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Five topographically organized fields in the somatosensory cortex of the flying fox: microelectrode maps, myeloarchitecture, and cortical modules.

Five somatosensory fields were defined in the grey-headed flying fox by using microelectrode mapping procedures. These fields are: the primary somatosensory area, SI or area 3b; a field caudal to area 3b, area 1/2; the second somatosensory area, SII; the parietal ventral area, PV; and the ventral somatosensory area, VS. A large number of closely spaced electrode penetrations recording multiunit activity revealed that each of these fields had a complete somatotopic representation. Microelectrode maps of somatosensory fields were related to architecture in cortex that had been flattened, cut parallel to the cortical surface, and stained for myelin. Receptive field size and some neural properties of individual fields were directly compared. Area 3b was the largest field identified and its topography was similar to that described in many other mammals. Neurons in 3b were highly responsive to cutaneous stimulation of peripheral body parts and had relatively small receptive fields. The myeloarchitecture revealed patches of dense myelination surrounded by thin zones of lightly myelinated cortex. Microelectrode recordings showed that myelin-dense and sparse zones in 3b were related to neurons that responded consistently or habituated to repetitive stimulation respectively. In cortex caudal to 3b, and protruding into 3b, a complete representation of the body surface adjacent to much of the caudal boundary of 3b was defined. Neurons in this area habituated rapidly to repetitive stimulation. We termed this caudal field area 1/2 because it had properties of both area 1 and area 2 of primates. In cortex caudolateral to 3b and lateral to area 1/2 (cortex traditionally defined as SII) we describe three separate representations of the body surface coextensive with distinct myeloarchitectonic appearances. The second somatosensory area, SII, shared a congruent border with 3b at the representation of the nose. In SII, the overall orientation of the body representation was erect. The lips were represented rostrolaterally, the digits were represented laterally, and the toes were caudolateral to the digits. The trunk was represented caudally and the head was represented medially. A second complete representation, PV, had an inverted body representation with respect to SII and bordered SII at the representation of the distal limbs. The proximal body parts were represented rostrolaterally in PV. Finally, caudal to both SII and PV, an additional representation, VS, shared a congruent border with the distal hindlimb representation of both SII and PV. VS had a crude topography, and receptive fields of neurons in VS were relatively large. Many neurons in VS responded to both somatosensory and auditory stimulation.

Animals

Measurement of current spread from microelectrodes when stimulating within the nervous system.

Tungsten stimulating microelectrodes have been tested in monopolar, bipolar and concentric configurations for the extent to which unwanted current spread occurred. Current spread from monopolar electrodes in close conformity with the predictions of the inverse square law, both in vitro in vivo. The bipolar and concentric configurations, tested only in vitro, had current-spread characteristics which did not follow the inverse square law so closely. The bipolar configuration gave little reduction in spread, compared wpith a monopolar electrode, but the concentric configuration did reduce the extent of stimulus spread. The extent of spread depends greatly upon a number of experimental variables, including the dimensions of the microelectrode tip. For studies requiring precise localization with low stimulus currents, it is advisable to determine the inverse square law constant for each experimental situation. For more generalized stimulation purposes within the mammalian C.N.S., employing monopolar microelectrodes of moderate tip size, brief pulses of 10 muA can be expected to stimulate myelinated axons with about 0.15 mm of the tip, and 100 muA WILL Stimulate with a radius of about 0.5 mm.

Animals

Active and passive properties of rabbit descending colon: a microelectrode and nystatin study.

