Combining Imaging and Electrophysiology to Visualize and Record Spreading Depolarizations in Mice

Summary

This protocol demonstrates the combination of imaging and electrophysiology to reliably detect spreading depolarizations in adult mice following a mild traumatic brain injury.

Abstract

Spreading Depolarizations (SDs) are massive events in the brain that often go undetected due to their slow propagation through gray matter. Because SD detection can be elusive, it is optimally confirmed using multiple methods. This protocol describes methods for combining imaging and electrophysiology to detect SDs in a manner that most laboratories can reliably and easily adopt. SDs occur following traumatic brain injuries, stroke, subarachnoid hemorrhages, ischemia, and migraine aura. Historically, SDs have been recorded using DC amplifiers, which can resolve the slow extracellular shift and the depression in high-frequency activity. However, DC amplifiers are nearly impossible to use for chronic in vivo recordings. This protocol employs a common AC amplifier for in vivo electrophysiology recordings to confirm high-frequency depression, along with non-invasive imaging necessary to detect the propagating wave of SD. These methods can be reliably adopted and/or modified for most experimental approaches to confirm the presence or absence of SDs following brain injury.

Introduction

Spreading depolarizations (SDs) are associated with various neurological conditions, including traumatic brain injury (TBI), stroke, subarachnoid hemorrhages, ischemia, and migraine aura¹. SDs have been implicated in the progression of tissue damage in strokes and TBIs². The exact mechanism underlying tissue loss progression remains unclear^3,4. However, SDs occur when the metabolic demands of neurons are not met with sufficient oxygen and glucose, as the function of the ATP-dependent sodium-potassium pump becomes inadequate to maintain ion gradients. To understan....

Protocol

All procedures were performed in accordance with the Institute's Animal Care and Use Committee (IACUC) at the University of New Mexico. C57BL/6J mice were obtained from a commercial source. Equal numbers of male and female mice were used within the age range of 8-14 weeks. All mice weighed approximately 20 g. Details of the animals, reagents, and equipment used in this study are listed in the Table of Materials. Supplementary File 1 contains the list of abbreviations.

Discussion

Spreading depolarizations are crucial in various disease models, including traumatic brain injury (TBI), stroke, and migraine¹. Accurate detection of SDs is essential for understanding their role in acute behavioral deficits and/or the activation of the neuroimmune system following mild traumatic brain injuries (mTBIs)^23,24. To mitigate the risk of missing SDs when using a single method, a multi-method approach is necessary for effective v.......

Materials

Name	Company	Catalog Number	Comments
#00-90 x .118 Machine Screw	US Micro Screw	00.90-118-M-SS-P, Amazon
0.8 mm diameter drill bit	PCB Drill Bit	WYTP21, Amazon
16-A Headstage	Intan	C3335
2.1 mm Burrs micro drill bit	Fine Science Tools	19007-21
AC amplifier	Intan	RHD recording system
C57BL/6J mice	Jackson Laboratory	Jax#000664
CCD camera	Mightex	TCE-1304-U
Extension lens	Zeikos	ZE-ETN
Flexible cyanoacrylate glue	Stick Fast	CA Flexible
Gold connector pin (female)	A-M Systems	520100
Gold connector pin (male)	A-M Systems	520200
ImageJ2/Fiji	Image J	NA
Impactor One	Leica	39463930
LED panel board	Supon	L122t
MATLAB	MathWorks	Institutional or individual activation code
Micro-Manager Camera Software	Image J	NA
Nail drill/polisher	Makartt Nail Drill Electric Nail File	JD700
Omnetic connectors	Omnetics	A79014-001
RHX Data Acquisition Software	Intan	NA
Stainless Steel uncoated wire (0.005")	A-M Systems	792800
Stereotaxic Frame	Kopf Intruments	1923-B
Teets 'cold cure' dental cement	A-M systems	525000, 526000
XLR camera lens	Nikon	NIKKOR 50 mm f/1.8-16