optical voltage recordings from naked-mole rat iNeurons

Last Tuesday (12/23) was my last day at work this year, and I ran my final experiment. Since starting the new project, the first goal has been to achieve optical voltage recording from a neuron. By filling neurons with what is called a voltage-sensitive dye (VSD), I have been working toward this goal.(In parallel, voltage recording using a GEVI—genetically engineered voltage indicator—on human iNeurons is being developed and is progressing thanks to our master student, Isabel de Vicente and our collaborator, Eline van Hugte, but it is not ready yet.)

To record optical voltage signals using VSD, the dye must first diffuse into the neuron from a patch-clamp pipette. In addition, the dye must be transported to the neuronal membrane in order to emit voltage-sensitive signals. This means that the neuron has to survive a long recording period. But in the end, it seems we have succeeded.

I recorded from a neuron derived from an iPSC culture of a naked mole-rat (NMR), prepared by our collaborator Sharon Gloudemans from Marijn Kuijpers’ lab. The neuron showed a firing pattern at room temperature, as shown in the figure below (left). After 40 minutes, the membrane of the neuron (top right, shown with the patch pipette) displayed clear fluorescence (bottom right).

By withdrawing the first pipette and placing a second pipette for patch-clamp recording in the cell-attached mode, and by applying voltage steps, the membrane potential of the cell was modulated, as shown in the figure below (bottom). The recorded emitted light reflected the temporal changes in the membrane potential (top).

Later, I was even able to do the re-patching of another cell, perform a second patch-clamp recording in the whole-cell configuration, and record optical voltage signals (below, optical signal is weak probably due to bleaching after repetitive illuminations).

Preliminary—but, oh well, it’s a start.