An improved simple tungsten microelectrode

Brain Resecrrch Eullerin,

Vol. 4,

pp. 285-286.

Printed in the U.S.A

An Improved Simple Tungsten Microelectrode T. R. VIDYASAGAR

AND G. W. PERRY’

Visual Sciences Laboratories, Department of Ophthalamic Optics University of Manchester Institute of Science und Technology, Munchester M60 lQD, U.K. (Received

26 September

1978)

VIDYASAGAR, T. R. AND G. W. PERRY. At1 improved simple trtrlgsten rnicroelecirotie. BRAIN RES. BULL. 4(2) 285-286, 1979.-An improvement to tungsten-in-glass microelectrode has been made which requires no special skills. It involves the introduction of a gel solution into the glass micropipette which surrounds and binds the tungsten wire along its full length. A significant damping of microphonic effects is produced and a good fit between tungsten and glass near their tips becomes less critical for success. The electrode was found to be very stable over long recording sessions. Borosilicate glass

Tungsten microelectrode Microphonic effects

Polyacrylamide

SINGLE unit studies in the mammalian central nervous system received a major boost when tungsten microelectrodes were invented by Hubel [l] in 1957. These were first lacquer coated except at the tip. The coating, however, is prone to peel off during penetrations of the brain, thus altering the resistance and tip size markedly. Further, varnishing is tedious and controlling the area of uninsulated tip is difticult. The next major improvement came with the development of glass-coated tungsten electrodes [2,3]. The glass provided insulation as well as greater strength to the electrode. Levick [2] introduced a sharpened tungsten wire into a borosilicate glass micropipette such that only its tip protrudes through the fine end of the pipette and cements the wire to the glass at the broad end of the pipette. Merrill and Ainsworth 131 actually pull the glass micropipette over the sharpened tungsten wire and thus ensure a close binding between the two. Both these methods produce excellent electrodes with controllable tip lengths; however, they have a few demerits. Success with Levick’s method [2] depends on ensuring that the relative shapes of wire and pipette are appropriate to provide a good mechanical bond. This unfortunately is not always easy to achieve. With Merrill and Ainsworth’s [3] technique removing the glass from a desirable length of the tip is not a simple procedure. Also, microphonic electrodes are a frequent problem. We report here a method that is considerably faster, does not require any special skill and provides electrodes with remarkable stability. In brief, out method is very similar to Levick’s [2] except that we introduce a solution into the glass micropipette prior to inserting the tungsten wire and which later sets into a solid gel binding strongly the tungsten and glass.

gel

Riboflavin

Photopolymerization

used to draw micropipettes on any standard electrode puller, with controls set to obtain a tip diameter of approximately 1.5 pm with relatively short tapers. The glass micropipette is filled with an acrylamide based solution (monomer solution), similar to that used for polyacrylamide gel electrophoresis. The solution is prepared as follows: 24 ml N HCI; 3.0 g Tris (hydroxymethyl)-aminomethane; 0.23 ml TEMED(NNN’N’tetramethylenediamine); 2.5 g Acrylamide and 0.052 g Bisacrylamide, pH 6.8. This gives a 10% gel solution. To this is added riboflavin (0.3 mg/lOO ml). A polythene catheter (Portex Limited, Hythe, Kent, England) of outer diameter 0.63 mm, sufficiently small to allow easy passage inside the glass micropipette, is used for filling and care is taken not to introduce air bubbles. The micropipette is then mounted onto the adjustable stage of a microscope and the etched tungsten wire onto a suitable stand. The sharpened end of the wire is inserted through the broad end of the pipette to protrude to the desired length through the tip. Details of this assembly are similar to those used by Levick 121. Solution dispensed from the catheter rarely fills to the very tip of the pipette: however, as the tungsten wire is advanced through the tapering section of glass the solution runs easily to the tip. The tungsten is then glued to the broad end of the pipette with cyanoacrylate adhesive (Pedal Products, Newhaven, Sussex) and the electrode placed 10-15 cm from a fluorescent light source (20 W). The polymerization of the acrylamide solution proceeds only in the presence of free radicals, produced here by the photodecomposition of riboflavin under fluorescent light. The solution turns to a solid gel within 15-30 min. The tips may then be plated with a gold cyanide solution and platinum black [ 31.

