| dc.description.abstract | Neuronal excitability is mediated by ion-selective transmembrane ion channels, through which ions are transported in response to changes in membrane potential. In particular, the rising phase of the action potential is caused by an influx of sodium ions through voltage-sensitive sodium channels. These channels are large glycoproteins that consist of a single major pore-forming polypeptide, the subunit of molecular weight 226 kDa, and 1 and 2 auxiliary subunits, each having a molecular weight of 31–33 kDa.
Sodium channels are found in almost all excitable cells of the nervous system, muscle, heart, and endocrine cells; they are instrumental in determining the threshold of action potential firing and play a vital role in its initiation and propagation. Excitability of a neuron can be modulated by regulating the effective number of sodium channels available for opening. Such modulation is possible due to fast and slow inactivation properties of the channel molecule. Once inactivated, the channels do not activate upon a subsequent depolarization. Thus, inactivating sodium channels are crucial in regulating the response of excitable cells to internal and external stimuli.
The modulation of the threshold of action potential firing is a powerful memory mechanism that depends solely on the intrinsic properties of a neuron, independent of its synaptic input. Loss of inactivation, or its slowing characterized by slower time constants of inactivation, is associated with neurological disorders such as epilepsy, hyperkalemic periodic paralysis, and cardiac arrhythmia, suggesting the importance of sodium channel inactivation kinetics in cellular functioning.
In the present work, the relation between previous channel activity and the availability of channels for subsequent activation was studied. We examined the effect of sustained depolarization, in both long (1–160 s) and short (10–1000 ms) duration ranges, on the detailed kinetics of activation, fast inactivation, and recovery from slow inactivation in rNav1.2a channel subunit expressed in Chinese Hamster Ovary (CHO) cells. The duration and amplitude of prolonged depolarization altered all steady-state and kinetic parameters in a pseudo-oscillatory fashion with time-variable period and amplitude, often superimposed on a linear trend. The pseudo-periodicity was confirmed using weighted wavelet (WWZ) analysis.
The half steady-state activation potential showed a reversible depolarizing shift of 5–10 mV, dependent on the duration of prior prolonged depolarization (1–160 s), while the half steady-state inactivation potential showed a hyperpolarizing shift of 43–55 mV. The time periods of slow pseudo-oscillation for most parameters, determined by WWZ analysis, lie close to 30 s, suggesting consistent oscillation in the channel that couples activation, fast, and slow inactivation.
A similar search for pseudo-periodic changes in the channel was made with short depolarizing prepulses of duration range 10–1000 ms. The steady-state activation and inactivation parameters, as well as the time constant of recovery from fast inactivation, depended on the shorter prepulse duration (100–1000 ms) and amplitude in a pseudo-oscillatory manner. However, in the 10–100 ms prepulse duration range, the time constant of recovery changed monotonically with prepulse duration, following a power relationship. Most parameters showed a time period of oscillation close to 225 ms.
Co-expression of the 1 subunit decreased the periods of oscillation (close to 22 s for +1 in steady-state activation parameters in the prolonged prepulse duration range), whereas a suppression of the oscillatory component in the steady-state inactivation parameters was observed in the shorter prepulse duration range.
This inherent pseudo-oscillatory mechanism within voltage-gated sodium channels may regulate neuronal excitability and account for epileptic discharges and subthreshold membrane potential oscillation. To verify this, history dependence of activation parameters was examined with paroxysmal depolarization shift (PDS)-type stimulus protocol rather than a sustained prepulse. Pseudo-oscillatory dependence of steady-state activation on the total time spent in the depolarized state was observed in this case as well.
When varying parameters of sodium channel activation and fast inactivation, subject to variable prepulse duration in both long and short duration ranges, were incorporated in a single neuron simulation, subthreshold oscillation and periodic burst-and-silence activity in action potential firing—similar to epileptic seizures—were observed. Thus, the history dependence mechanism of sodium channels offers a complex molecular memory mechanism intrinsic to the neuron, independent of synaptic activity.
Different signaling molecules that alter sodium channel kinetics, such as PKA, PKC, and G-proteins, may modulate this history dependence property. The effects of G-protein activation on the pseudo-oscillatory properties of sodium channels, in response to conditioning depolarization in both long and short duration ranges, were studied. While G-protein fine-tuned the period of pseudo-periodic oscillation of steady-state activation parameters in the longer duration range, it abolished the oscillation in steady-state inactivation parameters in the shorter duration range.
These findings suggest tight regulation of the history dependence and plasticity of sodium channel molecules by cellular signaling mechanisms. | |