The electrical properties of the basolateral membrane of rabbit descending colon were studied with microelectrode methods in conjunction with the polyene antibiotic nystatin. Two problems were examined: (i) the relative distribution of tight junctional, apical membrane and basolateral membrane resistances, and (ii) the ionic basis of the basolateral membrane potential. Intracellular K+ activity (K+) was measured using liquid ion exchanger microelectrodes ((K+) = 76 +/- 2 MM) and was found not to be in equilibrium with the basolateral membrane potential. In order to measure membrane resistances and to estimate the selective permeability of the basolateral membrane, the apical membrane was treated with nystatin and bathed with a K2SO4 Ringer's solution which was designed to mimic intracellular K+ composition. This procedure virtually eliminated the resistance and electromotive force of the apical membrane. Shunt resistance was calculated by two independent methods based on microelectrode and transepithelial measurements. Both methods produced similar results (Rs = 691 +/- 63 omega cm2 and 770 +/- 247 omega cm2, respectively). These findings indicate that the shunt has no significant selectivity, contrary to previous reports. Native apical membrane resistance was estimated as 705 +/- 123 V cm2 and basolateral membrane resistance was 95 +/- 14 V cm2. To estimate basolateral membrane selectivity, the serosa was bathed in a NaCl Ringer's solution followed by a series of changes in which all or part of the Na+ was replaced by equimolar amounts of K+. From measures of bi-ionic potentials and conductance during these replacements, we calculated potassium permeability and selectivity ratios for the nystatin-treated colon by fitting these results to the constant field equations. By correcting for shunt conductance, it was then possible to estimate the selective permeability of the basolateral membrane alone. Selectivity estimates were as follows: PNa/PK = .08 and PCl/ PK = .07 (uncorrected for shunt) and PNa/PK = .04 and PCl/PK = .06 (basolateral membrane alone). In a second set of experiments, evidence for an electrogenic Na+ pump in the basolateral membrane is presented. A small ouabain-sensitive potential could be generated in the nystatin-treated colon in the absence of chemical or electrical gradients by mucosal, but not serosal, addition of NaCl. We conclude that this electrogenic pump may contribute to the basolateral membrane potential; however, the primary source of this potential is "passive": specifically, a potassium gradient which is maintained by an "active" transport process. An appendix compares the results of nystatin experiments to amiloride experiments which were conducted separately on the same tissues. The purpose of this comparison was to develop a comprehensive model of colonic transport. The analysis reveals a leak conductance in the apical membrane and the presence of an amiloride-insensitive conductance pathway.

Animals

Biological and artificial ion exchangers: electrical measurements with glass microelectrodes.

Biological (stratum corneum) and artificial (cation-exchange resin beads, Bio-Rad AG 50W-X2) ion exchangers were impaled by glass microelectrodes filled with KCl solution. The electrical potential difference recorded in these structures in reference to the external bathing medium was shown to be dependent on the KCl concentration of both the external and the microelectrode filling solutions. The potentials were interpreted on the grounds of the fixed charge theory of membrane potentials as a consequence of two phase boundary potentials (Donnan potentials), one at the matrix-external solution interface and the other at the matrix-microelectrode solution interface. The contribution of a diffusion component for the recorded potential was considered.

Animals

Ion-selective microelectrodes suitable for recording rapid changes in extracellular ion concentration.

A method for fabricating double-barrel, ion-selective microelectrodes with fine tips (0.5-1.5 microns) and rapid response times is described. When made into K(+)-selective microelectrodes, the electrodes respond to changes in [K+]o with a time constant of 70-95 ms. The electrical response of these electrodes to common-mode voltages can be made to have a time constant of less than 2 ms, which minimizes electrical artifacts from field potentials. The application of these microelectrodes to the measurement of rapid, transient changes in retinal [K+]o is presented.

Animals

Optimization of glass microelectrode properties by response surface methodology.

Glass microelectrodes filled with electrolyte solutions are standard tools for electrophysiological studies. However, for any given application, there are limitations to the properties of the microelectrode, such as impedance and shank length, that can yield satisfactory results. The trial and error approach in pulling electrodes with the desired properties can be time consuming. The use of a response surface procedure which allows the experimenter to change more than one factor at a time and therefore determine the desired puller condition more efficiently is demonstrated. Also, design improvements for the World Precision Instrument, Model PUL-1, Microelectrode puller, used in this study are suggested.

Animals

Coincident recording and stimulation of single and multiple neuronal activity with one extracellular microelectrode.

This paper describes how an extracellular microelectrode may be used to stimulate neurons with brief, rectangular pulses and afterwards directly record the resultant activity. Two obstacles are the stimulus artifact lingering in the electrical circuitry and transient tip potentials (TTPs) arising from ion depletion at the electrode-tissue interface. Electronic switching between the stimulus source and the recording amplifier eliminates direct stimulus artifact from the electrical circuitry, although high but acceptable switching artifact remains. TTPs revert with time constants that are prominent in the desired recording (0.1-1 ms) and can reach 50 mV when more than 1 microA passes through a typical electrolyte-filled micropipette (for example 2-4 M omega, filled with 3 M NaCl, and placed in 0.1 M NaCl). They are always negative when cations flow into the tip, they are accompanied by a rise in microelectrode impedance, and they increase as a function of the resting electrode impedance, the duration and amplitude of applied current, and the dilution of the external electrolyte. TTPs were substracted by differential recording and stimulation through matched micropipettes (one in the brain and one in contiguous electrolyte) and in addition were reduced by pressure ejection of electrolyte. Directly elicited spikes (single or multiple) were detected about 0.5 ms after delivery of a rectangular stimulus pulse in the cerebellar cortex of pentobarbital-anesthetized rats. Typically, 3-4 units could be excited by less than 3 microA cathodal currents at any recording site. All-or-nothing properties, thresholds, and refractoriness to a second pulse within 2-4 ms verified the neuronal nature of the recorded signals. Complex wave forms, probably generated synaptically, were also seen. The technique of coincident extracellular recording and stimulation can be used as a universal search stimulus during microelectrode penetrations through the brain and in determining threshold-distance relations for extracellular stimulation. Where cell penetrations are unstable, it might be usefully substituted for intracellular technique in testing a neuron's behavioral or physiological influences or in exploring a cell membrane's response to drugs (in terms of excitability rather than voltage and impedance).

