NPI SEC-03M User manual

OPERATING INSTRUCTIONS AND
SYSTEM DESCRIPTION OF THE
SEC-03M
SINGLE ELECTRODE CLAMP
AMPLIFIER MODULE FOR EPMS
SYSTEMS
VERSION 1.8
npi 2014
npi electronic GmbH, Bauhofring 16, D-71732 Tamm, Germany
Phone [phone]; Fax: [phone]
[email protected]; http://www.npielectronic.com

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Table of Contents
About this Manual ................................................................................................................... 4
1.Safety Regulations .............................................................................................................. 5
2.EPMS-07 Modular Plug-In System .................................................................................... 6
2.1.General System Description / Operation..................................................................... 6
2.2.EPMS-07 Housing....................................................................................................... 6
2.3.EPMS-E-07 Housing ................................................................................................... 6
2.4.PWR-03D .................................................................................................................... 6
2.5.System Grounding ....................................................................................................... 7
EPMS-07 ..................................................................................................................... 7
EPMS-E-07.................................................................................................................. 7
2.6.Technical Data............................................................................................................. 7
EPMS-07 ..................................................................................................................... 7
EPMS-E-07.................................................................................................................. 7
3.Introduction......................................................................................................................... 8
3.1.Why a Single Electrode Clamp?.................................................................................. 8
3.2.Principle of Operation ................................................................................................. 10
Major Advantages of the npi SEC System .................................................................. 12
3.3.Advantages of the Modular SEC-03M System ........................................................... 12
4.SEC-03M System ............................................................................................................... 13
4.1.SEC-03M Components................................................................................................ 13
4.2.Description of the Front Panel..................................................................................... 14
5.Headstages .......................................................................................................................... 19
5.1.Standard and low-noise (SEC-HSP) headstages ......................................................... 19
5.2.Low-noise headstage (SEC-HSP)................................................................................ 21
6.Setting up the SEC-03M System ........................................................................................ 22
7.Passive Cell Model ............................................................................................................. 23
7.1.Cell Model Description ............................................................................................... 23
7.2.Connections and Operation ......................................................................................... 24
7.3.Connections and Operation ......................................................................................... 25
8.Test and Tuning Procedures ............................................................................................... 27
8.1.Headstage Bias Current Adjustment............................................................................ 27
8.2.Electrode Selection...................................................................................................... 28
8.3.Offset Compensation ................................................................................................... 28
8.4.Bridge Balance (in BR mode) ..................................................................................... 29
8.5.Switching Frequency and Capacitance Compensation (in switched modes) .............. 31
Criteria for the selection of the switching frequency .................................................. 31
8.6.Capacity Compensation - Tuning Procedure............................................................... 33
First part: basic setting................................................................................................. 33
Second part: fine tuning............................................................................................... 38
8.7.Testing Operation Modes ............................................................................................ 39
Current Clamp (in BR- or discontinuous CC mode) ................................................... 39
Voltage Clamp............................................................................................................. 39
9.Sample Experiments ........................................................................................................... 41
9.1.Sample Experiment using a Sharp Microelectrode ..................................................... 41
9.2.Sample Experiment using a Suction Electrode............................................................ 44
10.Tuning VC Performance.............................................................................................. 46
General Considerations................................................................................................ 46
Tuning Procedure ........................................................................................................ 47
11.Trouble Shooting ......................................................................................................... 48
12.Appendix ..................................................................................................................... 49

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12.1.Theory of Operation .................................................................................................... 49
12.2.Speed of Response of SEC Single Electrode Clamps ................................................. 50
12.3.Tuning Procedures for VC Controllers........................................................................ 50
Practical Implications .................................................................................................. 51
13.Literature about npi single electrode clamp amplifiers ............................................... 53
13.1.Paper in Journals.......................................................................................................... 53
13.2.Books........................................................................................................................... 64
14.SEC-03M Specifications – Technical Data................................................................. 65
15.Index ............................................................................................................................ 68

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About this Manual
This manual should help to setup and use SEC systems correctly and to perform reliable
experiments.
If you are not familiar with the use of instruments for intracellular recording of electrical
signals please read the manual completely. The experienced user should read at least chapters
1, 4, 8 and 10.
Important: Please read chapter 1 carefully! It contains general information about the safety
regulations and how to handle highly sensitive electronic instruments.
Signs and conventions
In this manual all elements of the front panel are written in capital letters as they appear on
the front panel.
System components that are shipped in the standard configuration are marked with ,
optional components with . In some chapters the user is guided step by step through a
certain procedure. These steps are marked with .
Important information and special precautions are highlighted in gray.
Abbreviations
Cm: cell membrane capacitance
Cstray: electrode stray capacitance
GND: ground
Imax: maximal current
Ra: access resistance
Rm: cell membrane resistance
REL: electrode resistance
SwF: switching frequency
Cm: time constant of the cell membrane
VREL: potential drop at REL

