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PubMed1990

Intracellular free magnesium in frog skeletal muscle studied with a new type of magnesium-selective microelectrode: interactions between magnesium and sodium in the regulation of [Mg]i.

The application of a new type of intracellular magnesium-selective microelectrode based on the neutral carrier ETH5214 to measure intracellular free magnesium ([Mg]i) in frog skeletal muscle fibers is reported. At room temperature (18-20 degrees C) the average values for [Mg]i was 0.93 mmol/l (pMgi = 3.03 +/- 0.42, SD; n = 38 experiments). The regulation of [Mg]i was studied by measuring [Mg]i and [Na]i with ion-selective microelectrodes during alterations of the membrane potential and the transmembrane sodium and magnesium gradients. Depolarization by increasing external [K] from 2.5 mmol/l to 12.5 mmol/l did not significantly influence [Mg]i. Increasing extracellular [Mg] from 1 mmol/l to 10 and 20 mmol/l caused a concentration-dependent rise in [Mg]i and a decrease in [Na]i, whereas removal of external magnesium did not affect [Mg]i. Removal of external [Na] caused an increase in [Mg]i and a decrease of [Na]i. The results show that [Mg]i in frog skeletal muscle is not in thermodynamic equilibrium and suggest that a Na/Mg exchange mechanism may be involved in maintaining low levels of [Mg]i.

Animals↗

PubMed1976

pH-sensitive glass microelectrodes and intracellular pH measurements.

1. Some properties of the open-tipped, uninsulated, pH-sensitive glass microelectrode were examined in several electrical experiments. 2. Based on these observations, technical and theoretical problems were considered for application to the pH measurement in small cells. 3. The intracellular pH, (pH)i, of the epithelial cell in rat duodenum measured was approximately 7.0. A reduction in (pH)i was apparent (about 0.3) with the addition of 20 mM-glucose to the bathing fluid. 4. It was concluded that with certain limitations such uninsulated, open-tipped microelectrodes may be successfully utilized for intracellular pH measurements.

Animals↗

PubMed1976

Studies on the origin of the tip potential of glass microelectrode.

1. Tip potential (TP) of glass microelectrodes filled with 3 M KCl increased remarkably with the increase in the storage period in 3 M KCl solution at 37 degrees C, while the electrode resistances decreased gradually. 2. The electrical conductivity through the thin glass wall near the tip was found to increase in parallel with the TP increase. 3. The e.m.f. across the thin glass wall in the tip region was directly measured. This seems to contribute to the TP generation of the microelectrode when the conductivity of the glass wall is significantly high in the tip region. 4. Effects of the acid treatment of glass employed and the acidification of fillant electrolyte solution suggested that fixed negative charges on the glass wall play a fundamental role in the TP formation. 5. Based on these experimental results, it was concluded that not only the diffusion potential through the tip pore but also the interfacial potential through the thin glass wall near the tip contributes to the TP generation, and the contribution of the latter increases with a long exposure period of the electrodes to electrolyte solution. 6. In this connection, technical problems related to reduction of the tip potential were also discussed.

Chlorides↗

PubMed1989

A new double-barrelled, ionophore-based microelectrode for chloride ions.

A new Cl- selective microelectrode based on the ionophore 5,10,15,20-tetraphenyl-21H,23H-porphin manganese(III) chloride is presented which discriminates better against HCO3- and several organic anions than electrodes containing the Corning 477913 ion-exchanger. Using a redesigned construction procedure, fine-tip double-barrelled microelectrodes were produced which had slopes of -52.4 +/- 0.6 mV (SE, n = 24), resistances of about 7.10(11) omega and a selectivity coefficient log KpotCLHCO3 of -1.40 +/- 0.03. Some electrodes showed a small unexplained sensitivity to pH greater than 7.6. When used to puncture cells of isolated S3 segments of rabbit renal proximal tubule during perfusion with HCO3- Ringer solution, the electrodes gave a membrane potential of -69.8 +/- 1.5 mV and an intracellular Cl- activity, [Cl-]i, of 35.3 +/- 2.6 mmol/l. Upon switching bath and lumen perfusions to Cl- -free solutions the "residual" [Cl-]i dropped to 1.20 +/- 0.03 mmol/l, while in similar measurements with ion-exchanger electrodes the "residual" [Cl-]i dropped only to 10.9 +/- 0.5 mmol/l. These observations demonstrate the superiority of the new electrode and prove that previously determined high [Cl-]i values in Cl- -free ambient solutions reflect interference problems rather than non-exchangeable intracellular chloride.

