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Microelectrode study of insulin effect on apical and basolateral cell membrane of frog skin: comparison with the effect of 1-deamino-8-D-arginine-vasopressin (dDAVP).

The standard microelectrode technique was used to electrophysiologically characterize the effect of insulin on both apical and basolateral cell membranes of the frog skin, a model Na-transporting epithelium. Insulin applied to the basolateral side of the epithelium stimulated the sodium transport as shown by both increased short-circuit current, Isc, (P less than 0.02) and transepithelial potential difference, Voc, (P less than 0.002). Potential difference across the apical cell membrane, Vo, decreased (P less than 0.002) as did the apical cell membrane resistance, Ro, (P less than 0.05). The driving force of sodium ions, ENa, increased after insulin (P less than 0.005). These findings confirm that insulin acts both to increase the apical cell membrane permeability for ions and to stimulate the sodium pump in the basolateral membrane. The effects of insulin were compared with those of a vasopressin analog (dDAVP), known to stimulate transepithelial sodium transport by increasing the permeability of the apical cell membrane for sodium ions. dDAVP applied at the height of insulin effect further stimulated transepithelial transport, but insulin applied at the height of dDAVP action did not. It is concluded that the direct stimulation of the sodium pump by insulin may not represent a decisive component in the stimulation of transepithelial transport across the frog skin. A more potent stimulus for sodium transport is obviously the increased permeability of the apical membrane for ions.

Animals↗

[A method of rapid beveling of microelectrodes using vibration].

The new technique is suggested for a rapid bevelling of glass and metallic microelectrodes by means of ends' friction on the fine-grained surface of a quartz plate vibrating at a frequency of the mechanical resonance of the system.

Electrophysiology↗

A microelectrode study of the effects of atracurium on neuromuscular transmission.

Using standard microelectrode recording techniques, we studied the effects of atracurium on neuromuscular transmission in a concentration range between 10(-13) and 10(-5) M in the rat diaphragm. Both intact and cut diaphragm preparations were used. Atracurium produced no significant alteration of miniature endplate potential (MEPP) frequency (P greater than 0.05). Increasing concentrations of atracurium (10(-6)-10(-5) M) caused a linear decrease in MEPP amplitude from 70.5 +/- 2.3% to 28.0 +/- 2.5% of baseline levels (P less than 0.05). In the cut diaphragm preparation, atracurium increased the degree of rundown and decreased quantum content of the endplate potential (EPP). The above observations suggest that atracurium interferes with neuromuscular transmission by, first of all, producing cholinergic receptor block and secondly, producing frequency-dependent inhibition of release of acetylcholine from the nerve terminal.

Animals↗

[Intracellular microelectrode study of the electrophysiological effects of contrast media].

Intracellular microelectrodes were used to study the electrophysiological effects on action and resting potentials, refractory periods and conduction velocities of four contrast media administered either as 1/100 and 1/1000 solutions or as 20 mg bolus injections. The products studied were Telebrix, which has high osmolality (2100 mOsm/kg) and three other contrast media with lower osmolatity (700 mOsm/kg), including Hexabrix, which contains sodium, and Iopamidol and Iohexol which are sodium-free. The parameters measured were not modified by Hexabrix and Telebrix, whereas Omnipaque and Iopamiron induced varied and significant changes in these parameters, notably in concentrations of 1/100 and as bolus injections. It is suggested that the latter contrast media should be used in non-cardiovascular explorations and in patients without confirmed heart disease.

Action Potentials↗

[A floating hydraulic micromanipulator for microelectrode research on brain neurons].

The article describes a simple floating hydraulic micromanipulator of original construction which works in positive-positive pressure and provides maximum stability of the microelectrode tip concerning the neuron in the moving brain. This work is achieved by fixing of the micromanipulator on the hinged suspension bracket and by setting of it on the surface of the object with the help of bearing site. Breathing and vascular brain moving don't influence the neuron activity lead.

Animals↗

Identification of the two cell groups in the longitudinal muscle layer of the earthworm Amynthas hawayanus by a single microelectrode with lucifer yellow-CH.