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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.
1) 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.
2) AC MAINS CONNECTION: While working with the 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.
3) 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.
4) 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.
5) HANDLING: Please protect the device from moisture, heat, radiation and corrosive
chemicals.

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2. EPMS-07 Modular Plug-In System
2.1. General System Description / Operation
The npi EPMS-07 is a modular system for processing of bioelectrical signals in
electrophysiology. The system is housed in a 19” rackmount cabinet (3U) has room for up to
7 plug-in units. The plug-in units are connected to power by a bus at the rear panel.
The plug-in units must be kept in position by four screws (M 2,5 x 10). The screws are
important not only for mechanical stability but also for proper electrical connection to the
system housing. Free area must be protected with covers.
2.2. EPMS-07 Housing
The following items are shipped with the EPMS-07 housing:
EPMS-07 cabinet with built-in power supply
Mains cord
Fuse 2 A / 1 A, slow
Front covers
In order to avoid induction of electromagnetic noise the power supply unit, the power switch
and the fuse are located at the rear of the housing.
2.3. EPMS-E-07 Housing
The following items are shipped with the EPMS-E-07 housing:
EPMS-E-07 cabinet
External Power supply PWR-03D
Power cord (PWR-03D to EPMS-E-07)
Mains chord
Fuse 1.6 A / 0.8 A, slow
Front covers
The EPMS-E-07 housing is designed for low-noise operation, especially for extracellular and
multi channel amplifiers with plugged in filters. It operates with an external power supply to
minimize distortions of the signals caused by the power supply.
2.4. PWR-03D
The external power supply PWR-03D is capable of driving up to 3 EPMS-E housings. Each
housing is connected by a 6-pole cable from the one of the three connectors on the front panel
of the PWR-03D to the rear panel of the respective EPMS-E housing. (see Figure 1, Figure 3).
A POWER LED indicates that the PWR-03D is powered on (see Figure 1). Power switch,
voltage selector and fuse are located at the rear panel (see Figure 2).
Note: The chassis of the PWR-03D is connected to protective earth, and it provides protective
earth to the EPMS-E housing if connected.

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Figure 1: PWR-03D front panel view Figure 2: PWR-03D rear panel view
Note: This power supply is intended to be used with npi EPMS-E systems only.
2.5. System Grounding
EPMS-07
The 19" cabinet is grounded by the power cable through the ground pin of the mains
connector (= protective earth). In order to avoid ground loops the internal ground is isolated
from the protective earth. The internal ground is used on the BNC connectors or GROUND
plugs of the modules that are inserted into the EPMS-07 housing. The internal ground and
mains ground (= protective earth) can be connected by a wire using the ground plugs on the
rear panel of the instrument. It is not possible to predict whether measurements will be less or
more noisy with the internal ground and mains ground connected. We recommend that you try
both arrangements to determine the best configuration.
EPMS-E-07
The 19" cabinet is connected to the CHASSIS connector at the rear panel.
The CHASSIS is linked to protective earth as soon as the PWR-03D is
connected. It can be connected also to the SYSTEM GROUND (SIGNAL
GROUND) on the rear panel of the instrument (see Figure 3).
Important:: Always adhere to the appropriate safety measures.
Figure 3: Rear panel connectors of the EPMS-E-07
2.6. Technical Data
19” rackmount cabinet, for up to 7 plug-in units
Dimensions: 3U high (1U=1 3/4” = 44.45 mm), 254 mm deep
EPMS-07
Power supply: 115/230 V AC, 60/50 Hz, fuse 2 A / 1 A slow, 45-60 W
EPMS-E-07
External power supply (for EPMS-E): 115/230 V AC, 60/50 Hz, fuse 1.6/0.8 A, slow
Dimensions of external power supply: (W x D x H) 225 mm x 210 mm x 85 mm

