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A shielded microelectrode suitable for single-electrode voltage clamping of neurones in the CNS.

The fabrication of a shielded microelectrode is described which has a time constant of about 3 microseconds, regardless of its depth in neural tissue. The shield is insulated from surrounding tissue with an insulation resistance which exceeds 1000 M omega, and the insulation will withstand +/- 15 V. Because of its narrow profile, the electrode is suitable for in vivo intracellular recordings from motoneurones and other large neurones of the CNS. The small time constant and the high breakdown resistance of the electrode make it suitable for use with single-electrode voltage clamp circuitry to voltage clamp motoneurone somata using switching rates in excess of 30 kHz.

Animals↗

Continuous measurement of pentylenetetrazol concentration by a liquid ion exchanger microelectrode.

A double-barrelled microelectrode is described, which permits the continuous measurement of the concentration of the epileptogenic agent pentylenetetrazol (PTZ). The electrode is based on the liquid potassium exchanger (Corning No. 477 317) and enables measurements of PTZ concentration in physiological salines down to 1 mM. The electromotive behaviour of the liquid membrane against PTZ cannot directly be described by the Nicolsky-Eisenman formalism. It is suggested that specific interactions of the PTZ molecule with the Corning ligand are involved in the potential generating mechanisms.

Animals↗

A simple method for making ion-selective microelectrodes suitable for intracellular recording in vertebrate cells.

A simple procedure for manufacturing Cl-, K+, and pH liquid membrane ion-sensitive microelectrodes is presented in detail. Electrodes suitable for recording from the specimen of interest are back-filled with a small amount of silane solution and heated for 5 min on a hot plate at a temperature between 400 and 500 degrees C, after which they are injected with the ion-sensitive resin. The procedure is adaptable to many different glass stocks, e.g., single-barreled, double-barreled, or theta glass, and can be used to produce electrodes having a wide range of tip sizes for recording either extracellular or intracellular ion activities. Another advantage of the method is speed; up to 10 electrodes can be prepared simultaneously, permitting over 40 functional electrodes to be made per hour.

Animals↗

Anti-adhesive coating for glass microelectrodes.

A simple procedure is described for coating the tips of glass micropipette electrodes with a durable thin film of a commercially available anti-adhesive agent (Antispread). This lowers the surface tension of the electrode glass, and helps reduce damage and distortion to tissue, by preventing the microelectrode from sticking permanently to cell membranes. In controlled trials, the coating actually increased the probability that nearby cells could be recorded from successively by about 30%. Electrode resistance increased only very slightly (on average by 8%) as a result of the coating. Coated electrodes yielded excellent long-term penetrations of small visual cells, and in addition allowed these to be dye-filled in the normal way. The coating may have applications in all situations where prevention of wetting by tissue is desirable.

Animals↗

An improved microelectrode resistance meter.

A device is described for measuring the resistance of micropipette electrodes. The useful range of electrode resistances that it can measure is 100 k omega to 1 G omega. It is more convenient to use than previously described or commercially available meters, especially for very high-resistance electrodes. Resistance of even the finest microelectrodes can be measured accurately while their tips are inserted by hand into a test solution. This eliminates the need for special holders, speeding and simplifying the screening of large numbers of electrodes. Test solutions are stored in interchangeable reservoirs, making it easy to characterize the resistance of an electrode in solutions of different resistivity. Test solutions can also be capped and removed when not in use to prevent evaporation. To protect very high-resistance electrodes from damage during measurement, the measuring current is low (only 300 pA on the highest resistance range) and the test voltage across the electrode is limited to +/- 1 V.

Electric Conductivity↗

Effects of temperature and temperature gradients on ion-sensitive microelectrodes.

Ion-sensitive microelectrodes are widely used in studies of mammalian tissues. Often the tissue is maintained at 37 degrees C, some 10-15 degrees C above room temperature. The temperature difference between the room and the preparation was found to be capable of altering the measured ion potential by as much as 10 mV. The change depended on 3 factors: the temperature dependence of the Nernst slope, the temperature dependence of the interference factor, and the thermoelectric potential induced by the temperature difference between the two ends of the ion-exchanger column. Certain combinations of these changes can cancel each other, resulting in spurious but apparently temperature-insensitive readings. The first two factors can produce errors when the temperature of calibration differs from the temperature of the tissue being measured. Serious errors in measurements of ion concentration can also occur, due to all 3 factors, if a temperature gradient exists across the ion exchanger column; this situation can easily occur when recording from exposed mammalian tissues. The use of a short ion-exchanger column will reduce but not eliminate effects due to a temperature gradient.