Previous electrophysiological studies have suggested the presence of two types of muscular cells in the longitudinal muscle of A. hawayanus. However, a definite classification of such cells as muscles has been hindered by lack of morphological evidence. This study identified these cells electrophysiologically and morphologically by using the same intracellular microelectrode to record cell activity and inject Lucifer Yellow-CH. Electrophysiological measurements clearly differentiated the two groups of cells. One group did not respond to nerve stimulation but was spontaneously active, whereas the other responded to nerve stimulation with endplate potentials and did not discharge spontaneously. Morphological examination revealed two very similar types of longitudinal muscle cells. These results clearly show that, although morphologically similar, there are two distinct groups of muscle cells in the longitudinal muscle layer of the earthworm.

Animals↗

Effect of bevelling on C1-selective microelectrodes: experimental and model study.

Bevelling of the ultra-fine tips of C1-selective microelectrodes (ME) based on liquid ion-exchanger Corning 477913 caused significant decrease in ME resistivity, increase in sensitivity dependent on its initial value and decrease in selectivity constant for bicarbonates (KC1,HCO3) from 10:1 to approximately 8:1. Bevelling did not affect KC1,NO3 which had value below 1 due to higher selectivity of the ion-exchanger for nitrates. The observed experimental alterations in ME behavior after bevelling were in agreement with those predicted by a relatively simple electrical model comprising of two conductive pathways in parallel having opposite in sign sensitivities.

Chlorides↗

[Microelectrode study of the cellular reactions of the taste bud in the frog Rana temporaria].

Microelectrophysiological studies reveal two types of cells in the taste bud of frog which differ by the level of their membrane potential. During vertical implantation of microelectrode through the apical part of the taste bud, the potential difference in the upper layer amounts to 15 mV. Further implantation of the electrode results in a stepwise decrease of the potential difference up to 27 mV. Cells of the deeper layer are located 12-24 micron lower from the apical surface. Stimulation of cells by solutions of chemical substances is accompanied by cell depolarization, its amplitude being proportional to stimulus concentration. The steepness of depolarization depends on the modality of the stimulus, being maximum for salts. The data obtained suggest that cells of the second layer, with a higher resting membrane potential level, are taste ones.

Animals↗

[Injection of glyoxylate ions from the recording microelectrode for the fluorescent microscopic detection of neuronal monoamines].

To make it sure that a physiological experiment is carried out on the monoaminergic neurone, we propose to fill the intracellular recording glass microelectrode with 2 M solution of potassium glyoxylate, with pH 6.0. After the experiment is over, glyoxylate ions are injected ionophoretically into the neurone, and then the specimen is dried in the air and heated to obtain fluorescence of a cellular monoamine, if it is present in the cell. Some details of the method are described.

Animals↗

Identification of membrane transport processes in renal cells, by means of liquid ion exchanger microelectrodes.

The development of liquid-ion-exchanger microelectrodes displaying tip diameters less than or equal to 1 micron has permitted direct measurement of the transmembrane chemical and electrochemical gradients of several permeant ion species in cells of small size. We have used Cl- and H+ resins to study the intracellular Cl- activity (alpha iCl) and cell pH (pHi) in the proximal tubule of Necturus kidney. These determinations were performed in association with perfusion of peritubular capillaries by several artificial solutions, in order to assess the dependence of alpha iCl and pHi on the composition of physiologic plasma constituents and selected inhibitors. The main findings are: Intracellular chloride activity, alpha iCl, is higher than the theoretical value predicted from electrochemical equilibrium. Peritubular application of SITS resulted in a decrease of alpha iCl and increase of pHi; these observations are taken to indicate that Cl- uptake is achieved across the basolateral membrane in exchange for HCO-3 by a mechanism sensitive to SITS. Na+ removal from peritubular fluid elicited a small reduction of alpha iCl, suggesting the presence of carrier-mediated Cl--Na+ cotransport from interstitium to cell, contributing to the rise of alpha iCl above equilibrium. In conclusion, two carrier-mediated processes (Cl-/HCO-3 exchange and Cl--Na+ symport) located at the basolateral membrane of the proximal tubule may account for the establishment of alpha iCl values above equilibrium, at steady state. The physiologic role of these carriers is discussed in relation to proximal electrolyte absorption.

4-Acetamido-4'-isothiocyanatostilbene-2,2'-disulfo↗

Triple-barrelled ion-sensitive microelectrode for simultaneous measurements of two extracellular ion activities.

