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⚡ Rapid Kindling Protocol Automation (Arduino + RTC-Based)

This repository provides firmware and documentation for automating the Rapid Kindling (RK) protocol used in epilepsy research. Unlike versions relying on microcontroller timing, this system uses a DS3231 Real-Time Clock (RTC) to guarantee long-duration precision.


🧠 Overview

Electrical stimulation is applied to the basolateral amygdala (BLA) in rodent models using surgically implanted electrodes. The Arduino-based system initiates stimulus trains at predefined intervals, following protocols described in:

  • Álvarez-Ferradas et al., Neurobiology of Disease, 2015
  • Morales et al., Frontiers in Cellular Neuroscience, 2014

The system includes:

  • RTC-controlled stimulation timing
  • Manual start/cancel via push button
  • LCD display for time and status feedback
  • TTL signal output for external trigger control

🔧 Hardware Requirements

Component Description
Arduino Uno / Nano Microcontroller
DS3231 RTC Module Precision timekeeping
I2C LCD (16x2, addr 0x27) Real-time display
Push Button Starts/cancels protocol (connected to D2)
Relay Output Triggers external stimulator (connected to D3)
Bipolar Stimulus Isolator Safely delivers stimulation current
Square-Wave Pulse Generator Defines waveform (frequency, amplitude)

✅ Optional add-ons: buzzer, SD logger, multichannel expansion


⏱️ Protocol Timing

The Arduino gates stimulation trains at key intervals after button-press initiation. Each train lasts 10 seconds. The exact timing replicates RK paradigms:

Elapsed Time Event
00:20:00 Stimulus 1
01:00:00 Stimulus 2
01:40:00 Stimulus 3
02:20:00 Stimulus 4
03:00:00 Stimulus 5
03:40:00 Stimulus 6

The real-time clock ensures consistent delivery over long durations.


⚡ External Triggering & Bipolar Stimulus Isolation

This system acts as an external trigger controller, not a waveform generator. The relay output sends TTL-level pulses to gate an external square-wave stimulator, configured with:

  • Frequency: 50–60 Hz
  • Pulse Duration: ~1 ms
  • Train Duration: 10 seconds
  • Current Amplitude: Subthreshold, based on post-discharge threshold (AD) testing

Stimulation is delivered via a bipolar stimulus isolator, ensuring:

  • 🔒 Electrical safety and tissue protection
  • 🧪 Artifact minimization during recording
  • 📏 Reproducible current delivery across sessions

This modular design separates timing logic (Arduino + RTC) from waveform control (external generator), following established methodology.


🖥️ Software Features

  • Manual start/cancel protocol interface
  • Chronometer-style time display on LCD
  • Status messages for each stimulation event
  • Modular functions for relay control and timing conversions
  • Cancelable protocol at any time via button press

📐 Wiring Summary

Signal Arduino Pin Description
Push Button D2 Start/cancel protocol
Relay Output (TTL) D3 Gates external stimulator
DS3231 RTC SDA/SCL Timekeeping
LCD Display SDA/SCL Displays feedback

🧬 Scientific Context

This system facilitates studies of epileptogenesis and neural plasticity, particularly involving:

  • After-discharges (ADs)
  • Seizure threshold monitoring
  • Kindling progression in limbic regions

Compatible with:

  • Optogenetic and chemogenetic stimulation
  • Electrophysiological recording systems
  • EEG and behavioral monitoring setups

✅ Ethical Compliance

All procedures follow ethical standards approved by the Bioethics Committee of the University of Valparaíso, including:

  • Minimizing discomfort and stress in animal models
  • Scientific justification for stimulation paradigms
  • Trained personnel for animal handling
  • International compliance for animal research

🧠 Extensions and Improvements

  • Interrupt-driven timing for enhanced stability
  • SD card logging of timestamps and stimulation events
  • Multi-channel expansion for bilateral stimulation
  • Integration with ESP32 or Teensy for wireless control
  • Cloud syncing or serial logging options

📄 License

Released under the MIT License. Free to use, modify, and redistribute for scientific or educational purposes.


🤝 Contributions

You're welcome to:

  • Improve timing logic or modularity
  • Integrate new hardware or data logging
  • Share experimental datasets or outcomes
  • Translate documentation and expand use cases

📎 References

  • Álvarez-Ferradas et al., Neurobiology of Disease, 2015
  • Morales et al., Front. Cell. Neurosci., 2014
  • Martorell et al., Neurobiology of Disease, 2020

👨‍🔬 Developed by: Felipe Guiffa Gómez
📧 Email: felipe.guiffa@postgrado.uv.cl
🌍 Valparaíso, Chile

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Automated stimulation sequence for rapid electrical kindling in rodent models using Arduino and LCD interface.

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