OPERATING INSTRUCTIONS AND SYSTEM DESCRIPTION OF
Contents
1. 4 microJECT 1 pipette connector 2 connector for EJECT pressure 3 connector for EXHAUST pressure optional 4 electrical connection Important In experiments with high pressures gt 1 bar it might be necessary to insert 2 seals in the pipette connector in order to prevent the pipette from moving in any direction Caution Pipettes that are not rigidly fixed in the pipette connector might be pressed out of the connector by high pressure Always adhere to the appropriate safety measures version 1 6 page 9 PDES 02TX User Manual 4 Operation Connect the gas source to the INPUT connector at the rear panel The gas source must be filtered by a 5 um filter element Important Do not connect sources of OXYGEN or COMBUSTIBLE gases The gas source MUST be dry and oil free Please use a degreaser and filter the gas source by a 5 um filter element Connect injection micropipette to the OUTPUT connector at the rear panel or external valve Connect the pressure tubing of the external valve to the OUT connector and the electrical connector to the VALVE connector at the rear panel Connect your ejection pipettes to the external valve using tubing and an ejection pipette holder Turn on pressure and adjust the pressure to the desired value using the pressure regulator Connect a digital timing unit to the INPUT BNC connector at the front panel For remote operation connect the remote control unit to the REMOTE CONTROL2. filtered dry and oil free gas source 5 bar max Note npi recommends an input pressure of 1 bar more than the maximum output pressure 5 CHASSIS This connector is linked to chassis 6 GROUND This connector is linked to the internal system ground 7 POWER IN connector Jack for connecting the external power supply Minimum 12 V 0 5 A for one channel and 12 V 1 A for two channel instruments inner pin positive version 1 6 page 7 PDES 02TX User Manual 3 External Valve External valve s for the PDES system are either mounted on a remote valve plate or in the microJECT micro valve pipette holder Remote Valve Plate The valves which are controlled by the PDES 02DX are located on the remote valve plate see Figure 3 Tubing and cable of the external valves are connected at the rear panel of the PDES system refer to chapter 2 3 Exhaust is usually not connected Figure 3 external valves on remote valve plate 1 electrical connection black 2 EJECT pressure tubing red 3 connector for ejection pipette 4 exhaust connector optional version 1 6 page 8 PDES 02TX User Manual microJECT micro Valve Pipette Holder The valve which is controlled by the PDES 02DX is located in the micro valve pipette holder called microJECT see Figure 4 Tubing and cable of the microJECT are connected at the rear panel of the PDES system refer to chapter 2 3 Exhaust is usually not connected Figure
3. 2 Pressure regulator Rotary knob for regulating the pressure of channel A 3 Manometer Display indicating pressure at channel A 4 ON EJECT LED Indicates the operational status of channel A red light channel has been turned on valve is closed green light valve is open 5 TIME BASE switch Switch to set the TIME BASE of channel A see also 7 The total ejection time is the reading of this potentiometer multiplied by the reading of the TIMER A potentiometer 7 6 Operation mode switch CONT The valve is continuously open TIMER The internal timer is used The timer can be started via a TTL HIGH signal at 9 or by pushing a pushbutton connected to 8 EXTERN The valve is open as long as the input signal at 9 is high OFF The PDES module is turned OFF the valve is closed LED 4 is also off The source pressure value is still shown at the manometer 3 but opening of the valve is disabled in OFF position see also chapter 3 7 Timer potentiometer Potentiometer to set the ejection time of channel A The total ejection time is the reading of this potentiometer multiplied by the reading of the TIME BASE switch 5 Note Due to tolerances of resistors and capacitors the time preset with the analog timer may vary up to 20 8 START button Push button for starting operation of channel A in TIMER or EXTERNAL mode 9 TTL INPUT connector BNC connector for external operation of channel A with a TTL HI signa
