELC ELC amplifier
R FB Value of feedback resistor, 10M: 10 M, 100 M: 100 M
Headstage cable to amplifier
Mounting plate
The electrode filled with electrolyte is inserted into an electrode holder (optional) that fits into
the BNC connector of the headstage or into an electrode holder adapter. The electrical
connection between the electrolyte and the headstage is established using a carefully
chlorinated silver wire. Chlorinating of the silver wire is very important since contact of silver
to the electrolyte leads to electrochemical potentials causing varying offset potentials at the
electrode, deterioration of the voltage measurement etc. (for details see Kettenmann and
Grantyn (1992)). For optimal chlorinating of sliver wires an automated chlorinating apparatus
(ACl-01) is available (contact npi for details).
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GND provides system ground and is linked to the bath via an agar-bridge or a Ag-AgCl
pellet. The headstage is attached to the amplifier with the headstage cable and an 8-pole
connector. The headstage can be mounted directly to a micromanipulator using the mounting
plate, a dove tail or a holding bar.
Important: The shield of the BNC connector is linked to the driven shield output and must not
be connected to ground. The headstage enclosure is grounded.
Caution: Please always adhere to the appropriate safety precautions (see chapter 1). Please
turn power off when connecting or disconnecting the headstage from the HEADSTAGE
connector!
7.2. Headstage Bias Current Adjustment
Caution: It is important that this tuning procedure is performed ONLY after a warm-up
period of at least 30 minutes!
The ELC-03XS is equipped with a voltage-to-current converter with a very high output
impedance which is connected to the recording electrode. The zero current of this unit is
tuned with the BIAS current potentiometer (#49, Figure 1).
The tuning procedure should be performed regularly (at least once a month) since the BIAS
current changes over time.
The tuning procedure is performed using high-value resistors and/or a cell model. It cannot be
performed with an electrode, since there are always unknown potentials involved (tip
potential, junction potentials).
Disconnected all input signals (except the headstage). Put the CURRENT (nA) switch
(10, Figure 1) to position 0. Set the operation mode to CC.
Set the BIAS control potentiometer to 5 (zero position).
Connect the P
EL
connector of the headstage to ground.
Note: This cannot be done with the cell model. Please use a wire to connect the input of
the BNC connector on the headstage to GND of the headstage. Do not use the shield of
the BNC connector since it is connected to driven shield.
Tune the OFFSET to zero using the OFFSET control.
Remember: The zero position of the OFFSET control is at 5!!
Remove the wire and attach the cell model or a resistor with a value of about 5 M across
the same connection.
The value displayed at the POTENTIAL DISPLAY is related to the BIAS current of the
headstage according to Ohm's Law. Cancel this voltage by tuning the headstage BIAS
current potentiometer until the POTENTIAL DISPLAY shows 000.
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7.3. Headstage with x1 / x100 current range (option)
Optionally, the ELC-03XS amplifier has a switch or TTL selectable extended current range of
x100. This is realized by reduction of the headstage feedback resistance from 100 M to 1
M. It enables the amplifier to generate higher currents and voltages with low resistance
electrodes, e.g. for electroporation, both in current clamp and in voltage clamp mode.
Figure 8: ELC-XS with switchable headstage, front panel view
The extended current range can be set at the front panel of the ELC-03XS with an additional
switch labeled RANGE x100 or with a TTL signal applied to the x100 INPUT (TTL) BNC
connector at the rear panel of the amplifier (see also chapter 4). If the switch is set to ON the
current range is extended to 12 µA. This mode is indicated at the headstage by an additional
LED (x100 ON, see Figure 10).
Important: In the VC x10 range which is usually used for generating high amplitude voltage
pulses suitable for electroporation of large molecules the x100 current range is automatically
activated allowing for instance 10 V pulses with 1 M electrodes.
Note: All basic operations and settings (e.g. bridge balance or BIAS adjustment) should be
performed in standard x1 mode, i.e. RANGE x100 switch set to OFF, because these are also
affected by the RANGE x100 mode!
