Potentials in frog cornea and microelectrode artifact

Tl, Davis; Jw, Jackson; Be, Day; Rl, Shoemaker; Rehm, Warren S.

doi:10.1152/ajplegacy.1970.219.1.178

Cited by 21 publications

(12 citation statements)

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“…In these experiments 0.1 M KC1 was chosen as a standard because of the small difference in the mobilities of the potassium and chloride ions, and because it is tolerated by the spherical cells. Also, the near-isotonicity of the solution was expected to minimize any matrix or "pre-tip" potential artifacts (Davis et al 1970;Nelson et al 1978). Table 4 summarizes the results of several simultaneous measurements with 0.1 M KC1 and a test electrolyte as well as similar measurements in which both electrodes (or barrels) were filled with identical solutions.…”

Section: Electrode Tip Potentialsmentioning

confidence: 99%

“…Large cell volumes will act to buffer the effects of electrolyte leakage and, in impalements of plant cells (which generally result in penetration of the central vacuole), the tonoplast could provide a barrier between the cytoplasm and salt from the electrode tip. However, most animal cells are small, with volumes of 20 pl or less, and in several cases are known to be subject to salt leakage artifacts (Nelson et al, 1978, Davis et al, 1970. Comparably sized plant cells, such as stomatal guard cells, also occur, the tonoplast properties of which remain uncertain (Moody & Zeiger, 1978).…”

Section: Rt [_ J Rt"mentioning

confidence: 99%

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KCl leakage from microelectrodes and its impact on the membrane parameters of a nonexcitable cell

1983

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Summary. Microcapillary electrodes filled with a variety of salt solutions, including 1 M KC1, have been used to measure the membrane potentials and resistances of spherical cells from the mycelial fungus Neurospora (cell diameters 15-25 gm, cell volumes 3-8 pl). During impalements with electrodes containing 0.3-1.0 M KC1, membrane potential and resistance decayed over a period of 3-10 min. In contrast, electrodes filled with 0.1 M KC1 gave stable membrane potentials of -180 mV and membrane resistivities of 40 k~ cm 2, values comparable to earlier results from the fungal hyphae.Salt leakage from 1.0 M KCl-filled electrodes (tip diameters 0.2-0.3 pro, resistances 50-75M~2) occurred at rates of 4~5 fmol sec-1, as indicated by direct intracellular measurements with ion-sensitive microelectrodes. Depending on cell size, such leakage rates could elevate cytoplasmic KC1 content at initial rates of 30 170 mM min -1, and actual values as high as 70 mM min-1 were observed. Salt leakage and changes in cytoplasmic KC1 concentration were reduced five-to sevenfold when impalements were made with electrodes containing 0.1 M KC1.The effects on cell membrane parameters of salt leakage from microelectrodes could be attributed to chloride ions. Substitution of the KC1 electrolyte with half-molar K2SO4 or NazSO4 and molar concentrations of K-and Na-MES [potassium and sodium 2-(N-morpholino)ethanesulfonate] gave stable membrane potentials in excess of -200 mV and membrane resistivities greater than 50 k~ cm 2, while the permeant anions NO~-and SCN-depressed the membrane parameters in a manner similar to that observed with 1 M KC1. Furthermore, modest elevation of cytoplasmic chloride concentration (below ca. 50 mM) affected both membrane potential and resistance in direct proportion to the concentration, and could be quantitatively described by the Constant Field Theory with a fixed membrane permeability (Pc1 ~ 4 x 10-8 cm sec-1). Higher cytoplasmic chloride levels produced a collapse of the membrane resistance and drastic depolarization in a fashion requiring large changes of membrane permeability.At least for cells with volumes of 10 pl or less, the standard practice of filling electrodes with 1 or 3 M KC1 should be abandoned. Half-molar (and lower) concentrations of K2SO 4 or Na2SO 4 are suggested as satisfactory replacements.