A method has been described by which triple-barrelled ion-sensitive microelectrodes for simultaneous measurements of two extracellular ion activities could be produced with a tip diameter of 1-2 microns. The following combinations with two ion exchangers could be achieved: Ca++/K+, Ca++/Cl-, K+/Na+. The mean steepness for the potassium electrode was 55.8 mV, for the calcium electrode 26 mV, for the chloride electrode 48.8 mV, and for the sodium electrode 55.2 mV. The change of steepness varied 4%-5%/day and some of the electrodes could be used up to 5 days. The mean response time for all electrodes did not amount to more than 200 ms. An example is given for simultaneous in vivo measurement of both extracellular potassium and calcium activity.

Calcium↗

pH distributions in transplanted neural tumors and normal tissues of BDIX rats as measured with pH microelectrodes.

The distribution of pH values was measured in transplanted neuroectodermal TV1A tumors and in brain and kidney of BDIX rats in vivo. Tissue damage during pH measurements could be minimized by the use of Hinke-type pH glass microelectrodes with maximum diameters of the pH-sensitive tips of less than or equal to 10 micrometers (sensitivity, 58 to 60 mV/pH unit at 37 degrees; response time (95%), less than or equal to 3 sec; drift, less than or equal to 0.01 pH unit/hr). The advantages and limitations of this technique are discussed in relation to other methods for the analysis of extracellular pH. In tumors weighing 1.0 to 2.5 g, pH values ranged from 6.8 to 7.1 (mean, 7.0). The pH distribution in tumors weighing 4 to 6 g was shifted to slightly lower values, with an average pH of 6.9 (range, 6.7 to 7.1). No marked pH differences were found between the tumors and normal tissues. The pH values measured in brain and kidney ranged from 6.6 to 7.3 (mean, 7.0) and 6.7 to 7.3 (mean, 7.1), respectively. Within single tumors, local pH variations in the range of 0.2 to 0.3 pH unit were observed. The local pH values measured in certain tumor areas are, however, sufficiently low to partially inhibit proliferation and colony formation in cultured malignant cells.

Animals↗

Double-barrell ion-sensitive microelectrodes with extra thin tip diameters for intracellular measurements.

A method has been described by which double-barrel ion-sensitive microelectrodes for intracellular measurements can be produced with a mean tip diameter of 0.1 micron. Potassium, calcium and chloride activity could be measured with these electrodes by means of ion exchangers and neutral ion carriers. The mean steepness for the potassium electrode is 50.16 mV, for the calcium electrode 24.7 mV and for the chloride electrode 39 mV. the drift of the electrodes varies between 2-4%/h. The mean response time amounts for the potassium electrode to 170 ms, for the calcium electrode to 350 ms and for the chloride electrode to 250 ms. With these electrodes it is possible to measure intracellularly cell membrane potential and ion activity simultaneously.

Action Potentials↗

Analysis of electroretinogram during systemic hypercapnia with intraretinal K(+)-microelectrodes in cats.

PURPOSE: Systemic hypercapnia alters the intraretinal pH and metabolism and profoundly affects the vitreal electroretinogram (ERG). Using intraretinal K(+)-selective microelectrodes (ISMs), the authors examined the effects of systemic hypercapnia on ERG in the intact cat eye in vivo. METHODS: Double-barreled K(+)-ISMs were used to record intraretinal, light-evoked [K+]o decrease, intraretinal ERG, vitreal ERG, and standing potential (SP) in the dark-adapted retina. Hypercapnia was induced by adding CO2 to the inhaled mixture of gas (without significant hypoxia). RESULTS: The standing potential was elevated, and the vitreal ERG c-wave increased in amplitude whereas the b-wave decreased during hypercapnia. Intraretinal recordings showed that the amplitudes of slow PIII and the transepithelial (TEP) c-wave decreased. The slow PIII decreased more than the TEP c-wave. The light-evoked [K+]o decrease also was reduced, but no change was noted in the subretinal [K+]o. CONCLUSION: The neural retina was more sensitive than the retinal pigment epithelium to the intraretinal acidification caused by systemic hypercapnia. However, the change in the SP and b-wave suggested complicated effects other than acidosis.

Animals↗

[Measurement of cytoplasmic Ca2+ activity in myocardia with Ca(2+)-sensitive microelectrode].