Animals↗

PubMed1984

Effect of external sodium on intracellular chloride activity in the surface cells of frog gastric mucosa. Microelectrode studies.

The intracellular chloride activity and its dependence on ionic substitutions in the bathing media was studied in individual surface cells of resting gastric mucosa using conventional and Cl- selective microelectrodes. When the tissue was perfused with control NaCl-Ringer the cell membrane p.d.'s, cell-lumen (psi cm) and cell-serosa (psi cs) were -40.9 +/- 0.6 mV and -66.8 +/- 0.5 mV (n = 175) respectively and the p.d. measured by the Cl- selective microelectrodes across the serosal membrane (psi csCl-) averaged -32.4 +/- 0.7 mV (n = 138). From these values an intracellular Cl- activity (acCl-) of 15.3 mmol/l can be estimated. The data indicate that chloride ion is distributed close to equilibrium at the luminal membrane while it is accumulated by an energy requiring step at the serosal membrane. Reduction (2 mmol/l) or absence of chloride from the luminal bath did not result in any detectable change of acCl-; on the other hand, after removal of Cl- from the serosal bath the intracellular Cl- activity fell to 7.1 mmol/l. When the tissue was exposed to serosal Na+-free Ringer (Na+ replaced by choline or TMA), although the acCl- remained unaffected, a marked reduction of the electrochemical gradient for Cl- at the serosal membrane was observed. These data indicate that: chloride is accumulated in the surface cells against its electrochemical potential difference at the serosal membrane; the luminal membrane has a negligible conductance to Cl-, while the serosal membrane represents a conductive pathway to chloride; the uphill entry of chloride at the serosal membrane seems to be, at least partially, Na+-dependent.

Animals↗

PubMed1980

Intracellular pH and surface pH in skeletal and cardiac muscle measured with a double-barrelled pH microelectrode.

The construction of a double-barrelled pH sensitive microelectrode for intracellular use is described. Repetitive measurements of intracellular pH were obtained in rat soleus muscle and sheep Purkinje fibres. They yielded pH values ranging between 7.1 and 7.2 in a CO2/HCO3- buffered medium at 37 degrees C. A lower pH value than that of the bulk solution was found at the surface of the cells using either double-barrelled or single-barrelled pH sensitive microelectrodes.

Animals↗

PubMed1979

Low impedance coaxial K+ selective microelectrodes.

A procedure for preparing coaxial K+ selective microelectrodes with a low longitudinal resistance (Re) of the liquid ion exchanger selective barrel is described. The low resistance was attained by inserting another microelectrode filled with 0.5 mol . 1-1 KCl into the ion-exchanger column (Corning 477317). The lower longitudinal resistance decreases the noise level and consequently increases the resolving power five times. This modification makes it possible to measure small and rapid K+ concentration changes.

Electrophysiology↗

PubMed1982

A simple method for constructing shielded, low-capacitance glass microelectrodes.

A new simple method is presented to produce shielded low-capacitance microelectrodes. A metal-shield is vapour-deposited on the inner surface of a glass pipette which is slid over the microelectrode proper and insulated at the tip by dipping in polystyrene. The unshielded protruding tip can be as small as 10 micrometers. A special advantage is the low capacitance between electrode and shield of approximately 0.16 microF/cm shield length.

Glass↗

PubMed1984

Low resting potentials in single isolated heart cells due to membrane damage by the recording microelectrode.

Single myocytes from adult rat hearts were prepared following the method of Powell and co-workers (9, 10, 11). Low resting potentials (Em) could be improved by three techniques. (i) Elevation of Cao to 7.2 mM which, however, mostly resulted in spontaneity and irreversible contracture. (ii) Pre-incubation in a "KB medium" (6). (iii) Use of suction pipettes instead of tapered microelectrodes for intracellular recordings (2). It is concluded that low Em measured previously (11) were due to membrane damage upon microelectrode impalement accompanied by insufficient healing of the membrane around the electrode insertion.

Animals↗

PubMed1988

Bias current modifies the selectivity of liquid membrane ion-selective microelectrodes.

A negative bias potential of up to -80 mV applied to the back of a liquid membrane ion-selective microelectrode containing classical "K+" ion-exchanger was found to make it more selective for millimolar concentrations of K+ over micromolar concentrations of choline, tetramethylammonium, tetraethylammonium and 5-hydroxytryptamine. Conversely, positive bias potential increased severalfold the sensitivity to micromolar concentrations of these ions while decreasing the sensitivity to K+. An increase in response amplitude for millimolar changes of ion concentration was also observed in neutral carrier electrodes for Na+, K+ and Ca2+ with negative bias potential. The various ions caused the resistances of the electrodes to change; these resistance changes contributed to the changes in response amplitude, but there were additional, unexplained, factors. The phenomenon was used to test if the signal from a K+ ion-exchanger microelectrode in extracellular space in bee retina was contaminated by substances other than K+.

