Abstract
Retrograde labelling of retinal ganglion cells with optic nerve transection often leads to degeneration of ganglion cells in prolonged experiments. Here we report that an intact optic nerve could uptake retrograde tracers applied onto the surface of the nerve, leading to high efficiency labelling of ganglion cells in the retina with long-term survival of cells. This method labelled a similar number of ganglion cells (2289±174 at 2 days) as the retrograde labeling technique from the superior colliculus (2250±94) or optic nerve stump (2279±114) after transection. This finding provides an alternative way to label retinal ganglion cells without damaging the optic tract. This will facilitate anatomical studies in identifying the morphology and connectivity of retinal ganglion cells, allowing secondary or triple labelling manipulations for long-term investigations.
Highlights
We occasionally found that retinal ganglion cells (RGCs) could uptake and be labelled by retrograde labelling dyes applied onto the surface of the intact Optic nerve (ON), suggesting for a possible way to label most of the RGCs without damaging the optic tract
Intact ON labelling leads to high efficiency of retrograde RGC filling We found that in our study, superior colliculus (SC) labelling and ON stump labelling are able to label most RGCs in the retina, as previously reported
Applying FG onto the surface of the ON was sufficient for labelling of RGCs after 2 days of labelling (Fig 1), with the same levels of fluorescence intensity and soma filling of the RGCs (Fig 2)
Summary
The use of animals followed the requirements of the Cap. 340 Animals (Control of Experiments) Ordinance and Regulations in Hong Kong. All the experimental and animal handling procedures were approved by the University of Hong Kong Animal Ethics Committee (committee of use of laboratory animals for teaching and research, CULATR, CULATR # 1937–09). The animals were housed with free access to food and water under a 12-hour light/12-hour dark cycle (7:00 AM–7:00 PM). For optic stump and intact ON retrograde labelling, 0.5% alcaine (AlconCouvreur, Puurs, Belgium) was applied to the eyes prior to the surgery, and antiseptic eye drops (Tobres [Tobramycin 0.3%]; lcon-Couvreur) were used to prevent infection after the procedures. Finadyne (0.025 mg/mL) in drinking water was applied for 7 days after surgery to relieve the pain when needed. All animals were sacrificed with overdose of pentobarbital at different time points
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