The improved neutral ligand ETH1001 Ca(2+)-sensitive microelectrodes (Ca-ISE, tip diameter 0.4-0.8 mumol.L-1) are highly ion selective and sensitive and therefore can be reliably used to measure cytoplasmic Ca2+ activity (alpha iCa) in myocardia. The resting alpha iCa in guinea-pig ventricular trabecula, canine ventricular myocardia and Purkinje fibers were respectively 0.19 +/- 0.01, 0.20 +/- 0.02 and 0.46 +/- 0.07 mumol.L-1. Three mumol.L-1 strophantin G increased the resting and dynamic myocardial alpha iCa by 0.18 +/- 0.02 and 6.69 +/- 2.09 mumol.L-1 respectively, as well as engendered triggered activity (TA). When pretreated with 100 mumol.L-1 dauricine (Dau), strophantin G could no longer increase alpha iCa and TA disappeared. It is suggested thus the applicability of Ca-ISE for measurement of myocardia alpha iCa and TA is obvious.

Alkaloids↗

Voltage clamping of a frog (Rana catesbeiana) taste cell with a single microelectrode.

1. Receptor currents in frog taste cells induced by taste stimuli of NaCl and HCl were studied in situ using a voltage clamp method with a single microelectrode. 2. Under voltage clamping of a taste cell at -20 mV an inward receptor current of a mean of 176 pA was induced by taste stimulation with 0.5 M NaCl and the input resistance was largely reduced during the stimulation. 3. A large sustained inward current was evoked by HCl stimulation but the input resistance was slightly reduced. 4. Increasing the holding potential in the negative direction linearly increased the inward receptor current by NaCl stimulation but did not remarkably change it by HCl stimulation.

Animals↗

Intraretinal study of cat electroretinogram during retinal ischemia-reperfusion with extracellular K+ concentration microelectrodes.

PURPOSE: Intraretinal electroretinograms (ERG) during retinal ischemia-reperfusion were analyzed to clarify which cells are affected in the cat dark-adapted eye. METHODS: Ischemia was induced by raising the intraocular pressure above arterial systolic pressure in vivo. Double-barreled microelectrodes were used to record the intraretinal ERGs and the light-evoked [K+]o decrease in the subretinal space. Vitreal ERGs, the standing potential and the transepithelial resistance were also recorded. RESULTS: After 10 minutes of ischemia, vitreal ERG b- and c-waves had recovered fully in 2 hours, and after 30 minutes of ischemia, recovered approximately 85% and 77% of their control amplitudes in 4 hours, respectively. At the early period of reperfusion after 10 minutes of ischemia we observed the supernormal c-wave (124% of its control amplitude, n = 10) with an elevated standing potential. The transepithelial potential (TEP) c-wave increased without changes of slow PIII and transepithelial resistance when the supernormal amplitudes occurred. The light-evoked [K+]o decrease in the subretinal space (90% retinal depth) in the area centralis recovered to about 90% of the control amplitude in 2 hours, and after 30 minutes of ischemia recovered to about 70% of the control amplitudes in 4 hours. The recovery courses of the slow PIII and TEP c-wave were almost the same as that of the light-evoked [K+]o decrease. CONCLUSIONS: The temporary supernormal amplitude of the ERG c-wave was originated from the retinal pigment epithelium (RPE) during the early period of reperfusion after 10 minutes of ischemia. The photoreceptors, Muller cell and RPE are responsible for the changes in the light-evoked [K+]o decrease, a slow PIII and TEP c-wave, respectively. Intraretinal recordings suggested that after ischemia of 10 to 30 minutes duration, the responses of photoreceptors, Muller cells and the RPE recovered with a similar time course. These findings suggest that in all cases, primarily photoreceptors were damaged.

Animals↗

[Measurement of human erythrocyte membrane potential with microelectrode].

The measurement of erythrocyte membrane potential (Ery-MP) in 189 healthy subjects using a microelectrode showed that the Ery-MP in 96 healthy subjects was -5.80 +/- 0.31 mV, and that in 93 healthy female subjects was -6.30 +/- 0.34 mV. Plasma lipid peroxide (P-LPO), erythrocyte lipid peroxide (Ery-LPO), plasma superoxide dismutase activity (P-SODA) and erythrocyte superoxide dismutase activity (Ery-SODA) were measured at the same time. After linear regression and correlation analysis, it was found that the Ery-MP value of healthy adults decreased as the P-LPO and Ery-LPO values increased, displaying a linear negative correlation, Ery-MP and Ery-SODA values formed a linear positive correlation.

Adolescent↗