Stephen Goldberg

Oriented extracellular channels and axonal guidance in the embryonic chick retina.

Abstract Transmission electron microscopy and serial reconstruction of lum sections were used to determine whether aligned extracellular channels precede the outgrowth of optic fibers in the embryonic chick retina. At stage 16, just prior to the migration of optic axons toward the optic stalk, extracellular spaces bounded by neuroepithelial cell processes, in the superficial (vitread) region of the retina, were aligned toward the optic stalk. The optic axons subsequently entered and grew within these spaces. After formation of the ganglion cell fiber layer (GCFL), the growth cones of new optic axons entered the most vitread portion of that layer. Hypertonic fixatives caused shrinkage of cell processes, resulting in intercellular separation. However, growth cone filopodia retained close contacts with neighboring glial cell endfeet and with optic axons in these solutions. This suggests that growth cones may be adherent to these structures. Transmission electron microscopy in conjunction with the use of hypertonic solutions may become a useful technique for assaying intercellular adhesivity.

Will central nervous systems in the adult mammal regenerate after bypassing a lesion? A study in the mouse and chick visual systems

Abstract We determined whether or not optic axons in the adult mammalian retina would regenerate well if allowed to bypass a lesion scar. In one series we produced microlesions with fine needles in the ganglion cell fiber layer of adult mouse retinas and later examined the retinas as silver-stained flat mounts to observe the behavior of optic axons that bypassed the lesion. Such axons continued to grow abortively, i.e., grew randomly and for only short distances, whether growing within a sector of Wallerian (anterograde) degeneration or in the neighboring zone of intact optic axon bundles. In a second series we produced optic nerve crushes in adult mice and observed the behavior of optic axons growing retrograde into the retina. These fibers similarly grew abortively whether in a zone of intact fiber bundles or when a retinal lesion was produced with the crush, in the sector of Wallerian degeneration. Retinal lesions in newly hatched chicks produced a comparable picture of abortive (short-distance) growth, but the optic and centrifugal fibers had a greater tendency to remain oriented within the ganglion cell fiber layer than did mouse axons. This improved orientation may be the consequence of the greater number of optic fibers in the chick retina and hence the greater opportunity for nonspecific contact guidance. The results indicate that blockage (by a lesion scar or myelin debris) and hypoxia are not the key causes of regenerative failure, as regeneration failed even when those factors were minimal.

Unidirectional, bidirectional and random growth of embryonic optic axons

Abstract Optic fibers were experimentally deflected from their normal courses in the ganglion cell fiber layer (GCFL) of the embryonic chick retina. Fibers deflected sclerad to the GCFL travelled randomly with no tendency to re-enter the GCFL. Fibers growing in the more sclerad portion of the GCFL travelled bidirectionally, i.e. either toward or away from the optic nerve, sometimes exiting through the retinal periphery. Fibers in the most vitread portion of the GCFL tended to grow unidirectionally, toward the optic nerve. The results indicate that the factors that steer optic fibers toward the optic nerve do not operate through the full thickness of the retina but are narrowly confined to the GCFL, possibly to its most vitread portion. The GCFL acts like a glue that entraps but does not attract optic axons.

Axon end-bulb swellings and rapid retrograde degeneration after retinal lesions in young animals

After making a lesion of the retina, anterograde (Wallerian) and retrograde degeneration of optic axons occurred more rapidly in newborn mice than in adults. Axon sprouting occurred only in adult mice, perhaps because retrograde degeneration may have been too rapid and severe for sprouting to occur in newborns. Retinal lesions in mice of any age produced end-bulb swellings initially on both sides of the lesion. In all animals, dense packing of lysosomes and other organelles occurred in end-bulbs on the side of Wallerian degeneration but did not occur in end-bulbs on the retrograde side, where accumulation of smooth endoplasmic reticulum was the most characteristic change. Retrograde end-bulbs appeared much like growth cones, which sprouted in adults, but degenerated in younger animals. Continuing daily enlargement of end-bulb swellings was noted on the Wallerian side of lesions in adults, but not in newborns. Such enlargement is believed to have resulted from retrograde axoplasmic transport and suggests that such transport may be greater in adults than in newborns.

Pyridine-Silver Methods for the Study of Optic Axons in Retinal Whole Mounts

The optic fibers in the retinas of diverse species may be selectively stained and viewed en bloc in the embryonic and adult states. Treat the eye as follows: 1) 50% pyridine for at least 16 hr, 2) distilled water 3--4 hr, 3) 20% H2O2 until the eye is a light brown, 4) 95% ethanol overnight, 5) 1.5% AgNO3 for 2--6 days at 37 C, 6) in water, remove the vitreous, then direct 0.25% pyrogallic acid in 1.25% formalin against the retina for 2--5 secs until the optic fibers are reduced to a coffee-copper color (1--4 minutes), 7) dissect the retina and mount flat on a glass slide, 8) cover with glycerin, apply a coverslip, and fix in place with nail polish. Variants for particular species are given. The technique offers an advantage over Golgi and methylene blue methods which tend to stain only a small percentage of fibers and frequently do not work at the earliest stages of development.

Progressive fixation of morphological polarity in the developing retina.

Abstract Fixation of morphological polarity may not be an all-or-none event. The optic fissure of the chick retina normally lies ventrally. When optic vesicles were rotated 180°, the fissures later appeared ventral provided the operations were performed prior to Hamburger-Hamilton stage 12. Operations beyond stage 12 resulted in dorsal fissures. The lengths of the fissures varied in a manner indicating a gradual rather than abrupt change from ventral to dorsal with progressive stages of operation.

Retina as a model system in paraplegia research: Pharmacologic studies

Abstract The retina may be a useful model system for the rapid testing of potentially neuroregenerative agents. We tested 24 different pharmacologic agents for their ability to promote regeneration of severed optic axons in the adult mouse retina. In this initial series of 738 mice, these agents were ineffective in reducing scar formation, stimulating axonal elongation, improving axonal guidance or retarding retrograde degeneration.