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Updated: Jun 2, 2026
Glaucoma-inducing Procedure in an In Vivo Rat Model and Whole-mount Retina Preparation
Published on: March 12, 2016
Elevation of p-NR2A(S1232) by Cdk5/p35 contributes to retinal ganglion cell apoptosis in a rat experimental glaucoma
Jie Chen1, Yanying Miao, Xiao-Han Wang
1Institutes of Brain Science, Institute of Neurobiology and State Key Laboratory of Medical Neurobiology, Fudan University, Shanghai 200032, China.
Abstract:
Glaucoma, mainly caused by high intraocular pressure (IOP), is characterized by apoptotic death of retinal ganglion cells (RGCs). We investigated the possible involvement of cyclin-dependent kinase 5 (Cdk5) and its activator p35, which have been implicated in a variety of neurological disorders, in RGC apoptosis in a rat experimental glaucoma model reproduced by blocking episcleral veins. Cholera toxin B subunit (CTB) retrogradely labeled RGCs displayed a dramatic reduction in number both in the central and peripheral retina on day 14 (D14) (P<0.05 vs. control), D21 (P<0.01 vs. control) and D28 (P<0.001 vs. control) after operation. Terminal dUTP nick end labeling (TUNEL)-positive cells were detected on D14 both in the central and peripheral regions, and numerous TUNEL-positive cells were found on D21 and D28 in both the regions (P all<0.001 vs. control). As compared with the control eyes, the expression level of Cdk5 was significantly increased on D21 (P<0.001), whereas that of p35 displayed a marked increase on D14 (P<0.01) and D21 (P<0.001). Meanwhile, both NR2A and p-NR2A(S1232) increased from D14 onwards (P<0.01 to 0.001). Co-immunoprecipitation indicated a direct interaction between Cdk5 and p-NR2A(S1232). Intraperitoneal injection of the Cdk5 inhibitor roscovitine remarkably inhibited RGC apoptosis (P<0.001 vs. vehicle group) and increased the number of CTB-labeled RGCs (P<0.05 to 0.01 vs. vehicle group) in whole flat-mounted retinas, which was accompanied by a significant decrease in expression levels of p35 and p-NR2A(S1232) (P all<0.01 vs. vehicle group). Our results suggest that elevation of p-NR2A(S1232) by Cdk5/p35 contributes to RGC apoptotic death in experimental glaucoma rats, which could be effectively ameliorated by inhibiting Cdk5/p35.
Insights
Cyclin-dependent kinase 5 (Cdk5) and its activator p35 contribute to retinal ganglion cell (RGC) apoptosis in glaucoma. Inhibiting Cdk5/p35 with roscovitine effectively reduced RGC death in an experimental glaucoma model.
Area of Science:
- Ophthalmology
- Neuroscience
- Cell Biology
Background:
- Glaucoma, often caused by elevated intraocular pressure (IOP), leads to retinal ganglion cell (RGC) apoptosis.
- Cyclin-dependent kinase 5 (Cdk5) and its activator p35 are implicated in neurological disorders and may play a role in RGC death.
Purpose of the Study:
- To investigate the involvement of Cdk5/p35 in RGC apoptosis in a rat model of experimental glaucoma.
- To explore the therapeutic potential of Cdk5 inhibition in mitigating glaucoma-induced RGC loss.
Main Methods:
- An experimental glaucoma model was created in rats by blocking episcleral veins.
- RGC apoptosis was assessed using Cholera toxin B subunit (CTB) labeling and Terminal dUTP nick end labeling (TUNEL).
- Expression levels of Cdk5, p35, NR2A, and phosphorylated NR2A (p-NR2A(S1232)) were analyzed.
- Co-immunoprecipitation was used to determine the interaction between Cdk5 and p-NR2A(S1232).
- The effect of the Cdk5 inhibitor roscovitine on RGC apoptosis and related protein expression was evaluated.
Main Results:
- Experimental glaucoma led to a significant reduction in RGC numbers and increased TUNEL-positive cells.
- Cdk5 and p35 expression levels were elevated in glaucoma eyes, alongside increased NR2A and p-NR2A(S1232).
- Cdk5 directly interacted with p-NR2A(S1232).
- Roscovitine treatment significantly inhibited RGC apoptosis and decreased p35 and p-NR2A(S1232) levels.
Conclusions:
- The Cdk5/p35 pathway, by elevating p-NR2A(S1232), contributes to RGC apoptotic death in experimental glaucoma.
- Inhibition of Cdk5/p35 demonstrates a promising therapeutic strategy for ameliorating RGC loss in glaucoma.
