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Calcium channels in the plasma membrane of root cells fulfill both nutritional and signaling roles. The permeability of these channels to different cations determines the magnitude of their cation conductances, their effects on cell membrane potential and their contribution to cation toxicities. The selectivity of the rca channel, a Ca2+-permeable channel from the plasma membrane of wheat (Triticum aestivum L.) roots, was studied following its incorporation into planar lipid bilayers. The permeation of K+, Na+, Ca2+ and Mg2+ through the pore of the rca channel was modeled. It was assumed that cations permeated in single file through a pore with three energy barriers and two ion-binding sites. Differences in permeation between divalent and monovalent cations were attributed largely to the affinity of the ion binding sites. The model suggested that significant negative surface charge was present in the vestibules to the pore and that the pore could accommodate two cations simultaneously, which repelled each other strongly. The pore structure of the rca channel appeared to differ from that of L-type calcium channels from animal cell membranes since its ion binding sites had a lower affinity for divalent cations. The model adequately accounted for the diverse permeation phenomena observed for the rca channel. It described the apparent submillimolar Km for the relationship between unitary conductance and Ca2+ activity, the differences in selectivity sequences obtained from measurements of conductance and permeability ratios, the changes in relative cation permeabilities with solution ionic composition, and the complex effects of Ca2+ on K+ and Na+ currents through the channel. Having established the adequacy of the model, it was used to predict the unitary currents that would be observed under the ionic conditions employed in patch-clamp experiments and to demonstrate the high selectivity of the rca channel for Ca2+ influx under physiological conditions.

The depolarization-activated, high-conductance "maxi" cation channel in the plasma membrane of rye (Secale cereale L.) roots is permeable to a wide variety of monovalent and divalent cations. The permeation of K+, Na+, Ca2+ and Ba2+ through the pore could be simulated using a model composed of three energy barriers and two ion binding sites (a 3B2S model), which assumed single-file permeation and the possibility of double cation occupancy. The model had an asymmetrical free energy profile. Differences in permeation between cations were attributed primarily to differences in their free energy profiles in the regions of the pore adjacent to the extracellular solution. In particular, the height of the central free energy peak differed between cations, and cations differed in their affinities for ion binding sites. Significant ion repulsion occurred within the pore, and the mouths of the pore had considerable surface charge. The model adequately described the diverse current vs. voltage (I/V) relationships obtained over a wide variety of experimental conditions. It described the phenomena of non-Michaelian unitary conductance vs. activity relationships for K+, Na+ and Ca2+, differences in selectivity sequences obtained from measurements of conductance and permeability ratios, changes in relative cation permeabilities with solution composition, and the complex effects of Ba2+ and Ca2+ on K+ currents through the channel. The model enabled the prediction of unitary currents and ion fluxes through the maxi cation channel under physiological conditions. It could be used, in combination with data on the kinetics of the channel, as input to electrocoupling models allowing the relationships between membrane voltage, Ca2+ influx and Ca2+ signaling to be studied theoretically.

A method is described for estimating rapid rate constants from the distributions of current amplitude observed in single-channel electrical recordings. It has the advantages over previous, similar approaches that it can accommodate both multistate kinetic models and adjustable filtering of the data using an 8-pole Bessel filter. The method is conceptually straightforward: the observed distributions of current amplitude are compared with theoretical distributions derived by combining several simplifying assumptions about the underlying stochastic process with a model of the filter and electrical noise. Parameters are estimated by approximate maximum likelihood. The method was used successfully to estimate rate constants for both a simple two-state kinetic model (the transitions between open and closed states during the rapid gating of an outward-rectifying K(+)-selective channel in the plasma membrane of Acetabularia) and a complex multistate kinetic model (the blockade of the maxi cation channel in the plasma membrane of rye roots by verapamil). For the two-state model, parameters were estimated well, provided that they were not too fast or too slow in relation to the sampling rate. In the three-state model the precision of estimates depended in a complex way on the values of all rate parameters in the model.