Photomembrane turnover in frog retina: light intensity and spectral correlates.

Light regulates membrane turnover in vertebrate rod photoreceptor cells. Rods shed membrane-filled tips immediately after light onset, with light inhibiting the dark priming phase but initiating the light induction phase. This study examines the intensities and wavelengths of light that control these two shedding requirements, and demonstrates unexpected situations where red or dim lights are simultaneously dark to the dark priming mechanism and light to the light induction process. Since shedding takes place immediately following darkness we asked if dim or red light could substitute for true darkness and dark prime the retinas: our results confirm this. White light, less than 0.7 microE m m-2 sec-1 (0.15 W m2 or 40 lx), allows dark priming, and even 15 microE m-2 sec-1 of red fluorescent light dark primes as effectively as true darkness. Conversely, bright white light and wavelengths from 480 to 560 nm inhibit dark priming, implying that dark priming inhibition is a photopic mechanism transduced by photopigment in the 502-cone. We also asked if dim or red light could induce shedding, substituting for the bright light usually employed: again, the results confirm thus. White light as dim as 0.15 microE m-2 sec-1 induces shedding and red light is an effective light trigger. This light induction is initiated at all wavelengths tested (420-640 nm), with a maximum effect between 540 and 600 nm. Finally, we find that retinas shed continuously in red or dim white light. These lights substitute both for the darkness necessary for dark priming and for the light of light induction, extending shedding from the 20 min dark-light transition period to hours or days. We also find that the dim, red light of natural dawn is as effective a shedding stimulus as the sudden onset of bright laboratory light.

Paracrine control of photomembrane removal.

Photomembrane turnover in vertebrate photoreceptors is regulated by light. Rod outer segments (ROS) shed membrane filled tips at light onset, during the coexistence of two light modulated processes: a dark priming factor and a light induction event. Transduction of these two signals is not direct but appears to involve the neural retina and diffusible paracrine molecules. I propose a model wherein three paracrines control this ROS tip shedding. Melatonin, a lipid soluble dark priming molecule, is synthesized in the dark by all photoreceptor cells, diffusing freely and separating the ROS disk membranes. A second paracrine, dopamine is released from the inner retina whenever light is absorbed by the 502 nm-cones, inhibiting melatonin synthesis. Third, a proposed trophic paracrine, "rostrophin", is released in the dark from internal horizontal cells, and stabilizes the photomembrane. Shedding occurs as rostrophin decreases in the presence melatonin; briefly at light onset or continuously in red or dim white light.

Light absorbed by 575-cones trigger rod disc shedding in the frog retina.

Photoreceptors periodically shed light-sensitive membranes; rods at light onset, cones at night. The spectral characteristics of light required to initiate shedding by 502-rods were studied in the frog retina. After 24 h of white light, animals were dark primed for 1 h then presented with 1 h of nearly monochromatic light to induce the shedding response. The light delivered a total photon flux of 2.5 or 0.5 microE/m2s. Following conventional fixation and plastic embedding, 1 micron sections were examined with light microscopy. The number of photoreceptor tips (phagosomes) shed by 100 consecutive rods were counted and plotted as a function of wavelength. Bright light induced at least 15 phagosomes per 100 rods at all wavelengths tested, 420-640 nm, and this shedding was more than doubled with light from 540-600 nm. When the light was dimmed, there was no shedding response except for this 540-600 nm window. This shedding peak closely corresponds to the absorbance curve of the frog's 575-cone photopigment and implies that the 575-cone can drive rod shedding. The broad background effect further indicates that all photoreceptors have an input and suggests that a luminosity cell, such as the internal horizontal cell, may be involved.

Ultrastructure of non-myelinated neurons during energy deprivation.

This paper examines the neuropathology of oxygen-glucose deprivation uncomplicated by stagnant conditions. Rabbit vagus nerves were pulled into a multi-compartment perfusion chamber, stimulated five times per second and deprived of energy by substituting nitrogen and deoxyglucose for oxygen and glucose in the Locke's perfusate. After incubation the compartments were perfused with gluteraldehyde solution, and the nerves were prepared for electron microscopy. Fixation in the compartments ensured precise cross and longitudinal sections which permitted quantitative comparisons. Although the action potentials ceased in 45 min, 1 h of energy deprivation did not significantly affect the ultrastructure. After 2 h of deprivation the axons were smaller and flattened and microtubules appeared packed together. In the smallest axons the microtubules were gone, the neurofilaments were compacted and the few mitochondria had a dense, homogenous appearance. By 4 h the shrinking was extreme, yet 8% were swollen much larger than any of the controls. Longitudinal views showed these ballooned areas were greatly expanded regions of the smallest axons. Both tiny and huge regions were devoid of microtubules and the swollen axons contained expanded mitochondria. Calcium is indirectly implicated in the pathogenesis by the concurrence of mitochondrial alteration as the microtubules disappear coupled with the known role of mitochondria in calcium regulation and the reported effect of high calcium on microtubual dissociation. It is suggested that axons first shrink as osmotically active molecules are used or washed out. After a time without energy the mitochondria can no longer regulate the intracellular calcium, microtubules dissociate, and calcium-activated phospholipases create osmotically active molecules. Finally, high-amplitude, disruptive swelling occurs.