Adult Zebrafish Electrophysiology: A Method to Measure the Extracellular Electrical Activity of Neurons In Vivo
Trascrizione
– Neurons propagate electrical impulses via action potentials. When the neuron is di resting state, there is a slower outflow of potassium ions than the inflow of sodium ions, which leads a the change di membrane potential. This causes the sodium ion channels a open and allow the inflow of sodium ions, generating an electrical impulse.
The method of studying such activities di a cell is called electrophysiology. Begin the procedure by inserting a secondary electrode tip into the nostril of an adult zebrafish. Next, insert the primary electrodes superficially, within the exposed optic tectum, a measure extracellular neural activity.
The primary electrode measures the extracellular membrane potential of all cells surrounding it. The signal represents the combined membrane potential of all these cells. The electrode detects the signal and transmits it a an amplifier– an oscilloscope– which presents a visual display of the membrane potential, and a computer.
Observe the readings and analyze the electrical difference between the primary and the secondary electrodes. The electrophysiological measurement is a comparison. In the example protocol, we will measure the effect of pharmacological compounds on the neural activity di an adult zebrafish.
– For electrophysiology recording, shut off the intubation stopcock and quickly disconnect the Leur lock connecting the intubation base a the tricaine drip. Move the intact intubation setup a the electrophysiology microscope, and connect it a the perfusion system. Turn on the habitat water and perfuse at a rate of 1 milliliter per minute for about one hour, di order a wash out the tricaine.
Using a micro-manipulator, position the secondary electrode so that the electrode tip can be inserted into the animal's nostril, or into the dip under the upper jaw. Subsequently, insert the primary electrode needle into the craniotomy opening. Then, insert the needle into the tissue, such that the tip is positioned fairly superficial within the optic tectum.
Deeper positions of the electrode may result di a small electrical signal. For data collection, gap-free mode at 5,000 hertz sampling rate, together with the lowpass filter of 0.1 kilohertz and a highpass filter of 1 hertz, are used. Begin the recording of the electrophysiological activity when the habitat water has perfused for a minimum of 45 minutes. Record a baseline of native activity for at least 15 minutes prior a the addition of experimental drugs.