Animals↗

PubMed1988

NH4+ ion-selective microelectrode based on the antibiotics nonactin/monactin.

A liquid-membrane microelectrode (less than or equal to 1 micron tip diameter) using macrotetrolide antibiotics as ion-selective components is described. The electrode shows selectivities of NH4+ over K+, Na+ and H+ of 3.8, 100 and 150, respectively. The stability and reproducibility of the sensor signal and the response time are determined in solutions with a typical intracellular ion background. The microelectrode does not suffer from significant interference by inorganic and organic inhibitors and lipophilic cations, but high concentrations of lipophilic anions may interfere considerably.

Anti-Bacterial Agents↗

PubMed1981

Some properties of KCl-filled microelectrodes: correlation of potassium "leakage" with tip resistance.

This study was undertaken in order to determine directly the rates of K leakage (JK) out of the tips of microelectrodes into a solution of 100 mM KCl (approximating the K concentration of the cell interior) and to relate these rates to the concentration of the filling solution and the tip resistance. The values of JK for electrodes filled with 3 M KCl having resistances of 16 and 30 M omega (when measured in 3 M KCl) were 10 and 5.5 fmol/sec, respectively. When the same electrodes were filled with 0.5 M KCl, the resistances (measured in 0.5 M KCl) increased to 62 and 115 M omega, respectively, and JK fell to 1.8 and 1.0 fmol/sec, respectively. These values are in reasonable agreement with what would be expected from theoretical considerations if leakage of KCl were the result of diffusion plus convective flow due to the hydrostatic pressure of the filling solution. We conclude that K leakage out of microelectrodes filled with 3 M KCl is unnecessarily high; leakage can be reduced fivefold by filling electrodes with 0.5 M KCl without incurring significant increases in tip or diffusion potentials or unmanageable tip resistances. Finally, the lowest rate of K leakage observed (1 fmol/sec) is still very considerable for the case of animal cells with an intracellular volume of approximately 1 pl and a K content of approximately 100 fmol. The finding of stable intracellular potentials, often for many minutes, in some tissues suggests that K which enters the cell rapidly diffuses into neighboring cells via high conductance intercellular communications.

Diffusion↗

PubMed2001

Voltammetric determination of iodide by use of an interdigitated microelectrode array.

An interdigitated microelectrode array (IDA) sensor has been applied to the determination of iodide in mineral water. It is based on reversible charge transfer in the redox system I2/2I- at a platinum microelectrode. The analytical signal from the IDA system was obtained by use of a bipotentiostat in dual mode. One segment of the IDA (generator) was polarized to the limiting current for oxidation of iodide to iodine in 0.1 mol L(-1) HClO4. The second segment (collector) was fixed at a potential value corresponding to the limiting current of iodine reduction. The geometrical arrangement of the IDA enables this transfer with high efficiency. Because the diffusion layer of both segments overlaps the iodide produced on the collector, the iodide diffuses back to the generator where it is reoxidized. Therefore, redox cycling will enhance the voltammetric signal of the IDA. The signal obtained with a vertically separated IDA was 20 times higher than that in the single mode. Because multiplication of the signal reduces the detection and determination limits, direct voltammetric determination of iodide in mineral water is possible.

Iodides↗

PubMed1999

A neutral carrier-based liquid membrane microelectrode for divalent putrescine cations.

A new ion-selective liquid membrane microelectrode, based on the neutral carrier 1,1'-bis(2,3-naphtho-18-crown-6), is described that shows the dependence of EMF on the activity of divalent putrescine cations a(Put), with the linear slope s(Put) = 26 +/- 3 mV/decade (mean +/- SD, N = 18), in the range 10(-4)-10(-1) M at 25 +/- 1 degrees C. Values of potentiometric putrescine cation selectivity coefficients of logK(Pot)(Put) (j) (mean +/- SD, N) are obtained by the separate solution method for the ions K(+) (1.0 +/- 0.4, 10), Na(+) (-1.2 +/- 0.4, 8), Ca(2+) (-2.3 +/- 0.5, 10) and Mg(2+) (-2.5 +/- 0.5, 7). The microelectrode can be applied for the direct analysis of the activities of free divalent putrescine cations in the range 5 x 10(-4) to 10(-1) M in an extracellular ionic environment. Established analytical methods, e.g. high performance liquid chromatography, determine the total concentration of the derivatives of free and bound putrescine.