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3. Introduction
npi electronic’s SEC (Single Electrode Clamp) systems are based on the newest
developments in the field of modern electronics and control theory (see also chapter 11).
These versatile current/voltage clamp amplifiers permit extremely rapid switching between
current injection and current-free recording of true intracellular potentials.
The use of modern high-voltage operational amplifiers and a new, improved method of
capacity compensation makes it possible to inject very short current pulses through high
resistance microelectrodes (up to 200 Mand more) and to record membrane potentials
accurately, i.e. without series resistance error, within the same cycle.
Although the system has been designed primarily to overcome the limitations related to the
use of high resistance microelectrodes in intracellular recordings, it can also be used to do
conventional whole-cell patch clamp recordings with suction electrodes or perforated patch
recordings. The whole-cell configuration allows to investigate even small dissociated or
cultured cells as well as cells in slice preparations in both current and voltage clamp mode,
while the intracellular medium is being controlled by the pipette solution.
3.1. Why a Single Electrode Clamp?
Voltage clamp techniques permit the analysis of ionic currents flowing through biological
membranes at preset membrane potentials. Under ideal conditions the recorded current is
directly related to the conductance changes in the membrane and thus gives an accurate
measure of the activity of ion channels and electrogenic pumps.
The membrane potential is generally kept at a preselected value (command or holding
potential). Ionic currents are then activated by sudden changes in potential (e.g. voltage-gated
ion channels), by transmitter release at synapses (e.g. electrical stimulation of fiber tracts in
brain slices) or by external application of an appropriate agonist. Sudden command potential
changes used to activate voltage-gated currents are especially challenging, because the
membrane will adopt the new potential value only after it’s capacitance (Cmin Figure 4and
Figure 5) has been charged. Therefore, the initial transient current following the voltage step
should be as large as possible to achieve rapid membrane charging. In conventional patch
clamp amplifiers, this requires a minimal resistance between the amplifier and the cell interior
– a simple consequence of Ohm’s law (U = R*I), i.e. for a given voltage difference (U),
the current (I) is inversely proportional to the resistance (R). In this context, R is the access or
series resistance (Rain Figure 4 and Figure 5) between the electrode and the cell interior. The
time constant for charging a cell is = REL*Cm.
Rais largely determined by certain electrode properties (mainly electrode resistance) and the
connection between the electrode and the cell. Typical Ravalues are between 5 to 10 M,
which results in a time constant of 0.5 to 1 ms for a cell with a membrane capacity of 100 pF,
i.e. the membrane needs roughly a millisecond to follow the command voltage step. Sharp
microelectrodes usually have much larger resistances (30 to 150 Mor even more).

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Figure 4: Model circuit for whole cell patch clamp recording using a suction electrode
Cm: membrane capacitance, Cstray: electrode stray capacitance, REL: electrode
resistance, Rm: membrane resistance
Figure 5: Model circuit for intracellular recording using a sharp electrode
Cm: membrane capacitance, Cstray: electrode stray capacitance, REL: electrode
resistance, Rm: membrane resistance

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Besides slowing the voltage response of the cell, Racan also cause additional adverse effects,
such as error in potential measurement. Ra, together with the membrane resistance (Rm) forms
a voltage divider (see Figure 4 and Figure 5). Current flowing from the amplifier to the
grounded bath of a cell preparation will cause a voltage drop at both, Raand Rm. If Ra<< Rm,
the majority of the voltage drop will develop at Rmand thus reflect a true membrane potential.
If, in an extreme case, Ra= Rm, the membrane potential will follow only one half of the
voltage command. In order to achieve a voltage error of less than 1% Ramust be more than
100 times smaller than Rm. This condition is not always easily to accomplish, especially if
recordings a performed from small cells. If sharp intracellular microelectrodes are used, it is
virtually impossible. If Rais not negligible, precise determination of the membrane potential
can be achieved only if no current flows across Raduring potential measurement. This is the
strategy employed in npi electronic’s SEC amplifier systems.
The SEC amplifiers inject current and record the potential in an alternating mode (switched
mode). Therefore, this technique is called discontinuous SEVC. This ensures that no current
passes through Raduring potential measurement and completely eliminates access resistance
artefacts.
After each injection of current, the potential gradient at the electrode tip decays much faster
than the potential added at the cell membrane during the same injection. The membrane
potential is measured after the potential difference across Rahas completely dropped (see
chapter 3.2). The discontinuous current and voltage signal are then smoothed and read at the
CURRENT OUTPUT and POTENTIAL OUTPUT connectors.
3.2. Principle of Operation
Figure 6: Model circuit of SEC systems