A Procedure for Implanting Organized Arrays of Microwires for Single-unit Recordings in Awake, Behaving Animals

Summary

Implanting organized arrays of microwires for use in single-unit electrophysiological recordings presents a number of technical challenges. Methods for performing this technique and the equipment necessary are described. Also, the beneficial use of organized microwire arrays to record from distinct neural subregions with high spatial selectivity is discussed.

Abstract

In vivo electrophysiological recordings in the awake, behaving animal provide a powerful method for understanding neural signaling at the single-cell level. The technique allows experimenters to examine temporally and regionally specific firing patterns in order to correlate recorded action potentials with ongoing behavior. Moreover, single-unit recordings can be combined with a plethora of other techniques in order to produce comprehensive explanations of neural function. In this article, we describe the anesthesia and preparation for microwire implantation. Subsequently, we enumerate the necessary equipment and surgical steps to accurately insert a microwire array into a target structure. Lastly, we briefly describe the equipment used to record from each individual electrode in the array. The fixed microwire arrays described are well-suited for chronic implantation and allow for longitudinal recordings of neural data in almost any behavioral preparation. We discuss tracing electrode tracks to triangulate microwire positions as well as ways to combine microwire implantation with immunohistochemical techniques in order to increase the anatomical specificity of recorded results.

Introduction

Electrophysiological recordings allow scientists to examine the electrical properties of biological cells. In the central nervous system, where electrical impulses serve as a signaling mechanism, these recordings are of particular importance for understanding neural function^1-2. During single-unit recordings in behaving animals, a microelectrode that has been inserted into the brain is able to record changes in a neuron's generation of action potentials over time.

While many techniques allow one to record brain activity, single-unit electrophysiology is one of the most precise methods by allowing resolution at the single neur....

Protocol

The utmost care must be taken to maintain aseptic conditions (as described in the Guide for the Care and Use of Laboratory Animals⁹) while preparing for and conducting the following procedure. The following protocol is in compliance with the Guide for the Care and Use of Laboratory Animals and has been approved by the Institutional Animal Care and Use Committee, Rutgers University. It is estimated that the subsequent procedures will require 3-6 hr to complete.

Implanting the Microwire Array:

1. Place animals under anesthesia using 50 mg/kg sodium pentobarbital (i.p) and administer 10 mg/kg of atropine methyl nitrate (IP; ....

Results

A list of Equipment used by this laboratory for recording electrophysiological signals can be found in Table 3. Following recovery from surgery, single-units are recorded by plugging a unity-gain headstage into the implanted connector. This headstage is connected via a cable to a commutator, which is capable of free rotation without breaks in the electrophysiological recording through the use of electrical slip rings. The commutator allows subjects to freely move while recording during behavior, which is.......

Discussion

Extracellular recordings represent a powerful experimental technique that can be incorporated into nearly any experimental preparation in neuroscience. Wires that have been implanted in organized arrays can be tracked as their shafts pass through the brain and into their target region (Figure 5A). When a small, post-experimental lesion is created at the noninsulated microwire tip to create a small iron deposit from the stainless steel wire, one can precisely mark the location of the uninsulated microwire.......

Materials

List of materials used in this article

Name	Company	Catalog Number	Comments
Table 1. List of Surgical Materials.
Gauze	Fisher (MooreBrand)	19-898-144
Cotton Swabs	Fisher (Puritan)	S304659
Nembutal (Pentobarbital)	Sigma Aldrich	P3761
Atropine Methyl Nitrate	Sigma Aldrich	A0382
Baytril (Enrofloxacin)	Butler Shein (Bayer)	1040007
Ketamine Hydrochloride	Butler Shein	SKU# 023061
Betadine (Povidone-Iodine)	Fisher (Perdue)	19-066452
Stereotax	Kopf	Model 900
Cauterizing Tool	Stoelting	59017
Dissecting Microscope	Nikon	SMZ445
Dental Drill	Buffalo	37800
Bacteriostatic Saline	Bulter Schein	8973
Jewlers Screws	Stoelting	51457
Microwire Array	Microprobes	Custom (Flexible)
Ground Wire	Omnetics	Custom Plug
Dental Acrylic	Fisher (BAS)	50-854-402
Absorbable Sutures	Fisher (Ethicon)	NC0258473
Puralube (Opthalamic Ointment/Lubricant)	Fisher (Henry Schein)	008897
Table 2. List of Surgical Instruments.
2x Microforceps	George Tiemann Co.	#160-57	Multi-use (e.g. clearing debris in skull window)
2x Forceps	George Tiemann Co.	#160-93	Multi-use (e.g. tying sutures)
6x Hemostats	George Tiemann Co.	#105-1125	Clamp and open incision
1x Small scissors	George Tiemann Co.	#105-411	Cut sutures after tying
1x Tissue forceps	George Tiemann Co.	#105-222	Holding tissue while suturing
1x Needle holder	George Tiemann Co.	#105-1259	Holding suture needle
1x Scalpel holder (with #11 blade)	George Tiemann Co.	#105-80 (w/ #105-71 blade)	Making skull incision
1x #22 Scalpel blade	George Tiemann Co.	#160-381	Shaving scalp
1x Surgical Spatula	George Tiemann Co.	#160-718	Scraping skull to clear tissue on skull
Machine/Jewelers Screws	Various	N/A	0/80 x 1/8”
Table 3. List of Equipment for Recording Electrophysiological Signals.
Microwire Array & Connector	Micro Probe, Inc. (Gaithersburg, MD)	N/A (Part No. based on array characteristics)	Cranially implanted in target recording region. Arrays are customized based on desired wire spacing, length, etc.
Unity-Gain Harness/Headstage	M.B. Turnkey Designs (Hillsborough, NJ)	Proj 1200	Initial amplification of neural signal; allows for propagation of small neural signals.
Commutator (and Optional Fluid Swivel)	Plastics One, Inc. (Roanoke, VA)	SL18C	Allows animals to freely rotate while propagating electrical signal to preamp
Pre-amplifier	M.B. Turnkey Designs (Hillsborough, NJ)	Proj 1198	Differentially amplifies neural signals against a reference electrode.
Filter and Amplifier	M.B. Turnkey Designs (Hillsborough, NJ)	Proj 1199	Band-pass filters and further amplifies the differentially amplified signal.
Acquisition Computer	EnGen (Phoenix, AZ)	N/A (Custom Build)	Runs software and hardware for behavioral and neural data acquisition.
A/D Card	Data Translation (Marlboro, MA)	DT-3010	Digitizes neural signals for computer sampling.
Digital I/O Card	Measurement Computing (Norton, MA)	PCI CTR-05	Acquires behavioral inputs and outputs