Animals

Adaptation of the Reitboeck method of multiple microelectrode recording to the neocortex of the waking monkey.

We adapted to the neocortex of waking monkeys a method for multiple microelectrode recording devised by Reitboeck. A sliding platform allows micropositioning of 7 electrodes independently, in 2 microns steps. Microelectrodes are quartz glass filaments (80 microns o.d.) with central metal cores (30 microns) of tungsten-platinum alloy. Filaments are drawn in a high temperature chamber, and ground to the desired form and tip size. The microdrive is held over the region to be explored, and the microelectrodes passed through 300 microns o.d. guide tubes fixed in implant thimbles of chosen size an and x - y arrangement of tubes, sealed by an O-ring into a small craniotomy opening. A microprocessor controlled recording system provides gain, noise and wave-shape filtering, impedance testing, and differential amplitude discrimination for each channel. Electrode movement is obtained via the microprocessor which displays and updates on the console terminal the electrode depth, impedance, and the channel assignment of each electrode. A second microprocessor based system is used to collect, buffer, and encode in real time all event data, which are transferred whenever convenient to a minicomputer that controls the experiment. Exploratory recordings were made in the posterior parietal, somatic sensory, and motor cortical areas. The system has now been used successfully in a number of investigations.

Action Potentials

Ovarian function after incision of the ovary by scalpel, CO2 laser, and microelectrode.

OBJECTIVE: The objective was to evaluate the relative effects of a standardized surgical incision performed with scalpel, CO2 laser, or microelectrode on ovarian tissue and function. DESIGN: Thirty sexually mature female rabbits were randomly assigned to one of three surgical groups. The injury consisted of a linear incision along the long axis of both ovaries, from the cortex to the hilum, followed by immediate unilateral oophorectomy for evaluation of the acute tissue damage. The opposite ovary was left in situ to evaluate subsequent healing, steroidogenesis, folliculogenesis, and ovulation. RESULTS: Minimal damage was observed after surgery with either laser or microelectrode. The ovarian stroma and follicles adjacent to the surgical site appeared normal both immediately and 15 days after all surgeries. Steroidogenesis, folliculogenesis, and luteinization were similar among the three groups. The number of ovulated oocytes after human chorionic gonadotropin was significantly reduced after scalpel and laser surgery compared with microelectrode (P less than 0.05). The former groups appeared to have a higher incidence of unruptured luteinized follicles with untrapped oocytes, which were often covered with a cellular fibrous membrane. CONCLUSIONS: These findings suggest that surgical trauma is well tolerated by the ovaries and does not impair subsequent ovarian function. However, rupture of the preovulatory follicle and its release of the oocyte may be influenced by the presence of ovarian adhesions and/or surgical trauma.

Analysis of Variance

Tapered tungsten fine-wire microelectrode for chronic single unit recording.

A new fine-wire microelectrode made from tungsten wire is described. The electrode design provides for strength as well as flexibility due to the stiffness of tungsten wire and a gradually tapering profile. Unlike previously-used fine wire electrodes, microelectrodes of the new design can be singly implanted under electrophysiological control into deep brain structures of larger animals such as cats. The flexibility of the fine diameter terminal portion of the microelectrode permits stable recording from caudal brain stem single units during vigorous behavioral activity.

Animals

Amperometric glucose microelectrodes prepared through immobilization of glucose oxidase in redox hydrogels.

