Ion permeation through single channels activated by acetylcholine in denervated toad sartorius skeletal muscle fibers: Effects of alkali cations

Abstract

The gigaohm seal technique was used to study ion permeation through acetylcholine-activated channels in cell-attached patches of the extrajunctional membrane of chronically denervated, enzyme-treated cells from the sartorius muscle of the toad Bufo marinus. The most frequently occurring channel type (greater than 95% of channel openings), provisionally classified as 'extrajunctional,' had a chord conductance of approximately 25 pS under normal conditions (-70 mV, 11 degrees C, Normal Toad Ringer's). The less frequently observed channel type (less than 5% of channel openings), classified as a 'junctional' type, had a conductance of 35 pS under the same conditions, and a similar null potential. In many patches, a small percentage (usually less than 2%) of openings of the extrajunctional channel displayed a lower conductance state. The shape of the I-V curves obtained for the extrajunctional channel depended on the predominant extracellular cation. For Cs and K, the I-V curves were essentially linear over the voltage range +50 to -150 mV across the patch, suggesting that the potential independent component of the energy profile within the channel was symmetrical. For Li, the I-V curve was very nonlinear, displaying a significant sublinearity at hyperpolarized potentials. Both an electrodiffusion and a symmetrical uniform four-barrier, three-site rate-theory model provided reasonable fits to the data, whereas symmetrical two-barrier, single-site rate-theory models did not. For the alkali cations examined, the relative permeability sequence was PCs greater than PK greater than PNa greater than PLi--a "proportional" selectivity sequence. This was different from the single channel conductance sequence which was found to be gamma K greater than gamma Cs greater than gamma Na greater than gamma Li implying that ions do not move independently through the channel. The relative binding constant sequence for the channel sites was found to be a "polarizability" sequence, i.e., KLi greater than KCs greater than KNa greater than KK. There was an inverse relationship between the relative binding constant and the relative mobility for the cations examined. Under conditions when the single-channel conductance was relatively high, the conductance at depolarized potentials was lower than that predicted by both electrodiffusion and rate theory models, suggesting that there was a rate-limiting access step for ions, from the intracellular compartment into the channel.

Threonine in the selectivity filter of the acetylcholine receptor channel

“…In the AChR channel studied here, the best conducted ion is Rb+ and not K+ as in Torpedo AChR. The conductance sequence Rb > K > Cs > Na differs slightly from that of other AChR channels previously studied in frog muscle (Quartararo et al, 1987) or in Torpedo electric organ (Konno et al, 1991). The aT264V mutation produces a decrease in the conductance for large cations changing the conductance sequence to K > Rb > Cs > Na.…”

“…The measured pipette potential was substracted from this potential. The zero current potential was corrected by the liquid junction potential between the two solutions, calculated using the ion mobilities (Hille, 1984;Quartararo et al, 1987), to obtain the reversal potential. The temperature was 18'C.…”

“…The receptor channel is permeable to monovalent as well as divalent cations (Adams et al, 1980). In most preparations, permeability for monovalent cations follows the water mobility (Hille, 1984;Quartararo et al, 1987;Konno et al, 1991) as for the divalent cations the opposite has been found. The conductance sequence measured by single-channel recording is different from the permeability sequence (Quartararo et al, 1987, Konno et al, 1991.…”

M2 Pore Mutations Convert the Glycine Receptor Channel from Being Anion- to Cation-Selective

“…Thus, this study demonstrates that there is conservation of the molecular determinants of selectivity between the cationic and anionic LGICs. In addition to this fundamental change in ion selectivity, another key property conferred upon the STM GlyR was the monovalent permeability sequence (Cs ϩ Ͼ K ϩ Ͼ Na ϩ Ͼ Li ϩ ), which is identical to that of the native nAChR (Adams et al, 1980;Quartararo et al, 1987;Konno et al, 1991;. This suggests that the profile of the STM GlyR pore constriction region resembles that of the native nAChR.…”