Exception: The electrode resistance test works well in x100 mode!!
If the ELC-03XS operates in x100 high current mode, all current related signals have to be
corrected by the factor of hundred:
- CURRENT OUTPUT SENSITIVITY (V / nA): 0.001, 0.002, 0.005, 0.01, 0.02, 0.05, 0.1
- STIMULUS INPUT: 100 nA / V (CC x1) or 1µA / V (CC x10)
- Holding CURRENT / gated CURRENT: 0 to 999 nA (CC x1) or 9.99 µA (CC x10)
- CURRENT FROM HEADSTAGE: 0.001 V / nA
- CURRENT DISPLAY: shows always nA, scaling is adapted automatically.
Figure 9: Current range switch
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Figure 10: ELC x1 / x100 switchable headstage
Important: Even if the ELC-03XS is modified for the x1 / x100 switchable headstage it can
be used as a standard amplifier with a standard, i.e. not switchable headstage. In this case the
amplifier will recognize that a standard headstage is connected, and the additional current
switch at the front panel and the TTL input at the rear panel are automatically disabled.
8. Introduction into Experiments
The ELC-03XS is capable to perform several types of experiments that are briefly introduced
in the following with special focus on loose-patch stimulation and recording. It is assumed
that the capacity of the electrode is compensated, the offset of the electrode is cancelled and,
for intracellular recordings in BRIDGE mode, electrode artifact is eliminated using the bridge
balance circuit. Particularly, when working with charged substances and approaching the cell
in VC mode, the current offset is cancelled using the PIPETTE HOLD POTENTIAL
potentiometer (33, Figure 1).
8.1. Recordings with the Differential Headstage (optional)
Extracellular measurements are mostly done in slices or in vivo, in noisy environments, where
distortions of the recorded signal caused by other instruments and the animal itself are very
common. Additionally, extracellular signals are very small and have to be amplified
enormously. The drawback is that noise is amplified as well. Therefore, the headstage of the
ELC 03X can be equipped with a differential input that minimizes noise pick-up. Differential
means, that the signal for the amplifier is the difference between the positive (+) (PEL) and
negative (-) (REF.) input of the headstage. This results in canceling of all common mode
signals (i.e. which both electrodes record, e.g. noise). For differential measurements, both
inputs of the headstage (REF. and P
EL
) are connected to microelectrodes using cables with
grounded enclosure or electrode holders. P
EL
is connected to the measuring electrode and
REF. to the reference electrode. The experimental chamber is grounded by an Ag-AgCl pellet
(or an AGAR bridge) connected to GND of the headstage (see Figure 11).
If differential measurement is not required (single-ended measurement configuration, see
Figure 11), the REF input must be connected to ground (GND). The amplifier is in an
undefined state, if the REF is left open, and can go into saturation making reliable
measurements impossible (for more details see Lalley et al., 1999).
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Figure 11: headstage connections, A: differential measurement, B: single-ended measurement
8.2. Extracellular Voltage Measurement
Extracellular measurements are usually done in the loose-patch configuration or with special
metal microelectrodes. Recordings with extracellular metal electrodes is simple. The electrode
is advanced into the region where the recordings will be made using a micromanipulator and
the signals are filtered and amplified (see chapter 5 in Lalley et al., 1999 for details) as
required. For loose patch recording the basic procedure is the following (Barbour & Isope,
2000, Nunemaker et al, 2003):
Approach the cell in VC mode and apply square voltage pulses to the electrode.
Contact the cell and establish the loose-patch.
Set the MODE OF OPERATION switch to OFF.
Set the required amplification of the POTENTIAL OUTPUT.
Set the HIGHPASS FILTER to the desired corner frequency, e.g. 0.3 Hz.
Set the LOWPASS FILTER to the desired corner frequency, e.g. 3 kHz.
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8.3. Extracellular Stimulation and Electroporation
Cells can be stimulated using current or voltage signals.
Stimulation with Current
Approach the cell in VC mode and apply square voltage pulses to the electrode.
