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W.H. Baldridge, R.M. Abdel-Majid, M. Archibald, F. Tremblay; Tracer Coupling of Neurons in the Rat Retina Inner Nuclear Layer Labeled by Fluorogold . Invest. Ophthalmol. Vis. Sci. 2003;44(13):4297.
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© ARVO (1962-2015); The Authors (2016-present)
Purpose:There is considerable evidence of tracer coupling between ganglion cells (GCs) and amacrine cells (ACs) in the vertebrate retina. However, coupling has for the most part only been observed when tracer is injected into GCs. When tracer is injected into ACs it is usually restricted to the injected cell or passes only to other ACs. One possible explanation is that those ACs injected with tracer are not those coupled to GCs. Therefore, we have taken advantage of the movement of Fluorogold (FG) from GCs to ACs to target those ACs "tracer" coupled to GCs for injection with Neurobiotin (NB) to determine if these ACs will show NB tracer coupling to GCs. Methods: Rat GCs were retrogradely loaded with FG by application to the superior colliculus. After 12 weeks the retinas of these animals were isolated, mounted on filter paper and maintained in oxygenated Ames medium. Neurons in the inner nuclear layer (INL) labeled with FG were identified using a filter set that also permitted visualization of microelectrodes filled with 1% Lucifer yellow and 4% NB. Cells were impaled under visual guidance and injected with NB by iontophoresis (+1 nA, 500 msec duration, 1 Hz, for 2 min). Thirty minutes following the final injection, the tissue was fixed in 4% paraformaldehyde for 2 hrs, washed and then incubated overnight in Streptavidin conjugated to Cy3 (1:100) in PBS containing 1% Triton X-100. Results: Two types of FG-labeled neurons were encountered in the INL of the rat retina. At low density but well labeled with FG were neurons which, following NB injection, were found to possess an axon that ramified to the nerve fibre layer. It was concluded that these neurons were GCs displaced to the INL. Some of these neurons showed tracer coupling with patterns similar to that observed for conventionally placed GCs. The other type of FG-labeled cells in the INL were present at greater density but were weakly labeled with FG. After NB injection these neurons were identified as ACs. Every FG-labeled AC injected with NB showed tracer coupling to other ACs in the INL but, under the injection conditions used, did not show coupling to GCs. Conclusions: The tracer NB does not pass from ACs to GCs, even when injected into ACs known to be coupled to GCs. This adds to the evidence that tracer coupling between GCs and ACs may be unidirectional. The tracer coupling of displaced GCs is consistent with the view that the connectivity of these cells is similar to conventional GCs despite the different placement within the retina.
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