This article details a method for juxtacellular recording and staining of neurons in awake rats. The technique allows for precise visualization of neural activity and mapping to brain anatomy.
Start with a restrained, awake rat in an experimental jig, with its head firmly fixed in place.
Open the previously implanted recording chamber and remove the silicone gel covering.
Clear away any regrown tissue to access the brain.
Add a drop of saline to maintain hydration.
Next, attach a pre-filled recording pipette containing Chicago blue dye to a micromanipulator.
Lower the pipette into the recording chamber until it reaches the target brain region.
The pipette makes contact with the neuron and records the signals from the neuron without penetrating it.
Listen to these neuronal signals as audible clicks, then deliver electrical pulses through the pipette.
These pulses inject the dye adjacent to the neuron, enabling precise staining of the target brain region.
These juxtacellular recording and staining techniques precisely visualize the recording site and allow accurate mapping of neural activity to brain anatomy.
Place the rat in the cloth sock, and transfer the cloth sock into a rigid tube on an appropriate experimental jig. Attach the head restraining plate on the rat to the corresponding piece on the jig. Next, place an 832 nut on the head restraining bolt that is implanted on the rat.
Subsequently, screw in a threaded stainless steel rod to the head bolt. After that, open the recording chamber, and remove the silicone gel. Clean the craniotomy using fine forceps if tissue has regrown in the craniotomy. Use isoflurane anesthesia during this process, if necessary.
Next, attach the pipette to the motorized micromanipulator. Move the pipette to the desired recording location in the anterior, posterior, and medial lateral axes. Then advance the pipette ventrally through the dura until it is approximately 500 micrometers dorsal to the intended recording location. Slowly advance the pipette while listening for spiking events on an audio monitor of the amplified voltage recorded. Once spiking events are identified, continue to move the pipette about 0 to 100 micrometers until the positive voltage deflections are greater than approximately 500 microvolts.
To label the recording site, set the current source to negative 4 microamps with two second pulses at 50% Duty cycle. And pass the current for four minutes to iontophoretic inject Chicago Sky Blue through the pipette.
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