Developing a Behavioral Box for Assessing Prepulse Inhibition and Neural Activity in Psychiatric Animal Models

This protocol outlines a straightforward method for evaluating animal models of schizophrenia designed for behavioral researchers without programming or electronics expertise. We created a cost-effective behavioral box to assess prepulse inhibition in rats affected by neurodevelopmental issues stemming from early life distress. Recent advancements in psychiatric animal models using the prepulse inhibition test focus on understanding sensory gating deficits seen in disorders like schizophrenia.

This includes genetic modifications, neuroimaging, and pharmacological interventions to investigate prepulse inhibition disruptions, ultimately improving diagnostics and therapies. We hope that our protocol will popularize the study of prepulse inhibition in animal models of schizophrenia, and labs that are studying molecular, behavioral, and physiological aspects of the disease can use it to contribute to the understanding of this complex psychiatric disorder. To begin, make an acrylic transparent box to accommodate a freely-behaving rat equipped with a wired headstage.

Use self-adhesive rubber pads to isolate the acrylic box from vibrations and secure the accelerometer below without contact with other parts. Using a soldering iron, solder a ribbon cable directly to the four pinholes on the accelerometer board. VCC, GND, SCL, and SDA.

Clean the center of the acrylic and the flat face board of the accelerometer. Then, connect the opposite end of the ribbon cable to the microcontroller by connecting VCC to 3.3 volts, GND to GND, SCL to SCL, and SDA to SDA. Next, construct a soundproof chamber with a frontal door using 80 centimeter edges of 15 millimeters thick medium-density fiberboard.

Position a speaker 20 centimeters above the acrylic box, ensuring that the headstage and cable have a full range of motion. Connect the audio out from the computer to the microcontroller board. Using a universal serial bus cable, connect the microcontroller board to the computer.

Then, open the integrated development environment software for the microcontroller board on the computer. After selecting the corresponding microcontroller board, COM port and baud rate, open the file sketch, compile, and upload the code to the microcontroller board. Click on the monitor's serial port to check for incoming data.

Then, close the development environment software. Now, position a digital sound level meter within the acrylic box where the animal will be placed. Play the acoustic stimuli and adjust the detection range on the sound level meter to calibrate the stimulus levels being sent by the computer.

Then, open the preferred open-source software for acquiring and saving data from the serial port. Configure the software settings, including the COM port, microcontroller board, and baud rate. Click Connect to read raw data or the chart view.

Then, click Save to start recording the experiments data. After implanting the electrode array in the rat's brain, carefully connect the headstage to the connector implant on the rat's skull. Then, power on the neuronal data acquisition system and confirm that it receives data from the acoustic stimuli through either the microcontroller or a TTL generator connected to the audio detector or microphone.

Turn on the open-source serial port recorder set up earlier. Now, use electrophysiology acquisition software to collect electrophysiological and accelerometer data. Expose the animal to white background noise for seven minutes, with random presentation of all types of pulses, repeating each pulse type 10 times.

At the end of the recording session, turn off the acquisition software. Then, carefully disconnect the headstage connector from the implant on the animal's skull. To perform electrophysiological and behavioral analysis, import the raw data containing brain activity, acceleration, and acoustic stimuli into a signal processing platform.

Extract EPICS centered on each acoustic stimulus. Then, normalize the means to enable comparison between animals. Set the response to a single pulse as the maximum response and normalize the reflex responses to other pulses relative to this value.

Finally, define the startle amplitude for pulse 5 as 100%and represent responses to other pulses as a function of the pulse 5 amplitude. Local field potential analysis indicated no significant power modulation in the prelimbic and infralimbic cortices post-stimulation, while areas like the ventral tegmental area, amygdala, nucleus accumbens, and hippocampus showed decreased power spectrum density in delta, theta, and alpha bands, with a notable beta modulation. Both commercial and DIY accelerometers demonstrated consistent amplitude and time-based data, with baseline readings and startle response amplitudes showing close alignment across devices.