Roy H. Steinberg

Recent demonstrations of survival-promoting activity by neurotrophic agents in diverse neuronal systems have raised the possibility of pharmacological therapy for inherited and degenerative disorders of the central nervous system. We have shown previously that, in the retina, basic fibroblast growth factor delays photoreceptor degeneration in Royal College of Surgeons rats with inherited retinal dystrophy and that the growth factor reduces or prevents the rapid photoreceptor degeneration produced by constant light in the rat. This light-damage model now provides an efficient way to assess quantitatively the survival-promoting activity in vivo of a number of growth factors and other molecules. We report here that photoreceptors can be significantly protected from the damaging effects of light by intravitreal injection of eight different growth factors, cytokines, and neurotrophins that typically act through several distinct receptor families. In addition to basic fibroblast growth factor, those factors providing a high degree of photoreceptor rescue include brain-derived neurotrophic factor, ciliary neurotrophic factor, interleukin 1 beta, and acidic fibroblast growth factor; those with less activity include neurotrophin 3, insulin-like growth factor II, and tumor necrosis factor alpha; those showing little or no protective effect are nerve growth factor, epidermal growth factor, platelet-derived growth factor, insulin, insulin-like growth factor I, heparin, and laminin. Although we used at least one relatively high concentration of each agent (the highest available), it is still possible that other concentrations or factor combinations might be more protective. Injecting heparin along with acidic fibroblast growth factor or basic fibroblast growth factor further enhanced the degree of photoreceptor survival and also suppressed the increased incidence of macrophages produced by either factor, especially basic fibroblast growth factor. These results now provide the impetus for determining the normal function in the retina, mechanism(s) of rescue, and therapeutic potential in human eye diseases for each agent.

Electron microscopic examination of the bases of adult rod and cone outer segments (rhesus monkey, ground squirrel, and grey squirrel) has led to a new model of disc morphogenesis. In this model the disc surfaces and disc rims develop by separate mechanisms and from separate regions of the membrane of the inner face of the cilium. This membrane is alternately specified into regions that will form either the disc surfaces or the disc rims. The disc surfaces develop by an evagination or outpouching of the ciliary membrane. The two surfaces of an evagination, scleral and vitreal, each form one of the surfaces of adjacent discs. The disc rim is initially specified as a region of ciliary membrane between adjacent disc-surface evaginations. This region grows bilaterally around the circumferences of adjacent discs, zippering together the apposed surfaces to form the rim and completed disc. At the same time it seals the plasma-membrane edges of the evaginations, which have become detached from the surfaces. Incisures form in rod discs by infolding of the rim and surfaces together, and they begin to form before the rim is completed around the disc perimeter. When a number of new discs are developing simultaneously the ciliary membrane at the base of an outer segment consists of a stack of rim forming and surface forming growth points. This model provides, in addition, for the continuous renewal of outer-segment plasma membrane. It also establishes a developmental basis for the structural uniqueness of the disc rim. Finally, it indicates an evolutionary relationship between the discs of vertebrate visual cells and the membrane specializations of invertebrate visual cells.

Abstract The distribution of rods and cones in the cat retina was studied by light microscopy. The rods and cones were counted from the area centralis to the temporal periphery in photomicrographs of transverse sections through the inner segments. A 16% correction for the effect of tissue shrinkage was applied to the densities obtained in fixed‐dehydrated tissue, by comparing these counts with those obtained from similar retinal areas in fresh tissue. The cone distribution was characterized by a steep increase in cone density centrally, which was elongated in the nasotemporal axis, and coincided approximately with the central increase in ganglion cell density (Stone, '65). Cone density in the area centralis peaked at 26,000–27,000/mm 2 , and fell to a plateau of 4000/mm 2 in the periphery and to less than 3000/mm 2 near the ora serrata. The density of rods was greater than the density of cones in all regions. The rods reached a maximum density of 460,000/mm 2 at an eccentricity of 10–15°, in a region which completely surrounded the central cone elevation. Centrally, the rod distribution was characterized by a sharp fall in density, reaching a low of 275,000/mm 2 at the point of peak cone density. Peripherally, there was a plateau of high rod density at above 400,000/mm 2 , out to about 30° temporal eccentricity. Beyond this plateau, rod density steadily fell, and reached a low of 250,000/mm 2 near the ora serrata. The rod/cone ratio reached a low value of 10.5–11.0 in the central region of maximum cone density. It then rose steeply to reach a plateau of about 65 in the periphery, while near the ora serrata there were 100 rods for each cone. The rods were arranged in well defined rows, and each rod was surrounded by six other rods. The cones were interspersed throughout the rod mosaic and were less regularly arranged than the rods. In fresh tissue, rod outer segment diameters ranged from 1.0 μ to 1.6 μ. rod diameters were smallest in the region of maximum rod density and increased, along with the decrease in rod density, both in the periphery and in the area centralis. Two thirds of the central decrease in rod density from the pericentral peak could not be accounted for by the increase in the percentage of space occupied by cones, and was consistent with a central increase in rod diameter. Receptor‐ganglion cell convergence was estimated by comparing receptor densities with the ganglion cell densities of Stone ('65). Cone‐ganglion cell convergence reached a minimum in the area centralis at the cone density peak, probably indicating the retinal region in which photopic acuity is at an optimum. Minimum rod‐ganglion cell convergence, as estimated from the rod/large ganglion cell ratio, reached a minimum at about 7.5° central to the peak of rod density, and at about the same eccentricity as the human scotopic acuity optimum.

Changes in extracellular Ca2+ concentration were directly measured in the rat cerebellum, using an ion-selective micropipette. Extracellular K+ was measured simultaneously with a second ion-selective micropipette. The potential reference barrels of the ion electrodes also provided fast field and slow potentials. During repetitive stimulation of the parallel fiber--Purkinje cell cerebellar circuit, extracellular Ca2+ fell to about 80% of base line concentration. During the spreading depression of Leão, extracellular Ca2+ fell to about 10% of base line; decreases of this magnitude also occurred during terminal anoxia. In all cases extracellular K+ increased substantially. These results show that extracellular Ca2+ is modulated during neuronal activity in the central nervous system and that under some conditions the Ca2+ change can be extreme. Given the well-established and antagonistic effects of reduce extracellular Ca2+ on axonal excitability and synaptic transmission, these results suggest that Ca2+ modulation in the brain cell microenvironment may be a significant parameter in the behavior of neuronal ensembles.