Charles Straznicky

Morphological classification of retinal ganglion cells in adultXenopus laevis

SummaryRetrograde transport of horseradish peroxidase (HRP) was used to characterise the soma and dendritic arborization of retinal ganglion cells in adultXenopus laevis toad. HRP was administered to the cut end of the optic nerve and the morphological characteristics of HRP-filled ganglion cells were analysed in retinal wholemount preparations using computer assisted morphometry. Ganglion cells were classified according to their soma size, dendritic branching pattern, dendritic field and the number of shaft dendrites. Ganglion cells were divided into 3 major classes on the basis of soma sizes and extent of dendritic field: large (soma size, mean 258.04 μm2±52.03 SD; dendritic field size 0.104 mm2±0.23), medium size (126.7 μm2±37.01; 0.041 mm2±0.013) and small (87.3 μm2±22.69; 0.0061 mm2±0.0035). A more detailed analysis allowed 12 morphologically distinct subgroups to be identified (Types I–XII). Quantitative studies showed that large cells comprise about 1%, medium size about 8–9% and the small cells over 90% of total ganglion cell population. The number of large and medium size ganglion cells corresponded well with the number of myelinated optic fibres and the number of small neurons with the number of unmyelinated optic fibres in the optic nerve. Large ganglion cells were correlated with Class 4 and 5, medium size ganglion cells with Class 3 and small ganglion cells with Class 1 and 2 functionally characterized ganglion cells in the frog retina (Maturana et al. 1960). The retinal distribution of large ganglion cells appear to suggest certain similarities to mammalian alpha type ganglion cells.

Serotonin synthesis and accumulation by neurons of the anuran retina.

Serotonin-synthesizing and serotonin-accumulating neurons were studied in the retinas of Xenopus laevis and Bufo marinus. All previously identified cell types exhibiting serotonin-like immunoreactivity (SLI) were labeled by intravitreal injection of 5,7-dihydroxytryptamine (5,7-DHT). They included two amacrine cell types (large and small) in both species, and one bipolar cell type in Xenopus. Incubation of retinas in culture medium in the ambient light reduced SLI in amacrine cells and enhanced the labeling in bipolar cells. After incubation, some photoreceptor cell bodies and large numbers of outer segments also displayed SLI in both species. Incubation with the serotonin-uptake inhibitor, fluoxetine, reduced immunolabeling in bipolar cells and outer segments to the level in the untreated retinas. Both large SLI and 5,7-DHT-accumulating amacrine cells in Xenopus and Bufo were labeled with an antibody raised against phenylalanine hydroxylase (PH), which binds to tryptophan 5-hydroxylase, one of the synthesizing enzymes for serotonin. Small SLI and 5,7-DHT-accumulating amacrine cells in both species represented two populations, one with and the other without PH-like immunoreactivity (PH-LI). The anti-PH antibody failed to label any SLI or 5,7-DHT-accumulating bipolar cells in Xenopus. These observations indicate that all large and some small SLI amacrine cells in the retinas of Xenopus and Bufo synthesize serotonin, while other small SLI amacrine, bipolar and photoreceptor cell bodies, and outer segments only accumulate serotonin.

A neurofilament protein antibody selectively labels a large ganglion cell type in the human retina.

An antibody (SMI-32) raised against the non-phosphorylated form of the neurofilament protein triplet (NFP) revealed immunoreactivity in the soma and dendritic arborization of a group of large ganglion cells in the human retina. In addition, a population of smaller somas was also faintly labeled with this antibody in the ganglion cell layer. The completely stained cells amounted to 44,000 and were non-uniformly distributed across the retina with a peak density of 100 cells/mm2 in the retinal periphery. The soma sizes increased about two-fold and dendritic field sizes about 3-fold with retinal eccentricity. The immunoreactive dendrites branched in the vitread sublamina of the inner plexiform layer. The dendritic branching pattern of these cells indicated that they correspond to the previously described shrub cells. Antibodies against NFP and neuropeptide Y showed colocalization of these markers in all of the completely stained cells.

GABA-like immunoreactive neurons in the retina of Bufo marinus: evidence for the presence of GABA-containing ganglion cells

gamma-Aminobutyric acid (GABA)-like immunoreactive (IR) neurons in the retina of the cane toad Bufo marinus were revealed using immunohistochemistry on retinal wholemount preparation and sectioned material. GABA-IR neurons included horizontal, bipolar and amacrine cells in the inner nuclear layer and small to medium sized cells in the ganglion cell layer. A few IR axons were seen in the optic fiber layer of the retina. Following the injection of the carbocyanine dye, DiI into the optic tectum ganglion cells were retrogradely filled. A small population of DiI-filled ganglion cells (2.8%) was found to be GABA-IR. GABA-IR neurons in the ganglion cell layer without DiI label were considered to be displaced amacrine cells of which 45.3% were GABA positive. It is proposed that GABA-containing ganglion cells may form an inhibitory projection to visual centers of the anuran brain.

The localization of motoneuron pools innervating wing muscles in the chick

SummaryMotoneuron pools supplying principal muscles of the shoulder girdle and wing were localized in the chicks 2–7 days post hatching with the use of retrograde axonic transport of horse radish peroxidase (HRP). HRP was injected into selected muscles and the animals sacrificed after 24 h survival. Labelled motoneuron pools representing individual muscles were found to be clustered in longitudinal columns along the brachial spinal cord segments 13–16. Muscles derived from the dorsal muscle mass were innervated by motoneurons located in the ventro-lateral portion of the horn, while those originating from the ventral muscle mass received innervation from neurons occupying the dorsomedial portion within the ventral horn of the spinal cord. The rostrocaudal extent of the motoneuron pools could be correlated with the proximodistal position of wing muscles.The observed orderly topographical relationships between clusters of motoneurons of the brachial spinal cord and the muscles they innervate will be used as baseline data for experiments where the limb innervation is perturbed.

