Abstract
Purpose::
Mixed-rod cone bipolar (Mb) cells of goldfish retina have large synaptic terminals that make 50-70 ribbon synapses mostly onto amacrine cells, rarely to ganglion cells and receive 300-400 synapses from GABAergic amacrine cells. However, ganglion cells receive relatively rare input from bipolar cells, insufficient to account for the extensive physiological input from Mb bipolar cells. Here we demonstrate the existence of dendrites that penetrate deeply into Mb bipolar cell terminals and identify the origin of the dendrites.
Methods::
Goldfish retinas were fixed in 4% paraformaldehyde and incubated in combinations of the following primary antibodies: anti-PanNa, anti-GAD67, anti-FRMF, anti-PKC, or anti-TH, anti-Chat, or anti-SRIF, anti-Geph, anti-GFAP, or anti-PKC. Additionally, ganglion cells were backfilled with crystals of tetramethylrhodamine dextran that were placed on the cut optic nerve for 4-6 days at 10°C. Retinas were processed for LM and EM immunocytochemistry and confocal microscopy.
Results::
Tissue viewed by electron microscopy (EM) revealed the presence of double membrane bound processes deep within Mb terminals. These processes were unlike amacrine cell intrusions described previously by this laboratory in that they deeply invaginated the terminals as opposed to the more superficial amacrine intrusions. There was no indication of membrane specializations on these invaginating processes, although rare vesicular fusion was observed. Dendrites of GABAergic amacrine cells and ganglion cells had processes that invaginated Mb terminals. In addition, an antibody against all voltage-gated sodium channels (PanNa) revealed PanNa-IR processes that deeply penetrated Mb terminals. Confocal analysis clearly showed processes that extended into and through the Mb terminal, like "worms in an apple."
Conclusions::
Dendritic processes penetrate Mb terminals without making synaptic contacts. There are at least two (amacrine and ganglion cells) and possibly a third source (PanNa-IR processes) of these intrusions. As there is no evidence of synaptic specializations, the invaginating dendrites may be sites of ephaptic transmission. These dendrites may provide a mechanism for the transmission of signals from Mb bipolar cells to ganglion cells and to a subtype of GABAergic amacrine cell that bypasses conventional ribbon synapses.
Keywords: retina: proximal (bipolar, amacrine, and ganglion cells) • bipolar cells • immunohistochemistry