Evidence for synergistic and complementary roles of Bassoon and darkness in organizing the ribbon synapse.

Ribbon synapses are tonically active high-throughput synapses. The performance of the ribbon synapse is accomplished by a specialization of the cytomatrix at the active zone (CAZ) referred to as the synaptic ribbon (SR). Progress in our understanding of the structure-function relationship at the ribbon synapse has come from observations that, in photoreceptors lacking a full-size scaffolding protein Bassoon (Bsn(ΔEx4/5)), dissociation of SRs coincides with perturbed signal transfer. The aim of the present study has been to elaborate the role of Bassoon as a structural organizer of the ribbon synapse and to differentiate it with regard to the ambient lighting conditions. The ultrastructure of retinal ribbon synapses has been compared between wild-type (Wt) and Bsn(ΔEx4/5) mice adapted to light (low activity) and darkness (high activity). The results obtained suggest that Bassoon and environmental illumination synergistically and complementarily act as organizers of the ribbon synapse. Thus, light-dependent and Bassoon-independent regulation involves initial SR tethering to the membrane and a basic shape transition of ribbon material from spherical to rod-like, since darkness induces these features in Bsn(ΔEx4/5) rod spherules. However, the tight anchorage of the SR via an arciform density and the proper assembly of SRs to the full-sized horseshoe-shaped complex depend on Bassoon, as these steps fail in Bsn(ΔEx4/5) rod spherules.

Synaptic vesicle alterations in rod photoreceptors of synaptophysin-deficient mice.

The abundance of the integral membrane protein synaptophysin in synaptic vesicles and its multiple possible functional contributions to transmitter exocytosis and synaptic vesicle formation stand in sharp contrast to the observed lack of defects in synaptophysin knockout mice. Assuming that deficiencies are compensated by the often coexpressed synaptophysin isoform synaptoporin, we now show that retinal rod photoreceptors, which do not synthesize synaptoporin either in wild-type or in knockout mice, are affected by the loss of synaptophysin. Multiple pale-appearing photoreceptors, as seen by electron microscopy, possess reduced cytoplasmic electron density, swollen mitochondria, an enlarged cell surface area, and, most importantly, a significantly reduced number of synaptic vesicles with an unusually bright interior. Quantification of the number of synaptic vesicles per unit area, not only in these, but also in all other rod terminals of knockout animals, reveals a considerable reduction in vesicles that is even more pronounced during the dark period, i.e., at times of highest synaptic activity. Moreover, activity-dependent reduction in synaptic vesicle diameter, typically occurring in wild-type mice, is not detected in knockout animals. The large number of clathrin-coated pits and vesicles in dark-adapted synaptophysin knockout mice is taken as an indication of compensatory usage of synaptophysin-independent pathway(s), and, conversely, in view of the overall reduction in the number of synaptic vesicles, as an indication for the presence of another synaptophysin-dependent synaptic vesicle recycling pathway. Our results provide in vivo evidence for the importance of the integral membrane protein synaptophysin for synaptic vesicle recycling and formation.

Synaptic ribbon dynamics in photoreceptors of mice.

The present study deals with structural plasticity of a special type of chemical synapse, the ribbon synapse. Near the presynaptic membrane ribbon synapses contain conspicuous electron-dense synaptic bodies which appear mainly as rod-like profiles under the transmission electron microscope. In addition, club-shaped and spherical profiles may be present, the function of which is unclear. To gain some insight into the significance of the latter structures we studied their presence in rod-type photoreceptor cells of BALB/c mice under different lighting conditions. Quantification revealed that the club-shaped and the spherical profiles showed a clear light/dark dependence. They were virtually absent at night and increased strikingly in number when the animals were exposed to light. When darkness was extended into the morning, the profiles remained low in number. As the rod cells diminish their neurotransmitter release during the light phase, the present findings are interpreted as signs of synapse inactivation.

Transient synaptic ribbons in the mammalian retina at unusual sites.

In the vertebrate retina the presence of synaptic ribbons (SRs) is well documented in two sites only, viz., in photoreceptor axon terminals in the outer plexiform layer and in bipolar cell axons in the inner plexiform layer. The present paper reports the presence of non-photoreceptor SRs in the outer plexiform layer of cattle and mouse, where they were seen in small numbers in thin cell processes near cone pedicles of light-adapted animals. They were never seen near rod spherules. Quantitative data obtained in mice killed at different time-points revealed that the SRs under consideration increased in number during day time and were absent during the dark phase. Moreover, under high light intensity of 10000 lux they were more frequent in number compared to 100-lux-exposed animals. It is concluded that the cell processes revealing the temporary presence of SRs are processes of flat bipolar cells which may provide a feedback to cones during the light phase.

Ultrastructural changes of photoreceptor synaptic ribbons in relation to time of day and illumination.

