Modulation of slow inactivation in class A Ca2+ channels by β-subunits

Abstract

1. β-subunit modulation of slow inactivation of class A calcium (Ca2+) channels was studied with two-microlectrode voltage clamp after expression of the α(1A)- (BI-2) together with β(1a)-, β(2a)-, β3- or β4-subunits in Xenopus oocytes. 2. On- and off-rates of slow inactivation were estimated from the kinetics of recovery from slow inactivation. Ca2+ channels with an α(1A)/β-subunit composition inducing the slower rate of fast inactivation displayed the faster rate of slow inactivation. The corresponding order of slow inactivation time constants (τ(onset)) was: α(1A)/β(2a), 33±3s; α(1A)/β4, 42±4 s; α(1A)/β(1a), 59±4 s; α(1A)/β3, 67±5 s (n≥7). 3. Recovery of class A Ca2+ channels from slow inactivation was voltage dependent and accelerated at hyperpolarized voltages. At a given holding potential recovery kinetics were not significantly modulated by different β-subunits. 4. Two mutations in segment IIIS6 (IF1612/1613AA) slowed fast inactivation and accelerated the onset of slow inactivation in the resulting mutant (α(A/IF-AA)/β3) in a similar manner as coexpression of the β(2a)-subunit. Recovery from slow inactivation was slightly slowed in the double mutant. 5. Our data suggest that class A Ca2+ channels enter the 'slow inactivated' state more willingly from the open than from the 'fast inactivated' state. The rate of slow inactivation is, therefore, indirectly modulated by different β-subunits. 6. Fast and slow inactivation in class A Ca2+ channels appears to represent structurally independent conformational changes. Fast inactivation is not a prerequisite for slow inactivation.