Home › Neuroscience › Optical Imaging of Neurons in the Crab Stomatogastric Ganglion with Voltage-sensitive Dyes
Neuroscience JoVE (Open Access) Citable · DOI

Optical Imaging of Neurons in the Crab Stomatogastric Ganglion with Voltage-sensitive Dyes

DOI: 10.3791/2567-v
What you'll learn
  • ✓ Prepare and load voltage-sensitive dyes into crab stomatogastric ganglion neurons
  • ✓ Perform high-resolution fluorescent imaging of neuronal membrane potential changes
  • ✓ Correlate optical voltage recordings with simultaneous extracellular electrical measurements
Protocol

Here we present the methodology for fast and high resolution fluorescent voltage-sensitive dye imaging of detailed activity of neurons in the crab stomatogastric ganglion.

Difficulty
advanced
Total time
~3–4 hours per preparation (dye loading and imaging)
Model organism
Crab (Cancer productus or similar decapod)

Steps

1
Prepare voltage-sensitive dye solution

Mix and dissolve voltage-sensitive dye to appropriate concentration for intracellular loading. Ensure dye is fully solubilized and ready for microinjection.

▶ 02:11
2
Dissect and isolate stomatogastric ganglion

Surgically expose and prepare the crab stomatogastric ganglion tissue. Set up extracellular recording electrodes to monitor neural activity during imaging.

▶ 03:18
3
Load voltage-sensitive dye intracellularly

Inject voltage-sensitive dye directly into individual neurons of the stomatogastric ganglion using microelectrodes. Allow time for dye equilibration within the cell.

▶ 03:58
4
Acquire high-resolution optical imaging data

Use fluorescence microscopy to record fast voltage-dependent changes in dye fluorescence from stained neurons with high spatial and temporal resolution.

▶ 06:35
5
Compare optical and electrical recordings

Analyze and correlate simultaneous optical voltage imaging data with extracellular electrode recordings to validate neuronal activity measurements.

▶ 08:03
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