The expression pattern and assembly profile of synaptic membrane proteins in ribbon synapses of the developing mouse retina

Abstract

In the present study, we generated a systematic overview of the expression pattern and assembly profile of synaptic membrane proteins in ribbon synapses of the developing mouse retina. Using indirect immunofluorescence microscopy, we analyzed the spatial and temporal distribution of 11 important membrane and membrane-associated synaptic proteins (syntaxin 1/3, SNAP-25, synaptobrevin 2, synaptogyrin, synaptotagmin I, SV2A, SV2B, Rab3A, clathrin light chains, CSP and neuroligin I) during synaptogenesis. The temporospatial distribution of these synaptic proteins was "normalized" by the simultaneous visualization of the synaptic vesicle protein synaptophysin, which served as an internal reference protein. We found that expression of various synaptic membrane proteins started at different time points and changed progressively during development. At early stages of development synaptic vesicle membrane proteins at extrasynaptic locations did not always colocalize with synaptophysin, indicating that these proteins probably do not reside in the same transport vesicles. Despite a non-synchronized onset of protein expression, clustering and colocalization of all synaptic membrane proteins at ribbon synapses roughly occurred in the same time window (between day 4 after birth, P4, and P5). Thus, the basic synaptic membrane machinery is already present in ribbon synapses before the well-known complete morphological maturation of ribbon synapses between P7 and P12. We conclude that ribbon synapse formation is a multistep process in which the concerted recruitment of synaptic membrane proteins is a relatively early event and clearly not the final step.

Calcium Channel-Dependent Molecular Maturation of Photoreceptor Synapses

“…Our anatomical data are in line with developmental findings suggesting that photoreceptor terminals first acquire basic synaptic properties during the first postnatal week, before becoming ribbon synapses during the second postnatal week [32] and completing their maturation after eye opening [54].…”

“…The rabbit anti-VAMP2 (Abcam) reacts with a single 19 kDa band (manufacturer's technical information). The expression pattern obtained here was consistent with previous reports [32].…”

“…Synaptobrevin 2 (VAMP2) is a synaptic vesicle-associated v-SNARE protein and is expressed in photoreceptor synapses [32]. The rabbit anti-VAMP2 (Abcam) reacts with a single 19 kDa band (manufacturer's technical information).…”

Transplantation of Embryonic and Induced Pluripotent Stem Cell-Derived 3D Retinal Sheets into Retinal Degenerative Mice

“…Close observation of the direct-contact pattern showed that the inner part of the graft retina (other than ONL) was being “stripped off,” or pushed aside, enabling the bare graft ONL cells to make a direct contact with host bipolar cells in the correct orientation (Figures S3A and S3A’). This could also imply that the graft photoreceptors in young grafts developed in a structured ONL/INL complex nearly to the stage of synaptogenesis (i.e., around postnatal days 4–7; von Kriegstein and Schmitz, 2003), but somehow they formed host-graft synapses instead of intragraft synapses that we also observed in the neighboring area of host-graft synaptogenesis (Figures S4E–S4J). In some host-graft border area, both the host and graft bipolar cells sent dendrite terminals over the graft photoreceptor terminals of the same area.…”

Phosphorylation of cysteine string protein in the brain: developmental, regional and synaptic specificity

“…A recent developmental study of synaptic vesicle protein expression in the mouse retina as assessed by immunohistochemistry compares favourably with our data (Kriegstein & Schmitz, 2003). Analogous to the development of the cerebellar cortex, the early postnatal retina comprises layers of neuronal precursors that migrate and differentiate to form distinct layers of neurons and synapses, with the majority of synaptogenesis occurring in the second postnatal week (Olney, 1968).…”