Animals↗

Multisite hippocampal slice recording and stimulation using a 32 element microelectrode array.

A technique has been developed in which a planar array of 32 microelectrodes, arranged in a 4 by 8 pattern with 200 micron separation, is used to record from and stimulate the hippocampal slice preparation at multiple sites. Control of media flow past the tissue is critical to observe signals and preserve viability. Active suppression circuitry is used to prevent device saturation due to large stimulation artifacts. The field potentials recorded are spatially unique and provide a 2-dimensional description of the underlying population activity in the various pyramidal strata and subpopulations. Multisite stimulation is also possible with the array, permitting the experimenter to quickly stimulate and record from brain slices in many spatial patterns.

Animals↗

Calibration of ion-selective microelectrodes: flow-system and analysis program for the IBM PC.

A menu-driven program--written in C and Assembly language--has been developed for use with an IBM PC or compatible computer for calibration of ion-selective microelectrodes. Available features include (i) user-logical commands, (ii) rapid, accurate construction and display of calibration curves (fitted by least squares non-linear regression), (iii) analysis of data (stored either on- or off-line), with correction for drift and conversion from voltage or length to ion concentration or activity, (iv) calculation of selectivity coefficients, (v) optional storage and use of activity coefficients, and (vi) output of data, calculations and graphics to printer and plotter. A simple chamber and valve-activated flow-system for rapid change of solutions with either manual selection or computer control is also described.

Microelectrodes↗

A multiwire microelectrode for single unit recording in deep brain structures.

A method is described by which a single shaft multiwire microelectrode can be fabricated efficiently. The resulting electrode can be attached to a commercial microdrive and used for single neuronal unit recording from one or more tracks in deep brain structures of anesthetized or awake animals. The electrode consists of a 30 gauge stainless steel cannula through which multiple strands of 13 micron insulated tungsten microwires are threaded. At the electrode tip the wires protrude 3-4 mm from the cannula and are cut individually at suitable offsets. The tip is stabilized and fixed to the cannula with cyanoacrylate. At the base of the electrode the wires are threaded through flexible plastic tubing that provides strain relief and are glued to individual pins of a miniature connector that plugs into a field effect transistor (FET) voltage follower. Good single unit recordings have been obtained routinely from the basal ganglia of awake, behaving monkeys with this electrode.

Action Potentials↗

A slim needle-shaped multiwire microelectrode for intracerebral recording.

The construction of a needle-shaped multiwire microelectrode is described. It can be made with simple mechanical tools. The presented electrode assembly consists of 12 insulated nichrome wires (core diameter 25 microns) which are embedded in epoxylite resin. The straight-cut wire tips are aligned lengthwise and have a relative spacing of 150 microns. Outer dimensions vary from 100 x 180 microns at the level of the 1st electrode channel, to 100 x 100 microns at the level of the 12th channel at the tip. The configuration of this electrode was determined by its application: the laminar analysis of evoked potentials in the cortex of the rat. However, the number of channels, the diameter of the (nichrome) wire which determines the surface area of these channels, and the channel spacing can be easily adjusted during construction to meet other experimental requirements, such as the recording of single-unit activity. The electrode which is composed of biocompatible materials is suited for the study of field potentials and multiple-unit activity, in both acute and chronic experiments, and can be used repeatedly. To demonstrate the performance of the electrode assembly, a depth profile of field potentials is presented, accompanied by the corresponding current source density distribution. The potentials were recorded in the somatosensory cortex of the rat following stimulation of the median nerve under ketamine anesthesia.

Animals↗

Simultaneous "real-time" electrochemical and electrophysiological recording in brain slices with a single carbon-fibre microelectrode.

Many previous studies have demonstrated the value of carbon-fibre microelectrodes (CFMs) for single-unit activity recording and for fast cyclic voltammetry. In this report we show that these two independent methodologies can be combined at a single CFM and used to study simultaneous electrochemical and electrophysiological events in brain slices. In superfused slices of rat locus coeruleus, dorsal raphe and substantia nigra, we were able to record stable electrophysiological signals and stimulated monoamine efflux for periods of at least 2 h, thereby allowing quantitative pharmacological interventions. The simultaneous recording of amine efflux and unit activity at the same locus facilitates comparison of drug effects at pre- and post-synaptic sites. Furthermore, the system described here uses commercially available instrumentation. The circuitry is described and examples of its application are shown.

Animals↗

Ion-selective microelectrodes and diffusion measurements as tools to explore the brain cell microenvironment.