Morphological characterization of substance P-like immunoreactive amacrine cells in the anuran retina.

Using substance P immunohistochemistry it was possible to demonstrate a class of morphologically homogeneous group of neurons in the inner nuclear layer (INL) of the retina of two anuran species: Xenopus laevis and Bufo marinus. The number of cells with substance P-like immunoreactivity (SP-LI) was about 250 and 800 in juvenile and 600 and 2500 in adult Xenopus and Bufo, respectively, SP-LI cells had a small soma with one primary dendrite having up to four slender branches, located in the vitreal sublamina of the inner plexiform layer (IPL). Mean dendritic field sizes were 0.12 and 0.30 mm2 in juvenile and 0.29 and 0.65 mm2 in adult Xenopus and Bufo, respectively. The density of SP-LI cells was 40/mm2 in juvenile and 24/mm2 in adult Xenopus compared with 20/mm2 in juvenile and 13/mm2 in adult Bufo. Nearest neighbour distance measurements indicated that SP-LI cells were randomly distributed across the entire retina in both species. The location and the morphology of SP-LI cells indicated that they correspond to a subclass of wide-field amacrine cells, similar to types 20 and 21 described by Golgi techniques in the cat.

Neuropeptide Y-like immunoreactive amacrine cells in the retina ofBufo marinus

Neuropeptide Y-like immunoreactive (NPY-LI) amacrine cells of the Bufo marinus retina were morphologically characterized, and their retinal distribution was established using immunohistochemistry on retinal wholemount preparations and sectioned material. The somas of NPY-LI amacrine cells were situated in the innermost part of the inner nuclear layer and their dendrites branched primarily in the scleral sublamina of the inner plexiform layer. A subgroup of the NPY-LI cells had dendrites in both the scleral and vitreal sublamina. All immunoreactive cells had large dendritic fields (average 0.5 mm2) that resulted in a high dendritic overlap across the retina. NPY-LI amacrine cells were evenly distributed across the retina, with an average density of 30 cells/mm2, although higher densities were observed at regions adjacent to the ciliary margin. The dendritic field size of the NPY-LI cells, together with the previously characterized substance P-like immunoreactive (SP-LI) amacrine cells, indicates that they belong to the class of wide-field amacrine cells. However, unlike the SP-LI neurons whose dendrites branch in the vitreal sublamina of the inner plexiform layer, the dendrites of the majority of the NPY-LI neurons branch in the scleral sublamina.

Immunocytochemical localization of parvalbumin- and neurofilament triplet protein immunoreactivity in the cat retina : colocalization in a subpopulation of AII amacrine cells

Using antibodies against parvalbumin and neurofilament triplet protein, colocalization of these two neuronal markers was revealed in all of type A horizontal cells and alpha ganglion cells and in a small number of AII amacrine cells of the cat retina. Besides the double-labeled neurons, parvalbumin alone was present in type B horizontal cells, in small numbers of starburst- and A13-like amacrine cells and in the somata of unidentified ganglion cells. The processes of the double- or single-labeled amacrine cells did not have a continuous retinal cover. Although the parvalbumin- and neurofilament-immunolabeled amacrine cells belonged to groups of neurons with well-defined cell morphologies, their neurochemical features differed from other AII, starburst and A13 amacrine cells. The presence of these cells may be due to an accidental expression of an unusual combination of neurochemical features during retinal development. It is also possible that these cells support the functioning of ganglion cells with rarely occurring complex receptive fields.

The effect of nerve growth factor on developing primary sensory neurons of the trigeminal nerve in chick embryos

SummaryThe response to nerve growth factor (NGF) of two sensory neuron populations of the trigeminal nerve was studied in chick embryos. NGF promoted neuronal survival and cellular hypertrophy in the Gasserian ganglia with minimal effect on the neuron population of the mesencephalic trigeminal nucleus. NGF induced prolific neurite outgrowth from cultured Gasserian ganglia, in contrast, cultured mesencephalic trigeminal neurons remained refractory to NGF treatment.The apparent lack of response of mesencephalic trigeminal neurons to NGF may be explained either by their derivation from placodal material rather than from the neural crest, or their lost sensitivity to NGF due to interaction with the local environment in the central nervous system.

Neuropeptide Y-like immunoreactivity in neurons of the human retina

The distribution of neuropeptide Y-like immunoreactivity (NPY-LI) was investigated in wholemounts and in transverse sections of the human retina. NPY-LI was localized to the soma and axonal processes of large ganglion cells (GCs) and to the soma and dendritic arborization of amacrine cells (ACs). NPY-LI GCs were unevenly distributed across the retina, the highest density of 875 cells/mm2 was found in the fovea centralis and the lowest density of 15 cells/mm2 in the peripheral retina. The total number of NPY-LI GCs in the retina was estimated to be about 85,000. The soma sizes of NPY-LI GCs increased from 116 microns 2 +/- 23 (s.d.) in the retinal centre to 251 microns 2 +/- 57 in the retinal periphery. The soma size of NPY-LI ACs was in the range of 40 and 50 microns 2. In transverse sections NPY-LI was seen to be localized to the optic fibre layer, to the somata of GCs, to the scleral sublamina of the inner plexiform layer (AC dendrites) and to the innermost part of the inner nuclear layer (somata of ACs). The gradients of soma sizes and retinal distribution of NPY-LI GCs were taken as an indication that they correspond to the class of large to very large GCs, previously identified in the human retina by Golgi impregnation.