Abstract
Purpose :
Photoreceptors are highly polarized retinal neurons that absorb light and generate an electrical signal through the process of phototransduction. Light-dependent changes in the membrane potential drive glutamate release from the synaptic terminal onto the second order neurons, bipolar cells (BCs) and horizontal cells (HCs). The photoreceptor’s synaptic ribbon facilitates a high rate of exocytosis (vesicle fusion), which is complemented by endocytosis (vesicle retrieval). Endocytosis commonly requires the GTPase dynamin, whose two isoforms, Dnm1 and Dnm3, are expressed specifically in neurons. Here, we evaluated the impact of deleting Dnm1 and 3 on rod photoreceptor structure and function.
Methods :
Dnm1- and Dnm3-floxed mice were bred with transgenic iCre75 mice to generate rod-specific single (rodDnm1-/- and rodDnm3-/-) and double knockouts (rodDnm1-/-/3-/-). ERG recordings were used to test photoreceptor function and signal transmission to BCs. Photoreceptor structure was evaluated by immunohistochemistry, confocal microscopy and electron microscopy.
Results :
Immunostaining confirmed the conditional deletion of Dnm1 and 3 from rods. In rodDnm1-/-/3-/- mice, scotopic ERG a-waves were normal at P90, consistent with normal outer nuclear layer thickness and rod outer segment length; whereas scotopic b-waves were significantly impaired, consistent with a thinning of the outer plexiform layer. Immunostaining of photoreceptor synaptic terminals with ribbon synapse-related proteins (RIBEYE and bassoon) and synaptic vesicle-related proteins (VGLUT1 and complexin-3) showed degeneration of rod terminals. Consistent results were obtained by electron microscopy. In contrast, cone pedicles and the cone-driven ERG in rodDnm1-/-/3-/- mice remained intact. At P90, rodDnm1-/-/3-/- retinas showed a reduced number of HCs (anti-calbindin) and an increase in neurite sprouting from rod BCs (anti- PKCα). No abnormalities of any sort were observed in single knockouts of either Dnm1 or 3, suggesting that their function in rods is redundant.
Conclusions :
Deletion of Dnm1 and 3 from rods causes functional impairment and degeneration of the synaptic terminal. This degeneration is accompanied by structural defects in postsynaptic neurons and loss of light-driven signals in rod BCs. Normal function of rod synapses requires dynamin-dependent endocytosis.
This abstract was presented at the 2022 ARVO Annual Meeting, held in Denver, CO, May 1-4, 2022, and virtually.