Contact the cell, establish the loose-patch and disconnect the voltage signal from the
COMMAND INPUT :10 mV connector.
Set the MODE OF OPERATION switch to CC or CCx10.
Set the HOLDING CURRENT to zero.
For stimulation:
Apply the stimulus signal to the STIMULUS INPUT 1 nA/V connector.
or
Adjust the stimulus amplitude with the HOLDING CURRENT potentiometer and set the
stimulus polarity using the switch aside. Gate the preset stimulus with a TTL signal linked
to the GATE TTL BNC connector.
Electroporation with Current
Electroporation can be done using the stimulation procedure, but usually the applied current is
much higher and the stimulus duration is shorter. Therefore, most electroporation experiments
are done in CCx10 mode.
Stimulation with Voltage
Approach the cell in VC mode and apply square voltage pulses to the electrode.
Contact the cell and establish the loose-patch.
For stimulation apply a voltage signal of the required amplitude and duration to the
COMMAND INPUT :10 mV connector.
or
Adjust the stimulus amplitude with the HOLDING POTENTIAL potentiometer and set
the stimulus polarity using the switch aside. Gate the preset stimulus with a TTL signal
linked to the GATE TTL BNC connector.
Electroporation with Voltage
Electroporation can be done using the stimulation procedure, but usually the applied voltage
is much higher and the stimulus duration is shorter. For stimuli with large amplitudes set the
MODE OF OPERATION switch to VCx10 enabling electroporation with up to 12 V.
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8.4. Intracellular Recording
Intracellular recordings can be performed in the whole-cell patch configuration and with sharp
microelectrodes.
Note: VC mode does not function properly with sharp microelectrodes, i.e. electrodes with
more than 10 M resistance.
Current Clamp Recording
The ELC-03XS can be used like a standard bridge amplifier.
Set the MODE OF OPERATION switch to CC and the BRIDGE MODE switch to the
upper position. The BRIDGE MODE LED lights up.
Compensate the electrode artifact using the BRIDGE BALANCE potentiometer.
After impaling the cell readjust the bridge.
If needed set an appropriate holding current using the HOLDING CURRENT
potentiometer and the HOLDING CURRENT polarity switch.
Apply stimuli to the cell using the STIMULUS INPUT 1 nA/V BNC connector.
Voltage Clamp Recording
The ELC-03XS can also be used like a whole-cell patch-clamp amplifier.
Approach the cell in VC mode and apply square voltage pulses to the electrode.
Contact the cell, set a holding potential using the HOLDING POTENTIAL potentiometer
and establish the whole-cell patch clamp configuration.
Set the amplifier to BR mode and adjust the BRIDGE. Only if the BRIDGE is adjusted
correctly the SERIES RESISTANCE COMPENSATION works with the correct value.
Switch to VC mode.
If necessary, adjust the shape of the voltage signal using the CAPACITY
COMPENSATION (VC) potentiometer (#22 and #27, Figure 1) and compensate for the
SERIES RESISTANCE.
Apply stimuli to the cell using the COMMAND INPUT :10 mV BNC connector.
Note: CAPACITY COMPENSATION works only for the electrode capacity, not for the
capacity of the cell membrane. Therefore, capacitive transients are always present when
square shaped pulses are applied in VC mode.
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9. Literature
General Recording Methods and Voltage Clamp Technique
Dietzel, I. D., Bruns, D., Polder, H. R. and Lux, H. D. (1992). Voltage Clamp Recording,
in Kettenmann, H. and R. Grantyn (eds.) Practical Electrophysiological Methods, WileyLiss, NY.
Lalley, P. M., Moschovakis, A. K. and Windhorst, U. (1999). Electrical Activity of
Individual Neurons in Situ: Extra- and Intracellular Recording, in: U. Windhorst and H.
