Retigabine holds KV7 channels open and stabilizes the resting potential

The anticonvulsant Retigabine is a KV7 channel agonist used to treat hyperexcitability disorders in humans. Retigabine shifts the voltage dependence for activation of the heteromeric KV7.2/KV7.3 channel to more negative potentials, thus facilitating activation. Although the molecular mechanism underlying Retigabine\u27s action remains unknown, previous studies have identified the pore region of KV7 channels as the drug\u27s target. This suggested that the Retigabine-induced shift in voltage dependence likely derives from the stabilization of the pore domain in an open (conducting) conformation. Testing this idea, we show that the heteromeric KV7.2/KV7.3 channel has at least two open states, which we named O1 and O2, with O2 being more stable. The O1 state was reached after short membrane depolarizations, whereas O2 was reached after prolonged depolarization or during steady state at the typical neuronal resting potentials. We also found that activation and deactivation seem to follow distinct pathways, suggesting that the KV7.2/KV7.3 channel activity displays hysteresis. As for the action of Retigabine, we discovered that this agonist discriminates between open states, preferentially acting on the O2 state and further stabilizing it. Based on these findings, we proposed a novel mechanism for the therapeutic effect of Retigabine whereby this drug reduces excitability by enhancing the resting potential open state stability of KV7.2/KV7.3 channels. To address this hypothesis, we used a model for action potential (AP) in Xenopus laevis oocytes and found that the resting membrane potential became more negative as a function of Retigabine concentration, whereas the threshold potential for AP firing remained unaltered

Development of a model for excitability studies using Xenopus oocytes

Action potentials (AP) are basic functional units of electrical signaling in excitable cells. These electrical signals are involved in many biological processes, including muscle contraction, synaptic transmission and hormone release. In general, the plasma membrane is polarized, displaying a difference in electric potential (membrane potential) with a negative intracellular voltage with respect to the extracellular space. During an AP, the membrane potential is momentarily canceled (depolarized) or reverted (anti-polarized) by a inwardly-rectifying current typically mediated by sodium-selective voltage-gated channels (VGC); the membrane potential is returned back (repolarize) to its initial voltage (resting potential) by a outwardly-rectifying current mediated by potassium-selective VGC. The temporal and electrical characteristics of APs depend on which VGCs are present in the membrane. Understanding the role of VGCs in AP generation in their native cells constitutes a difficult task, commonly riddled with the use of pharmacological agents to isolate each specific conductance. Here, we have developed a model to study cellular excitability using Xenopus oocytes. Spontaneous and evoked APs were readily recorded from oocytes expressing Nav1.4, Drosophila Kv1.1 (Shaker), human Kv7.2 and Kv7.3. These APs were around 5-ms long. However, in the absence of Shaker, the AP lasted about 50 ms. These observations indicated that we were able to modify the temporal characteristic of APs by removing the fast-activating Shaker. To further validate this model, we used the Kv7.2-7.3 agonist diclofenac seeking to decrease excitability. The addition of diclofenac drove the resting potential to more negative voltages and raised the threshold for excitation, effectively decreasing excitability. These results constitute proof of concept showing that this type of models can be used as functional scaffolds for the evaluation of pharmacological agents and the assessment of the effect of mutations in VGCs on the generation of bioelectrical signals

Retigabine holds KV7 channels open and stabilizes the resting potential

The anticonvulsant Retigabine is a KV7 channel agonist used to treat hyperexcitability disorders in humans. Retigabine shifts the voltage dependence for activation of the heteromeric KV7.2/KV7.3 channel to more negative potentials, thus facilitating activation. Although the molecular mechanism underlying Retigabine\u27s action remains unknown, previous studies have identified the pore region of KV7 channels as the drug\u27s target. This suggested that the Retigabine-induced shift in voltage dependence likely derives from the stabilization of the pore domain in an open (conducting) conformation. Testing this idea, we show that the heteromeric KV7.2/KV7.3 channel has at least two open states, which we named O1 and O2, with O2 being more stable. The O1 state was reached after short membrane depolarizations, whereas O2 was reached after prolonged depolarization or during steady state at the typical neuronal resting potentials. We also found that activation and deactivation seem to follow distinct pathways, suggesting that the KV7.2/KV7.3 channel activity displays hysteresis. As for the action of Retigabine, we discovered that this agonist discriminates between open states, preferentially acting on the O2 state and further stabilizing it. Based on these findings, we proposed a novel mechanism for the therapeutic effect of Retigabine whereby this drug reduces excitability by enhancing the resting potential open state stability of KV7.2/KV7.3 channels. To address this hypothesis, we used a model for action potential (AP) in Xenopus laevis oocytes and found that the resting membrane potential became more negative as a function of Retigabine concentration, whereas the threshold potential for AP firing remained unaltered

Retigabine holds KV7 channels open and stabilizes the resting potential

The anticonvulsant Retigabine is a K(V)7 channel agonist used to treat hyperexcitability disorders in humans. Retigabine shifts the voltage dependence for activation of the heteromeric K(V)7.2/K(V)7.3 channel to more negative potentials, thus facilitating activation. Although the molecular mechanism underlying Retigabine’s action remains unknown, previous studies have identified the pore region of K(V)7 channels as the drug’s target. This suggested that the Retigabine-induced shift in voltage dependence likely derives from the stabilization of the pore domain in an open (conducting) conformation. Testing this idea, we show that the heteromeric K(V)7.2/K(V)7.3 channel has at least two open states, which we named O(1) and O(2), with O(2) being more stable. The O(1) state was reached after short membrane depolarizations, whereas O(2) was reached after prolonged depolarization or during steady state at the typical neuronal resting potentials. We also found that activation and deactivation seem to follow distinct pathways, suggesting that the K(V)7.2/K(V)7.3 channel activity displays hysteresis. As for the action of Retigabine, we discovered that this agonist discriminates between open states, preferentially acting on the O(2) state and further stabilizing it. Based on these findings, we proposed a novel mechanism for the therapeutic effect of Retigabine whereby this drug reduces excitability by enhancing the resting potential open state stability of K(V)7.2/K(V)7.3 channels. To address this hypothesis, we used a model for action potential (AP) in Xenopus laevis oocytes and found that the resting membrane potential became more negative as a function of Retigabine concentration, whereas the threshold potential for AP firing remained unaltered