Fluorescence Angiography for Evaluation of Aneurysm Perfusion and Parent Artery Patency in Rat and Rabbit Aneurysm Models

Summary

We present a protocol to efficiently evaluate aneurysm perfusion and vessel patency of sidewall aneurysm in rats and rabbits, using fluorescein-based fluorescence video angiography (FVA). With a positive predictive value of 92.6%, it is a simple but very effective and economical method with no special equipment required.

Abstract

Brain aneurysm treatment focuses on achieving complete occlusion, as well as preserving blood flow in the parent artery. Fluorescein sodium and indocyanine green are used to enable the observation of blood flow and vessel perfusion status, respectively. The aim of this study is to apply FVA to verify real-time blood flow, vessel perfusion status and occlusion of aneurysms after induction of sidewall aneurysms in rabbits and rats, as well as to validate the procedure in these species.

Twenty sidewall aneurysms were created in 10 rabbits by suturing a decellularized arterial vessel pouch on the carotid artery of a donor rabbit. In addition, 48 microsurgical sidewall aneurysms were created in 48 rats. During follow-up at one month after creation, the parent artery/aneurysm complex was dissected and FVA was performed using an intravenous fluorescein (10%, 1 mL) injection via an ear vein catheterization in rabbits and a femoral vein catherization in rats. Aneurysms were then harvested, and patency was evaluated macroscopically.

Macroscopically, 14 out of 16 aneurysms in rabbits indicated no residual parent artery perfusion with totally occluded luminae, however 11 (79%) were detected by FVA. Four aneurysms were excluded due to technical problems. In rats, residual aneurysm perfusion was macroscopically observed in 25 out of 48 cases. Of the 23 without macroscopic evidence of perfusion, FVA confirmed the incidence of 22 aneurysms (96%). There were no adverse events associated with FVA. Fluorescein is easily applicable and no special equipment is needed. It is a safe and extremely effective method for evaluating parent artery integrity and aneurysm patency/residual perfusion in an experimental setting with rabbits and rats. FVA using fluorescein as a contrast agent appears to be effective in controlling patency of aneurysms and the underlying vessel and can even be adapted to bypass surgery.

Introduction

Evidence of complete aneurysm obliteration and parent artery integrity is of utmost importance in aneurysm surgery. There are several options to confirm parent artery patency and aneurysm occlusion, such as Doppler sonography, conventional cerebral angiography (DSA), computed tomography angiography (CTA) or magnetic resonance angiography (MRA)^1,2. However, these are expensive and time-consuming methods which are often not available in a laboratory setting. Furthermore, they may have relevant side effects such as radiation exposure or need for additional sedation of experimental animals to avoid movement artefa....

Protocol

The rodents were housed in an animal care facility and experiments were reviewed and approved by the Committee for Animal Welfare at the University of Bern, Switzerland (BE 108/16) and (BE65/16). All animals were maintained on a standard laboratory diet with free access to food and water. All animal experiments were conducted under careful consideration of the 3Rs (replacement, reduction and refinement). Ten female New Zealand White rabbits and 48 male Wistar rats were included. ARRIVE guidelines were followed strictly⁸.

NOTE: Twenty sidewall aneurysms were created in 10 rabbits by suturing a decellularized arterial ....

Results

Heart rate and blood pressure were monitored during surgery. Mean heart rate was 193/min in rabbits and 196/min in rats. The rabbits' body weight ranged 3.05-4.18 kg, and the rats weighed 335-690 g.

We were able to perform FVA in eight out of ten rabbits (Figure 1). Four aneurysm examinations in two rabbits were not recorded with the camera due to technical difficulties. No technical difficulties involving FVA in rats were reported. However, FVA could not be p.......

Discussion

FVA is a promising and uncomplicated method to examine vessels in rodents and can be performed with commercial devices and off-the-shelf equipment. FVA can be implemented during any surgery where intraoperative evaluation of vessel integrity is needed as the vessels need proper dissection first.

The authors preferred venous injection to arterial injection due to the lower risk of inadvertent events such as infection, ischemia and compartment syndrome¹². Intravenous inje.......

Materials

Name	Company	Catalog Number	Comments
For rabbits
Aluminium foil
Animal shaver
Black tape
Blue filter	Thorlabs MF475-35
Body warm plate
Camera	Sony NEX-5R
Catheter	22G Vasofix Safety
Disinfictant
Fluorescein sodium	Fluorescein Faure 10%
Glas plate
Green filter	Thorlabs MF539-43
Incontinence pad
Infusion pump	Perfusor Secura
Ketamine hydrochloride			any generic products
Needle	25G
Oxygen
Ringer's Solution
Sterile sheets
Surgical instruments			micro forceps, micro scissor, blunt surgical scissor
Surgical microscope	OPMI, Carl Zeiss AG, Oberkochen, Germany
Syringe 2ml, 5ml, 50ml
Tape
Three-way-stopcock
Torch light
Xylazin			any generic products
For rats
Aluminium foil
Animal shaver
Black tape
Blue filter	Thorlabs MF475-35
Body warm plate
Camera	Sony NEX-5R
Disinfictant
Fluorescein sodium	Fluorescein Faure 10%
Green filter	Thorlabs MF539-43
Incontinence pad
Isoflurane
Ketamine hydrochloride			any generic products
Medetomidine hydrochloride			any generic products
Needle	25G
Oxygen
Plate
Ringer's Solution
Sterile sheets
Surgical instruments			micro forceps, micro scissor, blunt surgical scissor
Surgical microscope	OPMI, Carl Zeiss AG, Oberkochen, Germany
Syringe 2ml, 5ml
Tape
Torch light