Vala Assay: #NT4
Cell Type: Human dopaminergic neurons derived from normal hiPSCs or hiPSCs with the A53T alpha-synuclein mutation introduced with genome editing
Main Goal of Assay: Determine the effects of test compounds on action potential activity in human dopaminergic neurons derived from induced pluripotent stem cells. Neurons communicate with target cells through transient membrane electrical depolarization called action potentials. Dysregulation of action potential frequency can impede neuronal communication and cause the calcium overload, oxidative stress, inflammation, and cell death behind neurodegenerative diseases.
This assay features dopaminergic neurons, which are affected in Parkinson’s Disease, derived from normal hiPSCs or from isogenic hiPSCs with the A53T alpha-synuclein mutation introduced with genome editing. The A53T alpha-synuclein mutation causes early-onset Parkinson’s Disease. Testing compound effects on action potentials in both cell types can provide insight into Parkinson’s disease mechanisms and identify potential Parkinson’s therapeutics.
This assay measures the fluorescence of voltage indicator FluoVolt at 60 Hz for 10 seconds to capture neuronal action potentials on a cell-by-cell basis. Vala’s CyteSeer® image analysis program then reports a range of parameters (percent of active cells, event frequency, mean and maximum peak amplitudes, peak width, etc.) to provide a comprehensive picture of how each compound affects neuronal action potentials. This assay can be run concurrently with the neuronal calcium transient assay #NT3 to compare compound effects on calcium and voltage activity.
CyteSeer® Data Readout
- Fluorescence channel #1 (DAPI): Hoechst (cell number, viability)
- Fluorescence channel #2 (green channel): FluoVolt (neuronal action potential parameters)