| Assay Method Information | |
| | In Vitro Assay (Protocol 1) |
| Description: | Compounds of Formula (I), were diluted in eight concentrations in the extracellular solution composed of physiological saline, buffered with HEPES (mM): NaCl, 137; KCl, 4; CaCl2, 3.8; MgCl2, 1; HEPES, 10; Glucose, 10; pH 7.4. The extracellular solution is composed of (mM) CsCl, 50; CsF, 90; MgCl2, 5; EGTA, 5; HEPES, 10; pH 7.2. The duration of exposure of each compound with the cells expressing Nav 1.7 or Nav 1.8 was at least five minutes and the assays were performed at room temperature.The measurements of the sodium currents of Nav 1.8 and Nav 1.7 were obtained using the voltage protocols described below:Protocol 1: A holding voltage of −90 mV was established followed by a voltage phase of 200 ms to −120 mV, followed by a voltage step of 10 mV for 1 second for Nav 1.8 or 0 mV for Nav 1.7, followed by a step of −100 mV for 20 ms, followed by a step of 20 ms for 10 mV for Nav 1.8 or 0 mV for Nav 1.7 (TP1A) before returning to the holding voltage of −90 mV.Protocol 2: A holding voltage of −100 mV was established followed by an inactivation voltage step at −40 mV for 8 seconds, followed by a −100 mV step for ms, followed by a 20 ms step for 10 mV for Nav 1.8 or 0 mV for Nav 1.7 (TP1A) before returning to the holding voltage of −100 mV.Protocols were repeated at a frequency of 0.1 Hz and current amplitude was quantified over the TP1A phase log. The variation in the peak amplitude of the current was evaluated according to the Formula described below, after exposure of the cells expressing the channels, at each concentration of the different compounds:%Block=(1-(ITP1A,molecule/ITP1A,basal)×100%,where |TP1A, basal, and |TP1A, molecule represent the entry peaks of sodium currents into TP1A prior to exposure to the compound and in the presence of the compound, respectively. |
| Affinity data for this assay | |
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