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Section: Electrode Tip Potentialsmentioning

confidence: 99%

Section: Rt [_ J Rt"mentioning

confidence: 99%

KCl leakage from microelectrodes and its impact on the membrane parameters of a nonexcitable cell

1983

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“…They reported that the stroma was negative to the solution bathing both surfaces and suggested that the C1-transport mechanism was located in the endothelium. Controversial opinions regarding the stromal negative level are mentioned in literature [2,10,12,14]. Davis et al [10] using glass microelectrodes filled with 3 M KC1 or Ringer's solution suggested that the negative stromal potential could be a KC1 junction potential, being the consequence of the presence of negative fixed charges in the stroma, possibly due to existence of acidic polysaccharides in that region.…”

Section: Discussionmentioning

confidence: 99%

“…This idea is reinforced by the observation that leakage of KC1 from broken microelectrodes into the external solution induces a rapid disappearance of PDi, which is restored by repeated changes of the external solution. Therefore, based on the previous results, a working hypothesis, analogous to that formulated by Davis et al [10] to explain a potential level in the stroma of frog cornea, was put forward to account for the genesis of PDi. It was assumed that PDi is a KC1 d{ffusion potential generated by diffusion of this salt from the tip of the microelectrode into the matrix of the stratum corneum, where K ions would have a higher mobility than C1 ions (at the high pH used in the eXperiments), probably as a consequence of negative fixed charges in the matrix of the stratum corneum.…”

Section: Substitution Of Na By K In the External Solutionmentioning

confidence: 99%

Negative potential level in the outer layer of the toad skin

Nunes

Vieira

1975

J. Membrain Biol.

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The isolated skin of the toad Bufo marinus ictericus when impaled from the outer surface by glass microelectrodes filled with 3 M KCl shows a voltage profile which is a continuous function of the depth of impalement. The superficial intraepithelial potential difference measured with reference to the external solution (PDi) is negative with NaCl-Ringer's solution on both sides of the skin, displaying a minimum of -26.7+/-3.6 mV at 6+/-2 mum. Null value is obtained at 19+/-3 mum, with positive values for deeper impalements. Indications of cell impalements (abrupt voltage and resistance jumps) were frequently observed at sites deeper than 25 mum from the outer surface. Measurements of the electrical resistance between the microelectrode and the external solution, made with single- and double-barreled microelectrodes, showed great discrepancies, which may be attributed to distinct pathways of different resistances in the stratum corneum. PDi measured at a depth of 5 mum was a logarithmic function of Na2SO4 or K2SO4 concentration in the external solution, increasing in negativity with a reduction in concentration. Substitution of Na by K in the external solution had only minor effects on PDi. Acidification of the external solution from pH 9 is accompanied by a reduction in the negative value of PDi. At pH 3 PDi was positive. PDi was interpreted as a diffusion potential at the tip of the microelectrode due to KCl diffusion from the electrode into the matrix of the stratum corneum. Differences in K and Cl mobilities, responsible for the origin of PDi, were attributed to fixed charges in the matrix of the stratum corneum, with density and polarity determined by their degree of proponation, controlled by the hydrogen ion concentration of the external solution. Skin potential, short-circuit current and their relationship to PDI were discussed.

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“…An important reason for the genesis of large emf's at the level of the microelectrode tip is its location within a fixed-charge bearing matrix, as is the case of cells of the stratum corneum [15] or the stroma of connective tissue [5]. The electrical potentials observed with 3 M KC1 microelectrodes in the cells of the stratum corneum were interpreted as due to KC1 diffusion from the microelectrode into the fixed-charge bearing protein matrix with K § and CI-transference numbers controlled by the matrix degree of protonation [-10, 15].…”

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confidence: 99%

Biological and artificial ion exchangers: Electrical measurements with glass microelectrodes

1978

J. Membrain Biol.

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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.

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Potentials in frog cornea and microelectrode artifact

Cited by 21 publications

References 0 publications

KCl leakage from microelectrodes and its impact on the membrane parameters of a nonexcitable cell

KCl leakage from microelectrodes and its impact on the membrane parameters of a nonexcitable cell

Negative potential level in the outer layer of the toad skin

Biological and artificial ion exchangers: Electrical measurements with glass microelectrodes

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