PURPOSE: Electron microscopic sections through rod and cone ribbon synapses reveal mainly rodlike synaptic ribbon profiles, but a few unusual spherical and club-shaped profiles also occur. To elucidate the meaning of the latter two forms, the authors have investigated these ribbon synapses at different times during the 24-hour cycle and under various lighting conditions. METHODS: The various types of ribbon profiles were counted, and their sizes were measured by means of transmission electron microscopy in retinas of male BALB/c mice exposed to 12 hours light (lights on at 6 AM) and 12 hours dark (LD 12:12), continuous light, or continuous darkness for 4 days. RESULTS: A 24-hour study of mice exposed to LD 12:12 showed that spherical and club-shaped profile numbers ranged from 0% to 29%, depending on the time of day. They reached a maximum at 3 hours after light onset, followed by a gradual decrease to approach zero at night and reappearing after light onset the next morning. After 4 days of continuous light, the spherical profiles were significantly decreased in number (examined at 9 AM). After continuous darkness, the spherical and club-shaped profiles were significantly reduced in number. Administration of 4 hours of light after 92 hours of continuous darkness restored the number of spherical and club-shaped profiles to normal values. The rodlike ribbon profiles were found to be longer in darkness than in light. In rod terminals containing spherical profiles, the rodlike ribbon profiles were shorter than in terminals without spherical profiles. CONCLUSIONS. The club-shaped and the spherical profiles were related to the turnover of the synaptic ribbons. Soon after light exposure in the morning, the synaptic ribbons formed distal swellings, giving rise to club-shaped profiles and a decrease in length. The swellings appeared to bud off, thus forming spherical synaptic bodies. This article discusses whether these changes are signs of degradation of spent ribbons, or whether they play a physiological role related to the inactivation of the ribbon synapses after light exposure.

Plasticity of retinal ribbon synapses.

Ribbon synapses differ from conventional chemical synapses in that they contain, within the cloud of synaptic vesicles (SV's), a specialized synaptic body, most often termed synaptic ribbon (SR). This body assumes various forms. Reconstructions reveal that what appear as rod- or ribbon-like profiles in sections are in fact rectangular or horseshoe-shaped plates. Moreover, spherical, T-shaped, table-shaped, and highly pleomorphic bodies may be present. In mammals, ribbon synapses are present in afferent synapses of photoreceptors, bipolar nerve cells, and hair cells of both the organ of Corti and the vestibular organ. Synaptic ribbons (SR's) are also found in the intrinsic cells of the third eye, the pineal gland, and in the lateral line system. The precise function of SR's is enigmatic. The prevailing concept is that SR's function as conveyor belts to channel SV's to the presynaptic membrane for neurotransmitter release by means of exocytosis. The present article reviews the evidence that speaks for a plasticity of these organelles in the retina and the third eye, as reflected in changes in number, size, shape, location, and grouping pattern. SR plasticity is especially pronounced in the mammalian and submammalian pineal gland and in cones and bipolar cells of teleost fishes. Here, SR number and size wax and wane according to the environmental lighting conditions. In the pineal SR numbers increase at night and decrease during the day. In teleost cones, SR's are in their prime during daytime and decrease or disappear at night, when transmitter release is enhanced. In addition to numerical changes, SR's may also exhibit changes in size, shape, grouping pattern, and location. In the mammalian retina of adults, in contrast to the developing retina, the reported signs of SR plasticity are subtle and not always consistent. They may reflect changes in function or may represent signs of degradation. To distinguish between the-two, more detailed studies under selected experimental conditions are required. Probably the strongest evidence for SR plasticity in the mammalian retina is that in hibernating squirrels SR's leave the synaptic site and accumulate in areas as far as 5 microns from the synapse. Changes in shape include the occurrence of club-shaped SR's and round SR's or synaptic spheres (SS's). SS's may represent a special type of synaptic body, yet belonging to the family of SR's, or may be related to the catabolism of SR's. SR number is regulated by Ca2+ in teleost cones, whereas in the mammalian pineal gland cGMP is involved. An interesting biochemical feature of ribbon synapses is that they lack synapsins. The presently reviewed results suggest to us that SR's do not primarily function as conveyor belts, but are devices to immobilize SV's in inactive ribbon synapses.

Morphometry of pineal synaptic ribbon profile numbers after cytochalasin D treatment.

Synaptic ribbons (SRs) are electron-dense, plate-shaped synaptic organelles, to which electron-lucent synaptic vesicles (SVs) are attached by tiny stalks. In the mammalian pineal gland SRs are dynamic organelles, waxing and warning in number under different physiological and experimental conditions. The way in which SRs are formed, or catabolized, is not known. Since it has been suggested that actin may be part of SRs, in the present study the effect of the actin-disrupting drug cytochalasin D (CD, 1 microgram/ml, for 4 h) was examined in cultured guinea pig and rat pineal glands. The glands were preincubated for 48 h so that intra-pineal sympathetic nerve fibres degenerate and no longer release noradrenaline which may distort the results. CD had no effect on SR profile numbers in guinea pigs, but decreased them in rats (p > 0.05). The nonsignificant depressive effect of CD in rats was verified in a second experiment. To clarify the issue, acutely cultured rat pineal glands were treated with CD for 4 h, without effect. The results taken together suggest to us that CD has no major effect on pineal SR profile numbers, but that in rats preincubation for 48 h makes them vulnerable to catabolic processes. In all the experiments, the electron-dense plate of the SRs was qualitatively unaffected. However, the SVs were often larger and more irregular in shape and the stalks linking the SVs with the SRs were less frequently seen in CD-treated glands. In guinea pig pineals, in which SRs frequently lie in groups and parallel to each other, neither the distance between neighboring SRs nor the thickness of individual SRs were affected by CD. It is concluded that actin is not a major component of the SRs and the connecting stalks.