Johansson (eds.) Modern Techniques in Neuroscience Research, Springer, Berlin, New
York
Ogden DC (1994) Microelectrode Techniques. The Plymouth Workshop Handbook,
Second Edition, The Company of Biologists Limited, Cambridge
Polder, H. R., M. Weskamp, K. Linz and R. Meyer (2004) Voltage-Clamp and Patch-
Clamp Techniques, Chapter 3.4, pp. 272-323 in: Dhein, Stefan; Mohr, Friedrich Wilhelm;
Delmar, Mario (Eds.) Practical Methods in Cardiovascular Research, Springer, Berlin,
Heidelberg and New York 2004.
Windhorst, U. and H. Johansson (eds.) Modern Techniques in Neuroscience Research,
Springer, Berlin, Heidelberg, New York.
Juxtasomal Filling, Loose-Patch Techniques (General)
Auger, C., & Marty, A. (2000). Topical Review: Quantal currents at singlesite central
synapses. J Physiol. 526.1, 3-11.
Barbour, B., & Isope, P. (2000). Combining loose cell-attached stimulation and recording.
J Neurosci.Methods. 103, 199–208.
Bureau, I., Shepherd, G. M. G. & Svoboda, K. (2004). Precise Development of Functional
and Anatomical Columns in the Neocortex. Neuron, 42, 789-801.
Joshi, S. & Hawken, M. J. (2006). Loose-patch-juxtacellular recording in vivo-A method
for functional characterization and labeling of neurons in macaque V1. J
Neurosci.Methods. 156, 37-49.
Khaliq, Z. M., & Raman, I. M. (2005). Axonal Propagation of Simple and Complex Spikes
in Cerebellar Purkinje Neurons. J Neurosci. 25, 454-463.
Klausberger, T., Marton, L. F., Baude, A., Roberts, J. D., Magill, P. J. & Somogyi, P.
(2004). Spike timing of dendrite-targeting bistratified cells during hippocampal network
oscillations in vivo. Nature Neuroscience 7, 41-47.
Nunemaker, C. S., DeFazio, R. A., & Moenter, S. M. (2003). A targeted extracellular
approach for recording long-term firing patterns of excitable cells: a practical guide.
Biol.Proced.Online. 5, 53-62. www.biologicalprocedures.com
Pinault, D. (1996). A novel single-cell staining procedure performed in vivo under electro-
physiological control: morpho-functional features of juxtacellularly labeled thalamic cells
and other central neurons with biocytin or Neurobiotin. J Neurosci.Methods. 65, 113-136.
Rathenberg, J., Nevian, T. & Witzemann, V. (2003). High-efficiency transfection of
individual neurons using modified electrophysiology techniques. J Neurosci.Methods. 126,
91-98.
Roberts, W. M., & Almers, W. (1992). Patch Voltage Clamping with Low-Resistance
Seals: Loose Patch Clamp. In: Rudy, B. & Iversen, L. E. (eds.). Ion Channels. Methods in
Enzymology 207, Academic Press San Diego.
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Strickholm, A. (1961). Impedance of a Small Electrically Isolated Area of the Muscle Cell
Surface. J Gen.Physiol. 44, 1073-1088.
Tracer injection (juxtasomal filling) and extracellular recording using ELC amplifiers
Bruno, R. M. & Sakmann, B. (2006). Cortex is driven by weak but synchronously active
thalamocortical synapses. Science. 312, 1622-1627.
Hoshi, H., Liu, W. L., Massey, S. C., & Mills, S. L. (2009). ON inputs to the OFF layer:
bipolar cells that break the stratification rules of the retina. J Neurosci. 29, 8875-8883.
Fuentealba, P., Begum, R., Capogna, M., Jinno, S., Marton, L. F., Csicsvari, J., Thomson,
A., Somogyi, P., & Klausberger, T. (2008). Ivy cells: a population of nitric-oxideproducing, slow-spiking GABAergic neurons and their involvement in hippocampal
network activity. Neuron. 57, 917-929.
Fuentealba, P., Tomioka, R., Dalezios, Y., Marton, L. F., Studer, M., Rockland, K.,
Klausberger, T., & Somogyi, P. (2008). Rhythmically active enkephalin-expressing
GABAergic cells in the CA1 area of the hippocampus project to the subiculum and
preferentially innervate interneurons. Journal of Neuroscience 28, 10017-10022.
