Laser-induced Choroidal Neovascularization

Summary: The choroidal neovascularization model is one of the most popular preclinical models to study pathologic neovascularization in vivo.

Model Description

In 1989, Dobi and colleagues were first to describe a rat choroidal neovascularization model (CNV), which was induced using a krypton laser. Later on, the same induction of choroidal neovascularization was successfully repeated in mice.

Rapid development of different in vivo imaging modalities turned this model into a powerful and validated tool for the screening of novel drug candidates with anti-angiogenic and/or anti-fibrotic properties. This model mimics multiple pathological features of exudative age-related macular degeneration (AMD).

The typical follow-up time is 2 weeks both in mice and in rats. The induction is unilateral leaving the contralateral eye as naïve control.

Currently, intravitreal or systemic (ip) administration of clinically approved aflibercept successfully can prevent vascular leak up to 7 days after the administration (Kaja et al., 2019).

Animal speciesMice, Rats
Method of inductionDiode laser
Follow-up periodTypically 7-14 days
Route of compound administrationTopical (e.g. eye drops), intravitreal or subretinal injections,  systemic
Read-outs1. In vivo imaging
– Fluorescein angiography (FAG)
– Spectral domain-ocular coherence tomography (SD-OCT)
2. Morphological assessment (histology, immunohistochemistry, electron microscopy, stereology)

Outcomes and Read-Outs

In vivo imaging

  • Fluorescein angiography (Heidelberg Spectralis systems, Heidelberg Engineering) is used to monitor clinically relevant vascular leak longitudinally throughout the whole follow-up period (Ragauskas et al., 2018).
Fig. 1. Serial FA imaging immediately after lasering on day 0. 1.
Fig. 1. Serial FA imaging immediately after lasering on day 0. 1. Infrared reflectance image just prior fluorescein administration. Blue arrow point to optic nerve head (ONH). 2. FA image acquired immediately after fluorescein administration. Images 3-18. Increasing retinal leakage with time (each image is taken 20 sec apart) in lasered sites (green arrows in the last image 18). 
  • SD-OCT imaging (Envisu R2210 and Envisu R2210, Bioptigen Inc./Leica Microsystems) provides confirmation of Bruchs’ membrane damage (successful induction), cross-sectional view of pathological features, and enables quantitative evaluation of the lesion (Ragauskas et al., 2018). 
Fig. 2. Visualization of CNV pathology in detail using SD-OCT imaging: follow-up on days 0 through 14 (each lesion individually in red, green and blue).
Fig. 2. Visualization of CNV pathology in detail using SD-OCT imaging: follow-up on days 0 through 14 (each lesion individually in red, green and blue). 

Histology/morphometry

  • Retinal and choroidal flatmounts serve for secondary validation of in vivo findings. We use isolectin B4 staining to detect and, subsequently, quantify neovascularization and collagen 1a for staining of fibrosis.
Fig. 3. Rat choroids stained with isolectin B4 (in greent), which helps to visualize CNV.
Fig. 3. Rat choroids stained with isolectin B4 (in green), which helps to visualize CNV. 

Authors' picture