Cations, Divalent↗

PubMed2006

Multicellular recordings of cultured brainstem neurons in microelectrode arrays.

Several vital systemic functions are controlled by the brainstem, which has been studied in a variety of experimental preparations and by various techniques, including in-vitro electrophysiological preparations. Although these in-vitro approaches have greatly advanced the understanding of brainstem neurons, most recording methods with microelectrodes and patch pipettes are invasive. To take advantage of in-vitro approaches but avoid their potential problems, we have studied brainstem neurons in microelectrode arrays (MEA). Neurons were isolated from the medulla oblongata and cultured in DMEM. Extracellular recordings were performed with no evident perturbations to the cellular environment. Neurons started firing after 24-48 h in culture, reached stable activity in 3-4 weeks, and retained this activity for at least 3 months. From their firing patterns, these neurons could be divided into tonic and bursting units. The latter could be further divided into regular and irregular bursters based on their burst intervals. Cells were stimulated or inhibited by exposure to 10% CO2. The stimulatory effect of CO2, though smaller, was still seen after selective ablation of serotonergic neurons or with low Ca++ and high Mg++ in the extracellular medium. Similar treatments had no significant effect on CO2-inhibited units. The abundance of units with respect to their firing patterns and CO2 responses, together with the long-term stable non-invasive recordings with no evident perturbation to cellular environments, suggests that MEA represent another promising in-vitro approach for studying brainstem neurons.

Animals↗

PubMed1985

Quantitative determination of methylamines using microelectrodes.

A new method for measuring methylamino compounds such as choline, trimethylamine, trimethylamine-N-oxide, betaine, L-carnitine, and dimethylamine is described. A glass microelectrode is used to quantify methylamines in concentrations ranging from 0.1 to 10.0 mM. Rapid time response and a good sensitivity are maintained by the microelectrode even when measurements are performed in solutions having high ionic strength and low pH. These characteristics make this assay suitable for use with conventional column chromatographic techniques of separation for these methylamines.

Calibration↗

PubMed1977

A new solid-state microelectrode for measuring intracellular chloride activities.

Solid-state microelectrodes from measuring intracellular Cl activity (alphaiCl) were made by sealing the tips of tapered glass capillaries (tip diameter 0.3 mum), coating them under vacuum with a 0.2-0.3 mum thick layer of spectrscopic grade silver, and sealing them (except for the terminal 2-5 mum of the tip) inside tapered glass shields. 106 microelectrodes had an average slope of 55.0+/- 0.6 m V (S,E,) per decade c hange in alphaCl. Tip resistance was (77.1+/- 3.1) x 10(9) omega(n=30). Electrode response was rapid (10-20 s), was unaffected by HCO3, H2PO4, HPO42 or protein, and remained essentially unchanged over a 24-h period. AlphaiCl in frog sartorius muscle fibers and epithelial cells of bullfrog small intestine was measured in vitro. In both tissues, alphaiCl significantly exceeded the value corresponding to equlibrium ditribution of Cl across the cell membrane.

Animals↗

PubMed1980

Neutral carrier ion-selective microelectrodes for measurement of intracellular free calcium.

This paper describes the making and testing of calcium-selective microelectrodes for measurement of intracellular [free Ca2+] levels. Pipettes of tip outer diameter down to 0.4 micron were siliconized by a novel and easy method of vapor treatment. The tips were filled with a sensor mixture using the neutral ligand and solvent of Oehme et al. (Oehme, M., Kessler, M. and Simon, W. (1976) Chimia (Aarau) 30, 204-206) but with very hydrophobic cations replacing Na+ in the salt component. This change improved electrode stability and greatly reduced hysteresis in the responses to changing [Ca2+] levels. Lowering the Ca2+ concentration in the internal electrolyte also increased electrode lifetime. Electrodes showed a Nernstian response to [Ca2+] down to 1 micro M free concentration in 0.1 M KCl, and usually a useful response to below 100 nM Ca2+. Selectivity for Ca2+ over Mg2+ and H+ was sufficiently high that typical free intracellular levels of Mg2+ and H+ caused negligible interference. Selectivity for Ca2+ over Na+ was adequate for cells with 10(-2) M free Na+, but higher levels could raise significantly the detection limit for Ca2+. Preliminary measurements of [free Ca2+] have been made in frog skeletal muscle, ferret ventricular myocardium, and early embryos of Xenopus laevis. Relative merits of Ca2+ microelectrodes and optical indicators are discussed.

Animals↗