Pulsing frequency (0.01-10):
Aim: Open hERG channels using different stimulation frequencies and observe frequency-dependent channel block.
In our experiments, we use the voltage-clamp technique, which allows us to apply and maintain a fixed membrane potential ("clamped") during the recording. Typically, square voltage pulses are applied, and the potassium current flowing through hERG channels is measured in response to changes in voltage. This approach enables the investigation of activation, deactivation, and recovery kinetics characteristic of the hERG current.
Choose frequency and apply voltage protocol in the presence of drug (ie 10uM Amiodarone)
Hz
Aim: Apply different drug concentrations and observe hERG channel block.
To evaluate how different drug concentrations affect the hERG channel, we construct dose-response curves by measuring the extent of current inhibition at each concentration. Cells expressing hERG channels are exposed to increasing doses of a test compound, typically during a defined voltage protocol that elicits stable outward potassium currents. The percent inhibition is plotted against the logarithm of the drug concentration. By fitting the data with a standard Hill equation, we can determine key pharmacological parameters such as the half-maximal inhibitory concentration (IC50) and the Hill coefficient, which provide insight into the potency and cooperativity of the drug-channel interaction.
Apply different drug's concentrations
Drug concentration (0.01-100):
hERG Channel Block: Understanding How Drugs Affect Cardiac Electrical Signals
hERG channels are crucial potassium channels in the heart that help repolarize the cardiac action potential. They allow potassium ions to exit the cell, resetting the electrical state after each heartbeat. Many drugs — including antiarrhythmics and unintended blockers — can inhibit hERG channels, potentially leading to dangerous heart rhythm disturbances such as long QT syndrome.
Unlike most voltage-gated channels that open quickly and inactivate slowly, hERG channels behave uniquely: they open slowly and inactivate rapidly during depolarization. During the repolarization phase, they recover quickly from inactivation and briefly pass a large outward potassium current. This current, known as the tail current, accelerates repolarization and helps stabilize the heart's rhythm.
To investigate hERG current, researchers use a voltage-clamp protocol that mimics a cardiac action potential. A typical protocol includes a depolarizing step to +20 mV (plateau phase), followed by a step to -40 mV to elicit maximal tail current, and then a return to -80 mV, the resting potential.
1. State-Dependent Block (Affinity for Open or Inactivated States)
Like other voltage-gated channels, hERG channels transition through multiple states — open, inactivated, and closed — in response to changes in voltage. Some drugs show a preference for binding to the channel when it is open or inactivated, a phenomenon known as state-dependent block. These drugs are more likely to bind during periods of high electrical activity, which can lead to enhanced block at faster heart rates.
2. Frequency-Dependent Block (Use Dependence)
The more frequently hERG channels cycle through open and inactivated states — such as during rapid heart rhythms — the more opportunities a drug has to bind. This leads to frequency-dependent block, where repetitive stimulation increases inhibition. This property is key when assessing a drug's proarrhythmic risk, particularly during exercise or stress when heart rate increases.
In summary, hERG channel blockers must be evaluated not just by whether they block the channel, but how, when, and under what conditions they do so. Understanding hERG channel dynamics and gating behavior helps predict a drug's cardiac safety and proarrhythmic potential.