Generating Local Cornu Ammonis 1 (CA1) Gamma Oscillations Using Tetanic Stimulations in a Hippocampal Brain Slice

Secure a brain slice in a recording chamber with a continuous perfusion of aCSF containing high magnesium that blocks N-methyl-D-aspartate or NMDA glutamate receptors.

Position the stimulating and recording electrodes in the hippocampal Cornu ammonis 1 or CA1 stratum radiatum.

Stimulate to obtain an electrical signal, confirming intact neuronal connections.

Reposition the stimulating electrode to the stratum oriens and the recording electrode to the stratum pyramidale, then insert them deeper.

Apply tetanic stimulations, high-frequency electrical pulses to induce excitatory neurotransmitter glutamate release from Schaffer collateral axons.

Glutamate binds to glutamate receptors on CA1 neurons, facilitating sodium influx and membrane depolarization.

This unblocks NMDA receptors, allowing sodium and calcium influx and enhancing neuron excitation.

The excited interneurons in the stratum oriens release the inhibitory neurotransmitter GABA, interacting with CA1 neurons.

This balance of excitatory and inhibitory inputs leads to synchronized controlled excitation of the CA1 neuronal network, generating rhythmic brain waves called gamma oscillations, crucial for attention and memory.

To mount the slice in the recording chamber, place a brain slice into a submerged recording chamber perfused with ACSF flowing at 1 to 2 milliliters per minute and heated to 32 degrees Celsius. The ACSF used for recordings differs from that in the holding chamber, as the magnesium concentration is increased from 2 millimolar to 4 millimolar.

Secure the slice by placing a semi-circular stainless steel weight on it with its strands parallel to hippocampal CA1. After that, increase the speed of perfusion to 8 to 10 milliliters per minute at 32 degrees Celsius.

Using a dissecting microscope, move the stimulating electrode to the middle of the stratum oriens. Then, move the recording electrode to the pyramidal layer as close to the stimulating electrode as possible. Subsequently, lower both the stimulating and recording electrodes 50 to 100 micrometers into the slice in order to generate the field EPSP of 1 millivolt in response to the 120 to 150 microamp test pulse.

To generate gamma oscillations, stimulate the tissue with a train of 20 times 0.1-millisecond pulses delivered at 200 hertz. This tetanic stimulus can yield reproducible responses when delivered every 5 minutes.