Functional and morphologic study of retinal hypoperfusion injury induced by bilateral common carotid artery occlusion in rats

Abstract

Retinal hypoperfusion injury is the pathophysiologic basis of ocular ischemic syndrome (OIS) which often leads to severe visual loss. In this study, we aimed to establish a rat model of retinal chronic hypoperfusion by bilateral common carotid artery occlusion (BCCAO) and observe changes in the retinal function and morphology. We found that model rats showed retinal arteriosclerosis, slight dilated retinal vein, small hemangiomas, hemorrhages, vascular segmental filling, and nonperfused areas after 2 weeks of BCCAO. In the model rats, the retinal circulation time was significantly prolonged by fluorescein fundus angiography (FFA), the latency of a and b waves was delayed and the amplitude was decreased significantly at each time point by electroretinogram (ERG), and the perfusion of the eyes continued to reduced. Morphologic and ultrastructural changes covered that the retinal ganglion cells (RGCs) presented obvious apoptosis and the thickness in the retinal layers were significantly thinner. Collectively, these findings suggested that BCCAO induced retinal hypoperfusion injury in the model rats, thus providing an ideal animal model for the study of OIS.

Figure 1

Retinal fundus photograph and FFA of rats. The operational requirements: Scope of photo was 30° field and the exposure level was at 50. (a) The tail - retinal circulation time at 1w. (b) The tail - retinal circulation time at 2w. (c) The tail - retinal circulation time at 4w. (d) The comparison of retinal arterial filling time in different groups. (e) The comparison of retinal venous filling time in different groups (n = 8 to 10, ^*P < 0.05 compared with sham group, ^#P < 0.01 compared with sham group).

Figure 2

Retinal scotopic ERG of rats. (a) Changes of latency and amplitude of a- and b-waves at 1,2 and 4 weeks. (b) The latency of the a-wave in different groups. (c) The latency of the b-wave in different groups. (d) The amplitude of the a-wave in different groups. (e) The amplitude of the b-wave in different groups (n = 8 to 10, ^*P < 0.05 compared with sham group, ^#P < 0.01 compared with sham group).

Figure 3

The blood perfusion of rats’ eyes. The operational requirements: constant temperature (23.0 + 1.0 °C), relative humidity (55.0 + 5.0%), no direct sunlight, no obvious air convection and no noise interference. (a) The laser speckle blood flow imager of the eyes’ surface in different groups. (b) Changes in blood perfusion of the eyes’ surface in different groups (n = 8 to 10, ^*P < 0.05 compared with sham group, ^#P < 0.01 compared with sham group).

Figure 4

Retinal histology and morphology in HE. The magnification of retinal images were 10 times (eyepiece) *40 times (objective) with a scale bar of 50 microns (50 μm). (a) Retinal morphologic of different groups at 1, 2 and 4 weeks. (b) The thickness of the RTL in different groups. (c) The thickness of the INL in different groups. (d) The thickness of the ONL in different groups (n = 8 to 10, ^#P < 0.01 compared with sham group).

Figure 5

The Changes in ultrastructure of retinal nerve cells. (a) RGC in sham rats contained large and unbroken nuclei and intact nuclear membranes (thin short white arrow), and rich mitochondria (thin short black arrow) and endoplasmic reticulum (long white arrow). (b) RGC in BCCAO rats showed serious karyopyknosis, chromatic agglutination (thick short black arrow), decreased or swelling mitochondria (thin short black arrow) and vacuolization of endoplasmic reticulum (long black arrow), surrounded with a large number of proliferating microglial cells (thick short white arrow). (c) ONL cells in sham rats were neatly and tightly arranged, with unbroken nuclei (short white arrow). (d) ONL cells in BCCAO rats were with shrunken and invaginated nuclear membranes (short white arrow), and the chromatin was uneven density (short black arrow).

Figure 6

Model of bilateral common carotid artery occlusion. Bilateral carotid arteries of model rats were bluntly separated with vagus nerves and then fastened with 5–0 silk sutures.

Figure 7

Electrodes connection of scotopic ERG. The left was a scotopic ERG check operation of rat, and the right with stopped operation was for clearer rendering.