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Connexin alpha1 and cell proliferation in the developing chick retina
1Department of Anatomy and Developmental Biology, Department of Physiology, University College London, Gower Street, WC1E 6BT, London, UK.
Insights
Reducing connexin alpha1 (connexin 43) expression in chick embryos significantly decreased eye and retina size by inhibiting neurogenesis, not increasing cell death.
Area of Science:
- Developmental biology
- Neuroscience
- Ophthalmology
Background:
- Connexin alpha1 (connexin 43) is highly expressed during eye formation in the cornea, lens, and neural retina.
- Gap-junctional communication plays a role in tissue development and cell communication.
Purpose of the Study:
- To investigate the role of connexin alpha1 in regulating cell proliferation during early eye development.
- To determine if connexin alpha1 influences neurogenesis in the developing retina.
Main Methods:
- A novel antisense oligodeoxynucleotide (ODN) technique was used to reduce connexin alpha1 expression in early chick embryos (embryonic days 2-4).
- Pluronic gel was applied to deliver antisense ODNs to one eye of chick embryos.
- Eye size, retinal area, apoptosis (TUNEL, propidium iodide), and mitotic figures were analyzed in treated and control eyes.
Main Results:
- Antisense ODN application significantly reduced connexin alpha1 protein levels for 24-48 hours.
- Treated eyes showed a marked reduction in diameter and retinal area compared to contralateral untreated eyes.
- Apoptosis was rare in both treated and untreated eyes, indicating reduced cell death was not the cause of smaller retinas. The number of mitotic figures in the ventricular zone of treated retinas was significantly reduced (53% of control).
Conclusions:
- Connexin alpha1-mediated gap-junctional communication is crucial for controlling the early wave of neurogenesis in the chick retina.
- Reduced connexin alpha1 expression inhibits cell proliferation, leading to smaller eye and retinal sizes during development.
Abstract:
During the formation of the eye, high levels of connexin alpha1 (connexin 43) are expressed within the tissues of the cornea, lens, and neural retina. In order to determine whether connexin alpha1 plays a role in the regulation of cell proliferation we have used a novel antisense technique to reduce its expression early in development (embryonic days 2-4). Application of Pluronic gel, containing antisense oligodeoxynucleotides (ODNs) to connexin alpha1, to one eye of early chick embryos results in a rapid and significant reduction of alpha1 protein which lasts for 24-48 h. Embryos grown for 48 h, after ODN application to one eye, showed a marked reduction in the diameter of the treated, compared to that of the contralateral untreated, eye. Sections cut from the treated eyes showed that the retina was also reduced in size. TUNEL labeling and staining with propidium iodide showed that apoptosis within the retinae of both treated and untreated eyes was rare and thus that the reduction in the area of the retina brought about by antisense ODNs directed at connexin alpha1 was unlikely to be the result of increased cell death. However, the number of mitotic figures in the ventricular zone of the antisense-treated retinae revealed by propidium iodide staining was significantly reduced (P < 0.0001) to 53 +/- 3.5% (n = 5) of that in the contralateral untreated control eyes. Embryos in which one eye was sham operated, treated with pluronic gel, or treated with sense ODN showed no significant changes in eye size or in the number of mitotic figures within the neural retina. These results point to a role for connexin alpha1-mediated gap-junctional communication in controlling the early wave of neurogenesis in the chick retina.