Glucose microelectrodes have been formed with glucose oxidase immobilized in poly[(vinylpyridine)Os(bipyridine)2Cl] derivative-based redox hydrogels on beveled carbon-fiber microdisk (7 microns diameter) electrodes. In the resulting microelectrode, the steady-state glucose electrooxidation current density is 0.3 mA cm-2 and the sensitivity is 20 mA cm-2 M-1. The current density and sensitivity are 10 times higher than in macroelectrodes made with the same hydrogel. Furthermore, the current is less affected by a change in the partial pressure of oxygen. The higher current density and lower oxygen sensitivity point to the efficient collection of electrons through their diffusion in the redox hydrogel to the electrode surface. These results contrast with those observed for enzyme electrodes based on diffusing mediators, where loss of the enzyme-reduced mediator by radial diffusion to the solution decreases the current densities of microelectrodes relative to similar macroelectrodes.

Enzymes, Immobilized

Liquid and solid-state Cl- -sensitive microelectrodes. Characteristics and application to intracellular Cl- activity in Balanus photoreceptor.

When intracellular chloride activity (aiCl) was monitored with chloride-sensitive liquid ion exchanges (CLIX) microelectrodes in Balanus photoreceptors, replacement of extracellular chloride (Cl0) by methanesulfonate or glutamate was followed by a rapid but incomplete loss of aiCl. When propionate was used as the extracellular anion substitute, CLIX electrodes detected an apparent gain in aiCl, while a newly designed Ag-AgCl wire-in glass microelectrode showed a loss of aiCl under the same conditions. This discrepancy in Cl- washout when propionate replaced Cl0 is explained by the differences in selectivity of CLIX and Ag-AgCl electrodes for native intracellular anions and for the extracellular anion substitute which also replaces Cli and interferes in the determination of aiCl. Both electrodes indicate that ECl approximately Em when the cells are bathed in normal barnacle saline, and both electrodes showed the rate of Cl washout (tau approximately 5 min) to be independent of Cli when Cl0 was replaced by glutamate. Details of Ag-AgCl microelectrode construction are presented. These electrodes were tested and found to be insensitive to the organic anion substitutes used in this study. Selectivity data of CLIX electrodes for several anions of biological interest are described.

Animals

Halothane mimics oxygen in oxygen microelectrodes.

The effects of halothane and enflurane on the polarographic measurement of oxygen with five platinum and three gold microelectrodes were examined. Oxygen microelectrodes were calibrated in saline solution equilibrated with either nitrogen (N2) or air, then either halothane, 1.0 per cent, or enflurane, 2.0 per cent, was added to the gas mixture. For each electrode, polarographic curves were determined during exposure to five equilibrating gas mixtures: N2, air, N2 plus halothane, air plus halothane, and N2 plus enflurane. Halothane variably increased the current produced (and therefore the estimated oxygen tension) at all polarizing voltages in saline solution equilibrated with either N2 or air. The effect was present in both conical platinum electrodes and recessed-tip gold electrodes and was not prevented by membrane coatings of polystyrol, Rhoplex or collodion. Enflurane did not alter the polarographic measurement of oxygen. It is concluded that tissue oxygen tension measurements, made with these microelectrodes and membranes, may be unreliable in the presence of halothane.

Calibration

Ability of the Ca2+-selective microelectrodes to measure fast and local Ca2+ transients in nerve cells.

The ability of the Ca2+-selective microelectrode to measure fast Ca2+ transients intracellularly is reviewed. In vitro, Ca microelectrodes can respond to Ca2+ injections with time to peaks as small as 40 ms. We present methods to improve the dynamic response of Ca microelectrodes and to make Ca-buffered solutions in high ionic strength. Examples of measurements of intracellular free Ca2+ [( Ca2+]i) transients in Aplysia neurons and in Limulus photoreceptors are shown. To show the validity of those measurements, simultaneous recordings of the Arsenazo III (AIII) absorbance and of the Ca-selective electrode potential were made in voltage-clamped neurons of the abdominal ganglion of Aplysia californica. Pressure injection of AIII to a concentration of 300-500 microM induced a rise in resting [Ca2+]i; injection of higher [AIII] led to buffering of [Ca2+]i transients. Both techniques responded to changes in resting [Ca2+]i in the same direction except that AIII showed an increase in absorbance in 0 [Ca2+]o. Voltage-clamp pulses transiently increased both the AIII absorbance and the Ca2+ electrode potential. Reducing or increasing the driving force for Ca2+ entry changed the magnitude of both signals in the right direction. Examples of spatial localization of [Ca2+]i increases and Ca2+ gradients within the cytoplasm were demonstrated using the Ca electrode. The use of optical techniques to measure local [Ca2+]i changes is briefly reviewed.

