Effect of Sulfhydryl Reagents on the Biophysical Properties of the Plasmalemma of Chara corallina

Abstract

The administration of the sulfhydryl reagent N-ethyl-maleimide (NEM) to internodal cells of Chara coralina caused alterations in the biophysical properties of the plasmalemma, as measured with electrophysiological and radioactive tracer techniques. The membrane potential depolarized to, or near, the calculated Nermst potential for potassium (EK) after 30 seconds' exposure to 0.1 millimolar NEM. During this time, the ATP level did not decrease below the control value, and the specific membrane resistance did not increase; only upon further exposure to NEM did the resistance approach the value observed in the dark. In the depolarized state, the membrane potential responded to changes in the external potassium concentration in the manner of a K -electrode, but it retained it's relative insensitivity to external sodium.These results are interpreted in the following manner. NEM causes a) an increase in the membrane permeability to K+ (ie. an increase in K+ conductance); and b) perturbation of the electrogenic transport system(s) of the plasma membrane. The latter effect is manifested in a manner that is not consistent with an inhibition of ATP catalysis by a voltage-dependent ATPase possessing conductance. The nonpermeant sulfhydryl modifier, pchloromercuribenzenesulfonic acid, appeared to affect membrane properties in a similar, but reversible, way.The biochemical and biophysical properties of the plasmalemma play an undisputed key role in the organization and function of membrane transport phenomena. Increasingly, heavy metals and sulfhydryl reagents have been used to probe the interrelationships of these processes (4-6, 12, 18, 31 amino-1,3-naphthalene disulfonic acid; Rm, specific membrane resistance (K _-cm2); gm, total membrane conductance; PK, potassium permeability. tion of the external medium. Because the electrophysiology and ionic relations of Chara corallina are relatively well understood and since certain transport processes in this alga appear to be differentially susceptible to -SH reactive agents (18), we investigated the biophysical properties in C. corallina before and after treatment with thiol-modifying reagents. This approach has allowed us to conclude that sulfhydryl reagents affect the passive permeability of the membrane by increasing potassium conductance and act upon the electrogenic transport processes of the cell membrane. Experimental Solutions. Solutions were prepared using high purity glass-distilled H20. The CPW contained 1.0 mm NaCl, 0.2 mm KCI, 0.2 mm CaSO4, 1.0 mm TAPS (pH 8.0); modifications to this medium will be detailed in the text. Bicarbonate-free solutions were prepared by bubbling solutions with air that had been passed through C02-absorbent material (Ascarite; Arthur H. Thomas Co.). All chemicals were of Analar reagent grade and, except for PCMBS, were not purified further. Stock (100 mm) NEM solutions were titrated to pH 7, using fresh NaOH solution, and stored at 4 C for a maximum of 4 days. Because hydrolytic ring opening in the NEM molecule occurs at alkaline...

Potassium Transport in Corn Roots

“…Thus, this NEM-induced reduction in the linear component is still consistent with uptake by facilitated diffusion. However, PCMBS has also been shown to depolarize the membrane potential to the K+ diffusion potential (22,23), and PCMBS had no effect on the linear component in high-salt roots, while consistently stimulating the linear component by 40% in low-salt roots.…”

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“…A brief exposure of Chara cells to NEM results in an opening of K+ channels within the plasmalemma (12). Thus, in our NEM-pretreated cells, the membrane potential would have been depolarized from -250 mv down to -150 mv (12).…”

“…8) shown to open the K+ channels in the plasmalemma of Chara (12). Since Lichtner et al (12) did their experiments at pH 8.0, we exposed cells to NEM at both pH 8.0 and 6.5. Cells that were given 1-or 2-min pretreatment in 0.1 mm NEM had 'Cl-influx values similar to those of the controls (Table II).…”

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“…4), perhaps by inhibiting an ATPase proton pump in the same way it affects uptake by plant cells (Giaquinta, 1979;Lichtner, Lucas & Spanswick, 1981). Indeed Beale (1992) has shown that PCMBS is a potent inhibitor of ATPase in a plasma membrane fraction from Phytophthora cactorum (Lebert & Cohn), a related member of the Peronosporales.…”