4. D j Electronic Instruments for the Life Sciences ease wade OPERATING INSTRUCTIONS AND SYSTEM DESCRIPTION OF THE PDES 02TX PNEUMATIC DRUG EJECTION SYSTEM with analog timers and analog manometers PDES 02TX LA PDES 02TX Sp J CHANNEL A VERSION 2 0 npi 2013 npi electronic GmbH Bauhofring 16 D 71732 Tamm Germany Phone 49 0 7141 9730230 Fax 49 0 7141 9730240 support npielectronic com http www npielectronic com PDES 02TX User Manual Table of Contents lie Safety Regulations ssis ore ie ea ihotea ia ea aea e ear aada ease 3 2 PDES System svcisdivsssdcuacessesense niea re a REE KR EEE E E aaee 4 Dell System LIESCT SEL N seess a E a N EA EOE EES 4 2 2 Description of the Front Panel wsccccissccaisissovsstavsesaciavecvessiscevaceaseasaaea Sedeedsacsssaceuscdeedenspees 5 2 3 Description ot the Rear Panel 252 20 neie e eg eee 7 Deg External Valven niaar e E AE R Ea a Som A ERE 8 Remote Valve Plate soreerccrois oiea a E A atate eatenaaetotes 8 microJECT micro Valve Pipette Holder sseeeeeeeeseeseeseeesessersseseresressrsrresresseseresresse 9 As Oper t oN sinsi es eE ear a ARE E a E EE E R EnS 10 Se MEGAN Se ite nnne lit cies a a aaah ie E itt e E E S 13 Microinjection into OOCYIES iicscsccsvesvacccdavsvonedtwayaatacciaieasebeaddaseasadeasunnaceassieens bebenateavers 14 Fast drug application with microJECT EHM eee eesceseseeeeeeceaeceseeeeeeeeneeeaeees 14 G2 Technical Mal
5. PUT Me CONTINUOUS TIMER mise pressure at ef OFE pipette tip MODE 200 ms 4 6 x10 sec 3 7 xl sec 2 8 x0 1 sec 1 9 x0 01 sec 0 1 10 TIME BASE TIMER A if the internal timer is set to 200 ms Figure 6 Operation modes of the PDES 2TX version 1 6 page 12 PDES 02TX User Manual 5 Literature Bickmeyer U Heine M Manzke T amp Richter D W 2002 Differential modulation of Ih by 5 HT receptors in mouse CA1 hippocampal neurons Eur J Neurosci 16 209 218 Eggermann E amp Feldmeyer D 2009 Cholinergic filtering in the recurrent excitatory microcircuit of cortical layer 4 Proc Natl Acad Sci U S A 106 11753 11758 Garaschuk O Milos R I amp Konnerth A 2006 Targeted bulk loading of fluorescent indicators for two photon brain imaging in vivo Nat Protocols 1 380 386 Guzman J N Hernandez A Galarraga E Tapia D Laville A Vergara R Aceves J amp Bargas J 2003 Dopaminergic modulation of axon collaterals interconnecting spiny neurons of the rat striatum J Neurosci 23 8931 8940 Hanganu I L amp Luhmann H J 2004 Functional Nicotinic Acetylcholine Receptors on Subplate Neurons in Neonatal Rat Somatosensory Cortex Journal of Neurophysiology 92 189 198 Kilb W Hanganu I L Okabe A Sava B A Shimizu Okabe C Fukuda A amp Luhmann H J 2008 Glycine receptors mediate excitation of subplate neurons in neonatal rat cerebral co
6. T coupling where the pressure source is connected an OUTPUT coupling where the injection pipette is connected and an EXHAUST coupling where a low retain pressure can be applied in order to avoid capillary effects at the tip of the injection pipette analogous to the retaining current used in iontophoretic systems This pressure must be generated by an additional pneumatic device Please contact npi electronic for details The injection pressure is preset by a precision pressure regulator The pressure is displayed on an analog standard or digital option manometer Additional pneumatic devices tubes fittings connectors filters etc are available Please contact npi electronic for details A calibration procedure allowing quantitative drug application is described by Hofmeier and Lux 1981 see chapter 5 version 1 6 page 4 PDES 02TX User Manual 2 2 Description of the Front Panel TIMER B 2 1 MONITOR TIME BASE MODE Q TIMING OFF 10 sex sec sec ON EJECT O lt x0 sex TIMER EXTERN x0 0 CONTINUOU CHANNEL B ipp HRE FE ON EJECT PDES 02TX 2 CHANNEL A Figure 1 PDES 2TX front panel view the numbers are related to those in the text below version 1 6 page 5 PDES 02TX User Manual PDES systems are available with one or two channels Since both channels are identical only channel A is described 1 POWER switch Switch to turn POWER on or off
7. ace 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 HIGH PRESSURE PRECAUTIONS When working with PDES systems and components always adhere to the appropriate safety measures for handling high pressure pneumatic systems Before using the PDES or any external pneumatic device please read manuals and instructions carefully e Always turn off pressure source before connecting or disconnecting the PDES system e Any component used tubes fittings etc must be specified for the maximum possible pressure
8. ay cates accion sa ceca E E tess sect taenctel es soos sect a E E sateen SAA 15 version 1 6 page 2 PDES 02TX User Manual 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 by trained staff only 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 Repl