The construction and application of liquid-membrane ion-selective microelectrodes (ISM) are described. Recommendations are provided for the selection of appropriate cocktails containing neutral carriers to form the liquid membrane to sense K+, Ca2+, H+ and Na+. The use of charged carriers to sense Cl- and the cation tetramethylammonium (TMA+) is discussed. A detailed protocol is given for constructing double-barreled electrodes (ion-sensor and reference barrel) with tips of 1 micron diameter or more for extracellular ion measurements. The primary results obtained with ISMs in the brain cell microenvironment are briefly surveyed. The theoretical basis for measuring diffusion properties of extracellular space is described. Such measurements enable the estimation of volume fraction (proportion of tissue that is extracellular space) and tortuosity (hindrance of diffusion due to cellular obstructions). A method is given for using TMA+ ISMs in combination with iontophoresis or pressure ejection of TMA+ from a nearby micropipette to measure diffusion properties.

Animals↗

A method for bevelling of microelectrodes by means of vibration.

A technique for the rapid bevelling of glass and metallic microelectrodes is described. The method utilizes end friction of the electrode tips on the fine-grained surface of a quartz plate vibrating at the mechanical resonance frequency of the system. A electromagnetic vibrator is supplied with alternating current at a frequency equal to the mechanical resonance frequency of the system (200-300 s-1).

Electromagnetic Phenomena↗

A simple and comprehensive method for the construction, repair and recycling of single and double tungsten microelectrodes.

A method for the construction, repair and recycling of tungsten-in-glass micro-electrodes is described. Essentially, the method employs insulating tungsten electrodes by collapsing borosilicate-glass capillaries onto tungsten wires. Procedures are described for either (1) removing excess glass insulation from the microelectrode tip or (2) fine adjustment and reshaping of the exposed electrode tip. This approach enables the controlled shaping of the electrode tip during preparation and/or reshaping the electrode tip or glass insulation of damaged electrodes. The method may be applied in the preparation of bipolar tungsten-in-glass electrode and tungsten electrode/micropipette assemblies. Advantages of the method are discussed.

Electrophysiology↗

Toward the ultimate metal microelectrode.

The performance of metal microelectrodes for stimulating and recording neuronal action potentials depends on precise control of their geometrical, electrical and mechanical properties. We describe a combination of materials whose properties approach fundamental physical limitations on achievable performance and reproducible fabrication techniques that provide probes with very small dimensions. Pure iridium wire is electrolytically sharpened, vapor-coated with Parylene-C insulation and the tip exposed using an automatically steerable UV laser. Electrochemical activation of the iridium increases the capacitance of the metal-electrolyte interface so that the overall impedance in the relevant frequency band (100-10,000 Hz) is dominated by the access resistance of the surrounding tissues.

Electric Conductivity↗

Marking microelectrode penetrations with fluorescent dyes.

Fluorescent dyes were used to mark and identify the tracks left by extracellular microelectrodes in neurophysiological experiments. Forty-two penetrations were made into the postcentral gyrus of 3 Macaque monkeys with electrodes coated with 1 of 5 fluorescent dyes (DiI, DiO, DiI-C5, PyPO, and Fast Blue). The electrodes were driven at rates ranging from 10 to 1000 microns/min, to a depth of about 4000 microns, where a small electrolytic lesion was made. Histological sections were viewed under fluorescent optics and the electrode tracks were reconstructed from the dye traces. Fluorescent traces (width 50-400 microns) were observed in 41 of 42 penetrations with 24 traces extending to the lesion site. Of the electrodes driven in less than 3 h, those coated with DiI (8/8) and DiI-C5 (8/8) left a trace to the lesion site, while 57% (4/7) of the DiO, 40% (2/5) of the Fast Blue and only 11% (1/9) of the PyPO tracks were fully marked. This method of marking penetrations can be used with any extracellular recording configuration, does not require tissue sections to be processed or stained, does not require electrical lesions, and causes no detectable tissue damage. Because the dyes fluoresce at different wavelengths, closely spaced tracks can be uniquely identified.

Animals↗

Evaluation of brain tissue O2 supply based on results of PO2 measurements with needle and surface microelectrodes.

Tissue PO2 distribution was measured in rat cerebral cortex during arterial normoxia and arterial hypoxia. The study was designed to examine the relationship between the PO2 histograms determined with surface electrodes in the superficial cortical cells and with needle electrodes in the cortical laminae below. Under normoxic as well as under hypoxic conditions the PO2 distributions of the compared brain regions were in close agreement. The results indicate that PO2 measurements in the brain cortex with surface microelectrodes give a correct view over the tissue oxygenation in cortical regions up to 500 microns below the measuring field.

Animals↗