METHOD

DISCUSSION

Tungsten wire of diameter 0.125 mm is sharpened by etching in a solution containing sodium nitrite and potassium hydroxide [2]. Borosilicate glass capillaries (Coming 7740; 2.0 mm outside diameter, 0.35-0.5 mm wall thickness) are

Polymerization of the gel solution is inhibited by atmospheric oxygen; consequently, solution which may coat the electrode tip as it protrudes is unlikely to polymerize and can be washed off when the electrode has set. Normally, ac-

‘Present address: Department

Copyright

of Physiology and Biophysics,

0 1979 ANKHO

School of Medicine, University of Miami, Miami. Florida 33152, U.S.A.

International

Inc.-0361-9230/79/020285-02$00.70/O

VIDYASACSAK AND PERRk

286

_

I

2fYIV

~~~ __

_

___

_____

i

Lo-5 s

FIG. 1. Responses obtained by using glass and gel coated tungsten microelectrodes. (Top) Extra&h&r recordingof a single action potential from a cell in the rat’s Lateral geniculate nucleus. (Bottom) Extracelhtlar recording of the habituating responses of a visual ceh in the intermediate layer of the rat superior coliicuius to repeated visual stimulation.

ryfamide solutions require degassing before photopolymerization will occur; however, we have found it unnecessary to remove dissolved oxygen from the solution when using the proportions of constituents suggested above. Nevertheless, degassing may occasionally be necessary. Other problems with solutions not ~lyrne~~~ may be encounte~d if riboflavin and acrylamide stocks are too old. Fresh constituents are highly desirable. A stock solution may be kept at 4°C and protected from light for as long as a month. Also, it is better to use the solution cold without warming, as the low temperature slows the exothermic ~lyme~~tion which may expand the tungsten wire. The speed of the polymerization reaction can be controlled by adjusting the concentration of TEMED and riboflavin. Similarly, the gel consistency can be increased or decreased by adjusting the acrylamide concentration, but we have found that with higher concentrations the tungsten wire is pushed further out of the electrode tip during polymerization. An alternative to photopolymerization using riboflavin is the chemical production of free radicals using ammonium persulphate. This reaction does not require fluorescent light and is controlled only by the concentra~n of persulphate ions. Consequently, the time in which to make the electrodes before the solution polymerizes is limited. (Polymerization occurs in 20-30 min with 18 mg of persutphate added to 25 ml of monomer solution.) Further, persulphate induced polymerization appears to be more resistant to inhibition by atmospheric oxygen and solution coating the tip of the electrode may polymerize and increase the electrode resistance. For these reasons we prefer and recommend photopolymerization using riboflavin.

Other gelling solutions were tried, such as gelatine, agar and agarose; but all these alternatives were found to he unsuitable because they required warming and cooling before setting, or were too viscous. The acrylamide solution remains in an aqueous state until the polymerization reaction is initiated and readily passes intu the glass micropipette. In their monomeric state, acrylamide and bisacrylamide are neurotoxic. However, when polymerized, the gel is quite inert and no adverse effects to neurones during recording sessions have been observed. The electrodes are very easy to make and provide excellent mechanical stability. The relative shapes of the tungsten wire and glass at their tips are not vital to success as the gap that might exist between them is ftied withget. However, for this very reason, the profile of the pipette along its tip tends to be broader and thus inferior to electrodes made accordiug to the method of Merrill and Ainsworth [3], especiahy for deep recordings; but the simplicity of technique may be more endearing. A major advantage of this electrode is the virtual elimination of microphonic effects. The tungsten is closely tethered to the gel along its full length and any tendency for oscillation with acoustic fedhack from the ampfiBer’s loudspeaker is greatly damped. The electrodes are regufariy used in our laboratory for recording single units from mammalian btains and stable extraceiiular recording sessions for as long as 6-8 hours on one cell are not unusual (Fig. 1).

ACKNOWLEDGEMENTS We thank David Carden and Ian Wright for helpful discussions.

REFERENCES 1. Hubel, D. M. Tungsten microelectrode

for recording from single units. Science 125: 549-550, 1957. 2. Levick, W. R. Another tungsten microelectrode. Med. Bioi. Effg. lo: 51&515, 1972.

3. Merrill, E. G. and A. Ainsworth. Glass-coated platinum plated tungsten microelectrodes. Med. Biol. Eng. X8: 642-672, 1972.