9. connector Turn channel on TIMER control The LED is red when no ejection occurs and green when the pressure is applied to the pipette Select the operation mode using the respective toggle switch at the front panel e Inthe CONT position the application can be controlled manually e In the EXT position the duration of the application is determined directly by the duration of the TTL pulse applied to TTL INPUT e In the TIMER position the duration is preset by the timer control and time base of the PDES REMOTE control The selected operating mode determines the way the REMOTE control see Figure 5 works e Inthe CONTINUOUS mode the remote control unit is disabled e In the EXTERN mode the valve is open as long the REMOTE control switch is activated cont continuous 4 temporarily e In the TIMER mode the internal timer is triggered by activation of the REMOTE control switch The red LED indicates the channel that has been turned on version 1 6 page 10 PDES 02TX User Manual Figure 5 PDES remote control Note Green LED indicates that the valve is not connected properly or continuous operation version 1 6 page 11 PDES 02TX User Manual Manual mode os CONTINUOUS mode on OFF MODE TNE CONTINUOUS mode off EXTERN pressure at A pipette tip EXTERNAL mode O a TTL INPUT IV CONTINUOUS TIMER EXTERN OFF VODE pressure at bar pipette tip MODE TIMER mode TTL IN
10. e Do not use any aggressive combustible or explosive gases Only gases such as air nitrogen carbon dioxide helium etc are allowed e Use only dry oil free filtered gases max 5 um filter is allowed Maximum input pressure must be below 5 bar 1 bar 100 000 Pa npi recommends an input pressure of 1 bar more than the maximum output pressure version 1 6 page 3 PDES 02TX User Manual 2 PDES System 2 1 System Description Pressure ejection is a convenient method for applying both ionic and non ionic solutions from micropipettes This method is popular for two reasons First it can apply to almost all solutions Second the amount of solution ejected can be monitored by viewing the tip of the micropipette under a microscope npi PDES units are designed for pressure ejection of drugs in physiological and pharmacological studies PDES systems can be equipped with high pressure outputs up to Pmax 4 bar 58 psi for drug application with fine tipped micropipettes or with low range pressure output Pmax 2 bar 29 psi for controlled droplet application Analog or digital timers and a remote control unit facilitate the use of these instruments Each PDES system is housed in a 19 cabinet that can be mounted in a rack It comprises of one or two channels Each channel consists of a precision pressure regulator an analog or digital pressure display electronically controlled valves control electronics and a regulated pow
11. er supply Ejection of drugs can be activated manually or via a TTL input The operational status is indicated by a red green LED Each channel can be equipped with an EXHAUST coupling whereby a low retain pressure can be applied analogous to the retaining current used in iontophoretic systems In order to speed up drug ejection PDES systems are available with the microJect option These systems are equipped with a micro valve located in an injection holder The micro valve has a switch time of less than 1 ms and a very small volume that has to be controlled resulting in application times in the ms range Mode of operation and timing is selected by controls at the front panel If the internal timers are used the eject time is preset by a control and the time base can be selected by a toggle switch Each channel can be turned off by using a switch which is combined with the timing control The operational status is indicated by a red green LED No light indicates that the channel has been turned off red means that the channel has been turned on green indicates that injection takes place The remote control unit is housed in a small box For each channel it has a red LED that indicates if the respective channel is turned on and a toggle switch with a temporary and a fix position for starting drug application The pressure connectors quick connect type are located at the rear panel of the instrument Each channel is equipped with an INPU
