Purpose Cone photoreceptors of the retina use a sophisticated ribbon-containing synapse to convert light-dependent changes in membrane potential into release of synaptic vesicles (SVs). We aimed to study the functional and structural maturation of mouse cone photoreceptor ribbon synapses during postnatal development and to investigate the role of the synaptic ribbon in SV release. Methods We performed patch-clamp recordings from cone photoreceptors and their postsynaptic partners, the horizontal cells during postnatal retinal development to reveal the functional parameters of the synapses. To investigate the occurring structural changes, we applied immunocytochemistry and electron microscopy. Results We found that immature cone photoreceptor terminals were smaller, they had fewer active zones (AZs) and AZ-anchored synaptic ribbons, and they produced a smaller Ca 2+ current than mature photoreceptors. The number of postsynaptic horizontal cell contacts to synaptic terminals increased with age. However, tonic and spontaneous SV release at synaptic terminals stayed similar during postnatal development. Multiquantal SV release was present in all age groups, but mature synapses produced larger multiquantal events than immature ones. Remarkably, at single AZs, tonic SV release was attenuated during maturation and showed an inverse relationship with the appearance of anchored synaptic ribbons. Conclusions Our developmental study suggests that the presence of synaptic ribbons at the AZs attenuates tonic SV release and amplifies multiquantal SV release. However, spontaneous SV release may not depend on the presence of synaptic ribbons or voltage-sensitive Ca 2+ channels at the AZs.

It is a commonly accepted view that light stimulation of mammalian photoreceptors causes a graded change in membrane potential instead of developing a spike. The presynaptic Ca 21 channels serve as a crucial link for the coding of membrane potential variations into neurotransmitter release. Ca v 1.4 L-type Ca 21 channels are expressed in photoreceptor terminals, but the complete pool of Ca 21 channels in cone photoreceptors appears to be more diverse. Here, we discovered, employing whole-cell patch-clamp recording from cone photoreceptor terminals in both sexes of mice, that their Ca 21 currents are composed of low-(T-type Ca 21 channels) and high-(L-type Ca 21 channels) voltage-activated components. Furthermore, Ca 21 channels exerted self-generated spike behavior in dark membrane potentials, and spikes were generated in response to light/ dark transition. The application of fast and slow Ca 21 chelators revealed that T-type Ca 21 channels are located close to the release machinery. Furthermore, capacitance measurements indicated that they are involved in evoked vesicle release. Additionally, RT-PCR experiments showed the presence of Ca v 3.2 T-type Ca 21 channels in cone photoreceptors but not in rod photoreceptors. Altogether, we found several crucial functions of T-type Ca 21 channels, which increase the functional repertoire of cone photoreceptors. Namely, they extend cone photoreceptor light-responsive membrane potential range, amplify dark responses, generate spikes, increase intracellular Ca 21 levels, and boost synaptic transmission.

No abstract

Mammalian cone photoreceptors enable through their sophisticated synapse the high-fidelity transfer of visual information to second-order neurons in the retina. The synapse contains a proteinaceous organelle, called the synaptic ribbon, which tethers synaptic vesicles (SVs) at the active zone (AZ) close to voltage-gated Ca2+ channels. However, the exact contribution of the synaptic ribbon to neurotransmission is not fully understood, yet. In mice, precursors to synaptic ribbons appear within photoreceptor terminals shortly after birth as free-floating spherical structures, which progressively elongate and then attach to the AZ during the following days. Here, we took advantage of the process of synaptic ribbon maturation to study their contribution to SV release. We performed whole-cell patch-clamp recordings from cone photoreceptors at three postnatal (P) development stages (P8–9, P12–13, >P30) and measured evoked SV release, SV replenishment rate, recovery from synaptic depression, domain organization of voltage-sensitive Ca2+ channels, and Ca2+-sensitivity of exocytosis. Additionally, we performed electron microscopy to determine the density of SVs at ribbon-free and ribbon-occupied AZs. Our results suggest that ribbon attachment does not organize the voltage-sensitive Ca2+ channels into nanodomains or control SV release probability. However, ribbon attachment increases SV density at the AZ, increases the pool size of readily releasable SVs available for evoked SV release, facilitates SV replenishment without changing the SV pool refilling time, and increases the Ca2+- sensitivity of glutamate release.