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  • 1
    ISSN: 1432-2013
    Keywords: K+ channels ; Ion permeation ; Chimeric K+ pore ; Single-site mutation ; Selectivity ; TEA blockade
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Abstract A conservative reversion at position 374 in a chimeric K+ pore, CHM, switched the preferred ionic conductance from K+ to Rb+. To understand how selectivity was switched, codons for 18 different amino acids were substituted at position 374 in each of two different K+ channels CHM and Kv2.1, the host channel for CHM. After injection of cRNA into Xenopus oocytes, less than half of the substituted mutants expressed functional channels. In both CHM and Kv2.1, channels with the substituted hydrophobic residues Val or Ile expressed Rb+-preferring pores while channels with the substituted polar residues Thr or Ser expressed K+-preferring pores. Val or Ile stabilized while Thr or Ser destabilized blockade by internal tetraethylammonium (TEA) confirming the importance of hydrophobic interactions for blockade. TEA blockade was dependent upon the charge carrier and was more effective in the presence of the ion having the larger conductance. The results are consistent with a model in which the side chains at position 374 form a filter for K+ and Rb+ ions and a site for blockade by internal TEA.
    Type of Medium: Electronic Resource
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  • 2
    Electronic Resource
    Electronic Resource
    Springer
    Cellular and molecular neurobiology 3 (1983), S. 381-395 
    ISSN: 1573-6830
    Keywords: Ca current ; voltage clamp ; ATP ; snail neuron
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary 1. Ca currents were examined with regard to their recovery from inactivation. The experiments were done on isolated nerve cell bodies ofHelix aspersa using a combined suction pippet, microelectrode method for voltage clamp, and internal perfusion. Ca currents were separated by suppressing K and Na currents. 2. The time course of recovery was determined by applying a test pulse at intervals ranging from 1 msec to 20 sec after prepulses varying from 20 to 3000 msec in duration. Each pair of pulses was preceded by a control pulse to ensure that the Ca currents had recovered before the next test pair was applied. Ba and Ca currents were compared and the effects of intracellular perfusion with EGTA, ATP, and vanadate were examined. 3. Ba currents recovered in two stages and this time course was well fit by a sum of two exponentials with amplitudes and time constants given byA 1 andτ 1 for the fast component andA 2 andτ 2 for the slow component. In Ba the time constants were unchanged when prepulse durations were prolonged from 70 to 700 msec, although the initial amplitudesA 1 andA 2, particularlyA 2, were increased. 4. Comparable influxes of Ca during the prepulse caused much more inactivation, but interestingly the recovery occurred at the same rate. The time course of Ca current recovery was also fit by a sum of two exponentials, the time constants of which were both smaller than the time constants of Ba current recovery. However, the time constants of Ca current recovery were increased markedly when prepulse durations were prolonged. Increasing the extracellular Ca concentration had a similar effect. 5. Increasing the Ba influx had no effect on the recovery time constants, and the Ba results are consistent with reversible inactivation gating of potential-dependent membrane Ca channels. The Ca results show that Ca influx enhances inactivation. Intracellular perfusion with EGTA resulted in less inactivation in the cast of Ca but it had no effect on Ba currents. Intracellular ATP increased the rate of recovery of Ca currents, and intracellular vanadate inhibited recovery. It is concluded that recovery of Ca channels depends upon both Ca influx and membrane potential and is modulated by agents which affect Ca metabolism.
    Type of Medium: Electronic Resource
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