Jul 22, 2022

Public workspaceThe Acute Effects of Gastric Electrical Stimulation on Nucleus of the Solitary Tract Neural Activity Assessed with Electrophysiological Recording V.1

DOI
https://dx.doi.org/10.17504/protocols.io.4r3l2okzjv1y/v1
  • Jiayue Cao1,
  • Kun-Han Lu2,
  • Xiaokai Wang1,
  • Zhongming Liu1
  • 1University of Michigan, Ann Arbor;
  • 2Purdue University, West Lafayette
Marlena S Pela
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DOI: https://dx.doi.org/10.17504/protocols.io.4r3l2okzjv1y/v1
Protocol Citation: Jiayue Cao, Kun-Han Lu, Xiaokai Wang, Zhongming Liu 2022. The Acute Effects of Gastric Electrical Stimulation on Nucleus of the Solitary Tract Neural Activity Assessed with Electrophysiological Recording. protocols.io https://dx.doi.org/10.17504/protocols.io.4r3l2okzjv1y/v1
License: This is an open access protocol distributed under the terms of the Creative Commons Attribution License,  which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited
Protocol status: Working
We use this protocol and it’s working
Created: July 21, 2022
Last Modified: July 22, 2022
Protocol Integer ID: 67243
Keywords: gastric electrical stimulation, effects of gastric electrical stimulation, acute effects of gastric electrical stimulation, solitary tract neural activity, nucleus of the solitary tract neural activity, neural activity in the nucleus, electrophysiological recording brief description, neural activity, electrophysiological recording brief description this protocol, nts neuron, solitary tract, polarities of the stimulus, current direction of the stimulus, stimulation, different neuron, stimulus parameter, different groups of neuron, neuron, pulse amplitude, stimulus parameters in term, bipolar electrode, stimulation trail, recording electrode, current through the bipolar electrode, pulse parameter, pulse polarity, amplitude, stimulus, nucleus, distal coil
Abstract
Brief description
This protocol describes the methods used to evaluate the effects of gastric electrical stimulation (GES) on neural activity in the nucleus of solitary tract (NTS). Briefly, the multi-unit activity (MUA) and local field potential are accessed through multi-channel electrophysiological recording. The stimulation is applied during the imaging acquisition. The configuration of VNS was varied in terms of pulse parameters (amplitude, width) and polarity (proximal or distal coil of a bipolar electrode connected as the cathode).

Experiment design
The purpose of this study is to study how NTS neurons respond to GES with different parameter settings. To do so, we varied the stimulus parameters in terms of pulse amplitude (PA: 0.2, 0.4, 0.6, 0.8, 1, 2, 3, 4, 5, 6mA), width (PW: 0.1, 0.2, 0.3, 0.4, 0.5ms), and the pulse polarity. The pulse polarity can also be viewed as the current direction of the stimulus. For example, when delivering current through the bipolar electrode, the anode and cathode setting has two options which can result in two polarities of the stimulus. We altered the amplitude, pulse width, and pulse polarity. A total of 100 different parameter settings were tested for a single trail. The recording electrode was moved around to record from different groups of neurons in a single animal. The stimulation trail can be repeated multiple times on each animal but for different neurons.
Materials
Recording device info:
  • 64 CHANNEL EPHYS SYSTEM (Tucker Davis Technologies, Alachua, USA)
  • PZ5 NEURODIGITIZER AMPLIFIER
  • RZ2 BIOAMP PROCESSOR
  • DAQ (National Instruments, Austin, USA).

Recording electrode info:
  • 32 silicon penetrating probe (NeuroNexus, Ann Arbor, USA)

Stimulation electrode specification:
  • Two-polar patch electrode from MicroProbe.
  • Contact is made of Platinum with the size of 1mm by 2 mm.
  • The distance between two contacts is 2 mm
  • Stainless steel leads 100 mm long.
Troubleshooting
Anesthesia Procedure
Anesthetize the animal with 5% Isoflurane mixed with 100% oxygen at a flow rate of 500-1000 mL/min for 5 minutes (or till the animal loses consciousness).
After the animal loses consciousness, deliver a subcutaneous bolus injection of 0.075 mg/kg dexmedetomidine solution (0.05 mg/mL, Zoetis, NJ, USA).
After five minutes, continuously deliver 0.1-0.5% isoflurane through a nose cone.
Fifteen minutes after the bolus, deliver a continuous subcutaneous infusion of the dexmedetomidine solution at 0.015 mg/kg/hour through an infusion line by a syringe pump.
Adjust the dose of isoflurane and dexmedetomidine to keep the heart rate between 250 beats/min and 350 beats/min, and respiratory rate between 30 times/min and 60 times/min (Lu, et al., 2012, https://doi.org/10.1073/pnas.1200506109).
Surgical preparation

Note
The animal first underwent surgery to implant the patch electrode on the stomach wall.

Following a toe-pinch test, made a ∼3 cm incision starting at 1 cm caudal to the xiphoid and moving 3 cm caudally.
After separating and retracting the skin and muscle layers, expose the ventral stomach such that the intersection of the greater curvature and the limiting ridge can be identified.
Suture a pair of electrodes (Microprobes, Gaithersburg, MD, USA) directly onto the forestomach wall.


Note
  • Pair of electrodes consisted of Pt/Ir foil adhered to a thin perylene substrate (a rectangular shape of 4 mm-by-2 mm sizes and 0.015 mm thickness)
  • This patch electrode was placed along the greater curvature at about 4 mm proximal to the limiting ridge.

Close the muscle and skin layers at the incision site with sutures.
Electrical Stimulation
After the electrode implantation, secure the animal in the stereotaxic frame (Stoelting Co, Wood Dale, IL, USA).
Made a midline incision through the scalp.
Localize the nucleus of solitary tract (NTS) at the connection between the occipital bone and spinal cord.
Open a 3 × 3 mm window at the end of the occipital bone, and dissect the tissue above the spinal cord to expose the dorsal surface of the brainstem.
Identify the obex was, and place the recording electrode on the left side of obex at AP from 50 µm to -10 µm

Secure micro-screws above the olfactory bulb, and use them as reference and ground.
Connect the recording electrode to a broadband recording system (Tucker Davis Technologies, Alachua, FL, USA) that amplifies and digitizes (at 25 kHz) the voltage between the recording site and the reference site (i.e., a micro-screw secured over the olfactory bulb).


Note
The electrode localization was initially guided by the stereotaxic frame and was then finalized based on responses to electric stimuli applied to the stomach wall.

Recording setup:

1. Recording details
  • Sampling rate = 24 kHz
  • AC coupling
  • No band-pass filter

2. Data saving
  • Raw data
  • Local field potential (optional)
  • Stimulation triggers
Stimulation parameters:
  • Waveform: biphasic pulses with delay
  • Stimulation amplitude: 0.2 mA, 0.4 mA, 0.6 mA, 0.8 mA, 1.0 mA, 2 mA, 3 mA, 4 mA, 5 mA, 6 mA
  • Stimulation pulse width: 0.1 ms, 0.2 ms, 0.3 ms, 0.4 ms, 0.5 ms
  • Inter-pulse duration: 50 m
Note
The stimulation parameter was alternated through the experiments. Mat data was recorded to specify the details of the stimulation parameters.