ION PERMEATION, DIVALENT ION BLOCK, AND CHEMICAL MODIFICATION OF SINGLE SODIUM-CHANNELS - DESCRIPTION BY SINGLE-OCCUPANCY AND DOUBLE-OCCUPANCY RATE-THEORY MODELS

Calcium ions, applied internally, externally, or symmetrically, have b een used in conjunction with rate-theory modeling to explore the energ y profile of the ion-conducting pore of sodium channels. The block, by extracellular and/or intracellular calcium, of sodium ion conduction through single, batrachotoxin-activated sodium channels from rat brain was studied in planar lipid bilayers. Extracellular calcium caused a reduction of inward current that was enhanced by hyperpolarization and a weaker block of outward current. Intracellular calcium reduced both outward and inward sodium current, with the block being weakly depend ent on voltage and enhanced by depolarization. These results, together with the dependence of single-channel conductance on sodium concentra tion, and the effects of symmetrically applied calcium, were described using single- or double-occupancy, three-barrier, two-site (3B2S), or single-occupancy, 4B3S rate-theory models. There appear to be distinc t,outer and inner regions of-the channel, easily accessed by external or internal calcium respectively, separated by a rate-limiting barrier to calcium permeation. Most of the data could be well fit by each of the models. Reducing the ion interaction energies sufficiently to allo w a small but significant probability of two-ion occupancy in the 3B2S model yielded better overall fits than for either 3B2S or 4B3S models constrained to single occupancy. The outer ion-binding site of the mo del may represent a section of the pore in which sodium, calcium, and guanidinium toxins, such as saxitoxin or tetrodotoxin, compete. Under physiological conditions, with millimolar calcium externally, and high potassium internally, the model channels are occupied by calcium or p otassium much of the time, causing a significant reduction in single-c hannel conductance from the value measured with sodium as the only cat ion species present. Sodium conductance and degree of block by externa l calcium are reduced by modification of single channels' with the car boxyl reagent, trimethyloxonium (TMO) (Worley et al., 1986) Journal of General Physiology. 87:327-349). Elevations of only the outermost par ts of the energy profiles for sodium and calcium were sufficient to ac count for the reductions in conductance and in efficacy of calcium blo ck produced by TMO modification.