Transfection using ELC amplifiers
Stan, A., Pielarski, K. N., Brigadski, T., Wittenmayer, N., Fedorchenko, O., Gohla, A.,
Lessmann, V., Dresbach, T., & Gottmann, K. (2010). Essential cooperation of N-cadherin
and neuroligin-1 in the transsynaptic control of vesicle accumulation.
Proc.Natl.Acad.Sci.U.S.A. 107, 11116-11121.
Extracellular recording using ELC amplifiers
Geis, C., Weishaupt, A., Hallermann, S., Grunewald, B., Wessig, C., Wultsch, T., Reif, A.,
Byts, N., Beck, M., Jablonka, S., Boettger, M. K., Uceyler, N., Fouquet, W., Gerlach, M.,
Meinck, H. M., Siren, A. L., Sigrist, S. J., Toyka, K. V., Heckmann, M., & Sommer, C.
(2010). Stiff person syndrome-associated autoantibodies to amphiphysin mediate reduced
GABAergic inhibition. Brain., doi: 10.1093/brain/awq253
Pangrsic, T., Lasarow, L., Reuter, K., Takago, H., Schwander, M., Riedel, D., Frank, T.,
Tarantino, L. M., Bailey, J. S., Strenzke, N., Brose, N., Muller, U., Reisinger, E., & Moser,
T. (2010). Hearing requires otoferlin-dependent efficient replenishment of synaptic
vesicles in hair cells. Nat.Neurosci. 13, 869-876.
Strenzke, N., Chanda, S., Kopp-Scheinpflug, C., Khimich, D., Reim, K., Bulankina, A. V.,
Neef, A., Wolf, F., Brose, N., Xu-Friedman, M. A., & Moser, T. (2009). Complexin-I is
required for high-fidelity transmission at the endbulb of held auditory synapse. Journal of
Neuroscience 29, 7991-8004.
Neef, J., Gehrt, A., Bulankina, A. V., Meyer, A. C., Riedel, D., Gregg, R. G., Strenzke, N.,
& Moser, T. (2009). The Ca2+ channel subunit ß2 regulates Ca2+ channel abundance and
function in inner hair cells and is required for hearing. Journal of Neuroscience 29, 10730-
10740.
ELC-03XS User Manual
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10. Technical Data
Headstage:
Input voltage range: ±12 V
Operating voltage: ±15 V
Enclosure: Size: 23 x 70 x 26 mm, grounded
Mounting plate: Size: 70 mm x 50 mm
on request
Holding bar: length 150 mm, 8 mm
Electrode connector: BNC with driven shield
Ground connector: 2.4 mm connector
Input resistance (CC): >1013 (internally adjustable)
Current range: ±120 nA max. (100 M feedback, x1 range)
±1.2 µA max(10 M feedback, x10 range)
±12 µA max(1 M feedback, x100 range)
Electrode parameter controls:
OFFSET: range ±100 mV, ten-turn control
PIPETTE HOLD POTENTIAL (in VC): range ±100 mV, ten-turn control
CAPACITY COMPENSATION: range 0 – 30 pF, ten-turn control
BIAS: range ±100 pA, ten-turn control
Bridge balance:
0-100 M adjustable with ten-turn control
Electrode resistance test:
Sensitivity 1 mV / M application of square current pulses ±1 nA
Display: 3 ½ digit, XXX M, activated by key switch
(same as POTENTIAL display)
Bandwidth and speed response (CC mode, optimal capacity compensation):
Full power bandwidth (REL = 0 M): >30 kHz, rise time (10% - 90%)
<10 µs (REL = 100 M)
<5 µs (REL = 10 M)
Outputs:
Output impedance: 50
Max. voltage: ±12 V
Current output: BNC connector, sensitivity 0.1…10 V/nA,
Current output sensitivity: Rotary switch, 0.1, 0.2, 0.5, 1, 2, 5, 10 V/nA
Current display: 3 ½ digits, XX.XX nA, resolution 10 pA
Current LP filter: 4-pole BESSEL filter (other options available)