12. l 5V In TIMER mode the internal timer is triggered by the TTL signal In EXTERN mode the valve is open as long as the TTL signal is HI 10 TIMING MONITOR BNC connector providing a TTL signal that indicates the status of the regulated injection valve of channel A HI valve open LO valve closed version 1 6 page 6 PDES 02TX User Manual 11 REMOTE CONTROL connector Connector for the REMOTE CONTROL unit see Figure 5 2 3 Description of the Rear Panel CHANNEL B vA ae tj Figure 2 PDES 2TX rear panel view the numbers are related to those in the text below The following connectors are located at the rear panel Connectors are mentioned only for CHANNEL A Since both channels are identical only one channel is described 1 VALVE connector optional Connector for the electrical connection of the VALVE located in a remote valve box or in the microJECT Important This connector is not used for systems without a remote valve or microJECT 2 OUT connector Connector for connecting the injection micropipette or for connecting the pressure tubing of the external valve on the remote valve plate The EJECT pressure is supplied to this connector 3 EXHAUST connector optional Connector for connecting the HOLD pressure source If no HOLD pressure is connected the system is exhausted as long as no pressure is applied to the pipette 4 SOURCE IN connector Connector for connecting the
13. rtex J Neurophysiol 100 698 707 Kolbaev S amp Draguhn A 2008 Glutamine induced membrane currents in cultured rat hippocampal neurons Eur J Neurosci 28 535 545 Langer J amp Rose C R 2009 Synaptically induced sodium signals in hippocampal astrocytes in situ J Physiol 587 5859 5877 Lin B J Chen T W amp Schild D 2007 Cell type specific relationships between spiking and Ca2 i in neurons of the Xenopus tadpole olfactory bulb Journal of Physiology 582 163 175 Marandi N Konnerth A amp Garaschuk O 2002 Two photon chloride imaging in neurons of brain slices Pflugers Arch 445 357 365 Partridge L D H U Zeilhofer and D Swandulla 1998 Combined Whole Cell and Single Channel Current Measurement with Quantitative Ca2 Injection or Fura 2 Measurement of Ca2 in P M Conn Ed Ion Channels Part B Methods in Enzymology Vol 293 Academic Press SanDiego Ponimaskin E Dumuis A Gaven F Barthet G Heine M Glebov K Richter D W amp Oppermann M 2005 Palmitoylation of the 5 Hydroxytryptamine4a Receptor Regulates Receptor Phosphorylation Desensitization and beta Arrestin Mediated Endocytosis Molecular Pharmacology 67 1434 1443 Qiu J Yao S Hindmarch C Antunes V Paton J amp Murphy D 2007 Transcription factor expression in the hypothalamo neurohypophyseal system of the dehydrated rat upregulation of gonadotrophin inducible transcription fac
14. t Pressure range input pressure max 5 bar 1 bar 10 Pa pressure control range 0 4 bar or 0 2 bar display range 0 4 bar or 0 2 bar Input and output couplings quick connect type Tubes input 6 mm OD output 4mm OD Status EJECT time indicator red green LED red light channel has been turned on valve is closed green light valve is open Mode selection three position toggle switch TIMER CONTINUOUS EXTERN TTL Minimum pressure pulsed width 100 ms Analog timer standard version 0 001 0 1 0 01 1 s 0 1 10 s and 1 100 s linear control range set by toggle switch time base selector 0 01 0 1 s 1 s 10 s Pulse input standard BNC connector TTL LO 0 2 5 V HI 3 5 V input impedance 10 KQ Timing output standard BNC connector TTL LO 0 2 5 V HI 3 5 V Power r eq uirements 1 channel DC 12 V 0 5 A min 2 channels DC 12 V 1 0 A min Remote control option Toggle switch and status LED for each channel size 2 5x4x10 cm version 1 6 page 15
15. tor 1 mRNA is mediated by cAMP dependent protein kinase A J Neurosci 27 2196 2203 Stepanyan R amp Frolenkov G I 2009 Fast adaptation and Ca2 sensitivity of the mechanotransducer require myosin XVa in inner but not outer cochlear hair cells Journal of Neuroscience 29 4023 4034 Wolfram V amp Juusola M 2004 The Impact of Rearing Conditions and Short Term Light Exposure on Signaling Performance in Drosophila Photoreceptors Journal of Neurophysiology 92 1918 1927 version 1 6 page 13 PDES 02TX User Manual Microinjection into Oocytes Wetzel C H Oles M Wellerdieck C Kuczkowiak M Gisselmann G amp Hatt H 1999 Specificity and sensitivity of a human olfactory receptor functionally expressed in human embryonic kidney 293 cells and Xenopus Laevis oocytes J Neurosci 19 7426 7433 Fast drug application with microJECT EHM Courjaret R Troger M amp Deitmer J W 2009 Suppression of GABA input by Al adenosine receptor activation in rat cerebellar granule cells Neuroscience 162 946 958 Heck N Kilb W Reiprich P Kubota H Furukawa T Fukuda A amp Luhmann H J 2007 GABA A receptors regulate neocortical neuronal migration in vitro and in vivo Cereb Cortex 17 138 148 version 1 6 page 14 PDES 02TX User Manual 6 Technical Data Source gas air nitrogen helium etc no aggressive combustible or explosive gases filtered by a 5 um filter elemen
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