Animals

Use of palladium touch microelectrodes under field conditions for in vivo assessment of dental plaque pH in children.

The aim of this study was to assess the applicability of palladium touch microelectrodes, connected to battery-run pH meters, for in vivo plaque pH measurements in children. The pH was assessed in 20 7-year-old and in 19 14-year-old caries-active and caries-inactive rural Kenyan children. The resting pH was measured at non-carious interproximal and occlusal sites and in open dentine cavities. Independent repeated measurements were performed at given sites at intervals of 15 s and 5 min and on different days. The resting plaque pH varied widely among the children, and there was no significant difference between caries-active and caries-inactive groups. The most striking feature was the considerable erratic fluctuations of pH at a given site with time, both in resting and in sucrose-challenged plaque. These fluctuations were sensitively recorded by palladium touch microelectrodes. After a sucrose rinse, not all sites in the same mouth behaved in a similar fashion, and thus the classical 'Stephan curve' was not always apparent. In conclusion, the palladium touch microelectrodes are highly applicable for plaque pH measurements in children, even under extreme field conditions.

Adolescent

The measurement of intracellular sodium activities in the bullfrog by means of double-barreled sodium liquid ion-exchanger microelectrodes.

A double-barreled Na+-selective microelectrode was constructed with monensin as a liquid ion exchanger. The HCl-treated monensin was dissolved in a solvent (Corning 477317) at 10% (weight/weight). Internal reference solution of its ionic barrel was mixture of 0.49 M NaCl and 0.01 M KCl, the pH being adjusted to 3 with 0.1 M citrate-HCl buffer, whereas that of the PD barrel was 0.5 M KCl. Average slope and selectivity ratio (Na+/K+) tested on 10 different microelectrodes were -57.5 +/- 1.87 mV/P(Na) (SEM) and 6.7 +/- 0.44, respectively. The electrical resistance was an order of 10(10) ohm and the response time was less than 10 sec. Using this microelectrode, a free flow micropuncture experiment was carried out in the bullfrog kidney and the intracellular Na+ activity as well as the membrane PD was determined on the proximal tubular cell. Average value (+/- SEM, n = 15) for the intracellular Na+ and K+ was 20.7 +/- 1.56 mEq/L and 61.2 +/- 1.16 mEq/L, respectively, and -68.7 +/- 0.88 mV for the peritubular membrane PD. There was a significant negative correlation between Na+ and K+ activities within the cell, i.e., the lower the ionic activity of cellular Na+ was, the higher the cellular K+, and vice versa, the sum of these two being kept nearly constant. The above finding may be somehow related to the isosmosis in the reabsorptive process across the proximal tubular epithelium.

Animals

Comparison of two approaches to measurement of electrical impedance of glass microelectrodes designed for evaluation of temperature changes in biological tissues.

We proposed a temperature sensitive microelectrode for rapid measurements of temperature at the cellular level. In principle, the electrical impedance of the tip of the microelectrode changes with temperature. We designed an impulse measurement system (STEP) sensitive to the above changes of impedance. The system is based on a presettable negative input impedance of the current to a voltage converter. We compared the efficiency of the new STEP with the currently used RAMP system. We found following advantages of the STEP system: i) the danger of high voltage oscillations which could mechanically destroy the microelectrode tip is eliminated; ii) this system provides the opportunity to set the maximum sensitivity of the system according to the measured temperature interval. Moreover, the STEP method makes it possible to measure the resistance by using a sinusoidal stimulation signal which has to be preliminarily compensated by a rectangular signal. The shortest sampling period of the new system represents 0.1 ms with a resolution higher than 0.1 K and sensitivity better than 30 mV/K.

Animals

[Successive detection of cortex oxygen partial pressure with oxygen microelectrodes in cats].

Brain tissue PO2 (PtO2) was successively measured with the new polarographic oxygen microelectrodes in the parietal cortex of ketamine anesthetized cats. This type of oxygen microelectrodes was made from pure platinum wire 0.5 microns in diameter pulled in glass with tip diameter of less than 1.0 micron. Results showed that PtO2 in the superficial gray matter (0-1,000 microns) was markedly higher than that in the white matter (1,001-3,000, 3,001-5,000 microns) in 10 cats. No significant change was observed during successive detecting of PtO2 within 3.0 hours in the cats. It was suggested that we can directly understand the oxygen supply and metabolism of living brain tissue by using the new oxygen microelectrodes.

Animals