attenuation: -24 dB/octave,
corner frequencies (Hz): 20, 50, 100, 200, 300, 500, 700, 1k, 1,3k, 2k, 3k,
5k, 8k, 10k, 13k, 20k
Potential output x1: BNC connector, sensitivity 1 V/V
Potential output: BNC connector, sensitivity 10…1k V/V
Potential output gain: Rotary switch, 10, 20, 50, 100, 200, 500, 1k
Potential output resolution in AC: 50 µV
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Potential LP filter: 4-pole BESSEL filter (other options available)
attenuation: -24 dB/octave,
corner frequencies (Hz): 20, 50, 100, 200, 300, 500, 700, 1k, 1,3k, 2k, 3k,
5k, 8k, 10k, 13k, 20k
Potential HP filter: 1-pole filter, (other options available)
attenuation: -6 dB/octave
corner frequencies (Hz): DC, 0.1, 0.3, 0.5, 1, 3, 5, 10, 30, 50, 100, 300,
500, 800, 1k, 3k
Telegraph potential LP filter -8…+7 V, 1V/step
Telegraph potential HP filter -8…+7 V, 1V/step
Telegraph current filter -8…+7 V, 1V/step
Telegraph potential output sensitivity +1…+7 V, 1 V/ step
Telegraph current output sensitivity +1…+7 V, 1 V/ step
Digital displays:
Display mV/M 3 ½ digits, XXXX mV or XXX M
Display current 3 ½ digits, XX.XX nA
Inputs:
Input impedance analog 100 k
Input range ±12 V
Input impedance digital (TTL) 10 k
Input range TTL 0-5 V
Current stimulus input CC via BNC connectors, sensitivity 1 nA / V
Current stimulus input CCx10 via BNC connectors, sensitivity 10 nA / V
Step gate input via BNC connector (TTL)
Gated stimulus CC with ten-turn control of holding current
resolution: 10 pA, range: ±10 nA
Gated stimulus CCx10 with ten-turn control of holding current
resolution: 100 pA, range: ±100 nA
Polarity selectable with toggle switch
Voltage command input VC via BNC connectors, sensitivity: 10 mV
Voltage command input VCx10 via BNC connectors, sensitivity: 1 mV
Step gate input via BNC connector (TTL)
Gated stimulus VC with ten-turn control of holding potential
resolution: 1 mV, range: ±1 V
Gated stimulus VCx10 with ten-turn control of holding potential
resolution: 10 mV, range: ±10 V
Polarity selectable with toggle switch
Dimensions:
19” rackmount cabinet
19” (483 mm), 10” (250 mm), 3.5” (88 mm)
Power requirements:
115/230 V AC, 60/50 Hz, fuse 0.4/0.2 A, slow, 25 W
Weight: 4.5 kg
ELC-03XS User Manual
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version 2.2 page 37
Index
basic settings 22
bias current adjustment 27
BRIDGE unit 18
CAPACITY COMPENSATION (VC) unit
15
cell model 23
connections and operation 24
description 23
COMMAND set unit 13
CURRENT set unit 11
Differential Headstage (optional) 29
ELC-03XS
Components 6
Optional Accessories 6
electrical connections 22
Electroporation 31
Experiments 29
Extracellular Stimulation 31
Extracellular Voltage Measurement 30
Front Panel View 8
Grounding 21
headstage 26
elements 26
HEADSTAGE connector 9
Intracellular Recording 32
Literature 33
MODE SELECT connectors 20
MONITORING OUTPUT connectors 19
OSCILLATION SHUT OFF unit 13
PENETRATION / BUZZ unit 9
POTENTIAL OUTPUT FILTER 14
Rear Panel 19
Safety regulations 3
SEAL TEST 21
SERIES RESISTANCE
COMPENSATION unit 16
Technical Data 35
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