Optical signals from neurons with internally applied voltage-sensitive dyes

Area of Science:

• Neuroscience
• Biophysics
• Optical Imaging

Background:

• Monitoring neuronal electrical activity, such as action potentials and subthreshold potentials, in individual neurons is crucial for understanding neural function.
• Traditional methods often lack the spatial or temporal resolution to accurately track signal propagation in fine neuronal processes.
• Previous optical methods using voltage-sensitive dyes showed limitations in sensitivity, particularly for intracellular monitoring in distal neuronal structures.

Purpose of the Study:

• To develop and validate a highly sensitive optical method for monitoring neuronal electrical activity in real-time.
• To investigate the generation and spread of action potentials and subthreshold potentials in individual snail neurons.
• To determine the site of action potential initiation and calculate propagation velocities in axonal segments.

Main Methods:

• Experiments were conducted on neurons from the snail Helix aspersa ganglia.
• Neurons were stained using intracellular pressure injection of voltage-sensitive dyes.
• A fast, multiple-site optical monitoring system with a silicon photodiode array was employed, focusing on fluorescence measurements with the novel styryl dye JPW1114.

Main Results:

• Absorption-based dye testing proved insufficiently sensitive for distal neuronal processes.
• The newly synthesized dye JPW1114, used in fluorescence mode, provided a ~50-fold improvement in signal-to-noise ratio compared to previous methods.
• Simultaneous recordings from cell bodies and axonal branches allowed determination of action potential initiation sites and calculation of propagation velocities (0.07–0.53 m/sec).
• The method successfully recorded 10 mV hyperpolarizing electrotonic responses and their decline with distance.

Conclusions:

• The novel voltage-sensitive dye JPW1114 and fluorescence-based optical monitoring system offer unprecedented sensitivity for studying neuronal electrical activity.
• This technique enables detailed analysis of action potential initiation and propagation dynamics in neuronal processes.
• Future modest improvements in sensitivity could facilitate systematic studies of synaptic potential spread and summation in single neurons.