Interrelationships between Cellular Density, Mosaic Patterning, and Dendritic Coverage of VGluT3 Amacrine Cells

AUTHORS

Patrick W. Keeley, Mikayla C. Lebo, Jordan D. Vieler, Jason J. Kim, Ace J. St. John and Benjamin E. Reese

ABSTRACT

Amacrine cells of the retina are conspicuously variable in their morphologies, their population demographics, and their ensuing functions. Vesicular glutamate transporter 3 (VGluT3) amacrine cells are a recently characterized type of amacrine cell exhibiting local dendritic autonomy. The present analysis has examined three features of this VGluT3 population, including their density, local distribution, and dendritic spread, to discern the extent to which these are interrelated, using male and female mice. We first demonstrate that Bax-mediated cell death transforms the mosaic of VGluT3 cells from a random distribution into a regular mosaic. We subsequently examine the relationship between cell density and mosaic regularity across recombinant inbred strains of mice, finding that, although both traits vary across the strains, they exhibit minimal covariation. Other genetic determinants must therefore contribute independently to final cell number and to mosaic order. Using a conditional KO approach, we further demonstrate that Bax acts via the bipolar cell population, rather than cell-intrinsically, to control VGluT3 cell number. Finally, we consider the relationship between the dendritic arbors of single VGluT3 cells and the distribution of their homotypic neighbors. Dendritic field area was found to be independent of Voronoi domain area, while dendritic coverage of single cells was not conserved, simply increasing with the size of the dendritic field. Bax-KO retinas exhibited a threefold increase in dendritic coverage. Each cell, however, contributed less dendrites at each depth within the plexus, intermingling their processes with those of neighboring cells to approximate a constant volumetric density, yielding a uniformity in process coverage across the population.

The Relationship Between Macula Retinal Ganglion Cell Density and Visual Function in the Nonhuman Primate

AUTHORS

Kwame Antwi-Boasiako; Louvenia Carter-Dawson; Ronald Harwerth; Margaret Gondo; Nimesh Patel

ABSTRACT

Purpose: Loss of ganglion cell inner plexiform layer (GCIPL) and visual sensitivity in the macula region are known to occur at all stages of glaucoma. While both are dependent on the underlying retinal ganglion cells (RGCs), the relationship between structure and function is modest. We hypothesize that the imprecise relationship is due to a lack of direct correspondence between in vivo measures and RGC counts, as well as the relatively large stimulus size used by standard perimetry, which exceeds spatial summation.

Methods: The relationship between optical coherence tomography (OCT)–derived GCIPL thickness and corresponding inner cell density from retinal flat mounts was determined for four nonhuman primates with varying stages of neuropathy. Normative data for 10-2 threshold using Goldman size I to V stimuli were established for 10 animals, 4 of which were then followed longitudinally with OCT and perimetry. The relationship between GCIPL volume, which incorporated stimulus size after removal of residual thickness, and differential light sensitivity was determined for both experimental glaucoma and healthy eyes.

Results: Peak inner retinal cell density was 63,052 ± 9238 cells/mm2 in the healthy eye. Cell density was related to both GCIPL thickness and eccentricity (R2 = 0.74, P < .01). For all 10-2 eccentricities, size III stimuli were greater than the critical area (P < 0.01). Based on the structural and histologic relationship, the critical area corresponds to approximately 156 RGCs.

Conclusions: The relationship between cell density and GCIPL thickness is dependent on retinal eccentricity. For 10-2 perimetry, perimetric loss, especially at earlier stages of neuropathy, may best be detected using size II or smaller stimuli.

A Novel Resorbable Composite Material Containing Poly(ester-co-urethane) and Precipitated Calcium Carbonate Spherulites for Bone Augmentation—Development and Preclinical Pilot Trials

AUTHORS

Claudia Rode, Ralf Wyrwa, Juergen Weisser, Matthias Schnabelrauch, Marijan Vučak, Stefanie Grom, Frank Reinauer, Adrian Stetter, Karl Andreas Schlegel, Rainer Lutz

ABSTRACT

Polyurethanes have the potential to impart cell-relevant properties like excellent biocompatibility, high and interconnecting porosity and controlled degradability into biomaterials in a relatively simple way. In this context, a biodegradable composite material made of an isocyanate-terminated co-oligoester prepolymer and precipitated calcium carbonated spherulites (up to 60% w/w) was synthesized and investigated with regard to an application as bone substitute in dental and orthodontic application. After foaming the composite material, a predominantly interconnecting porous structure is obtained, which can be easily machined. The compressive strength of the foamed composites increases with raising calcium carbonate content and decreasing calcium carbonate particle size. When stored in an aqueous medium, there is a decrease in pressure stability of the composite, but this decrease is smaller the higher the proportion of the calcium carbonate component is. In vitro cytocompatibility studies of the foamed composites on MC3T3-E1 pre-osteoblasts revealed an excellent cytocompatibility. The in vitro degradation behaviour of foamed composite is characterised by a continuous loss of mass, which is slower with higher calcium carbonate contents. In a first pre-clinical pilot trial the foamed composite bone substitute material (fcm) was successfully evaluated in a model of vertical augmentation in an established animal model on the calvaria and on the lateral mandible of pigs.

Cell-autonomous role of Presenilin in age-dependent survival of cortical interneurons

AUTHORS

Jongkyun Kang, Jie Shen

ABSTRACT

Mutations in the PSEN1 and PSEN2 genes are the major cause of familial Alzheimer’s disease. Previous studies demonstrated that Presenilin (PS), the catalytic subunit of γ-secretase, is required for survival of excitatory neurons in the cerebral cortex during aging. However, the role of PS in inhibitory interneurons had not been explored.

To determine PS function in GABAergic neurons, we generated inhibitory neuron-specific PS conditional double knockout (IN-PS cDKO) mice, in which PS is selectively inactivated by Cre recombinase expressed under the control of the endogenous GAD2 promoter. We then performed behavioral, biochemical, and histological analyses to evaluate the consequences of selective PS inactivation in inhibitory neurons.

IN-PS cDKO mice exhibit earlier mortality and lower body weight despite normal food intake and basal activity. Western analysis of protein lysates from various brain sub-regions of IN-PS cDKO mice showed significant reduction of PS1 levels and dramatic accumulation of γ-secretase substrates. Interestingly, IN-PS cDKO mice develop age-dependent loss of GABAergic neurons, as shown by normal number of GAD67-immunoreactive interneurons in the cerebral cortex at 2–3 months of age but reduced number of cortical interneurons at 9 months. Moreover, age-dependent reduction of Parvalbumin- and Somatostatin-immunoreactive interneurons is more pronounced in the neocortex and hippocampus of IN-PS cDKO mice. Consistent with these findings, the number of apoptotic cells is elevated in the cerebral cortex of IN-PS cDKO mice, and the enhanced apoptosis is due to dramatic increases of apoptotic interneurons, whereas the number of apoptotic excitatory neurons is unaffected. Furthermore, progressive loss of interneurons in the cerebral cortex of IN-PS cDKO mice is accompanied with astrogliosis and microgliosis.

Our results together support a cell-autonomous role of PS in the survival of cortical interneurons during aging. Together with earlier studies, these findings demonstrate a universal, essential requirement of PS in the survival of both excitatory and inhibitory neurons during aging.

Ultrastructural effects of nerve growth factor and betamethasone on nerve regeneration after experimental nerve injury

AUTHORS

Leman Sencar, Mustafa Güven, Dilek Şaker, Tuğçe Sapmaz, Abdullah Tuli, and Sait Polat

ABSTRACT

Peripheral nerve injuries (PNI) are an important health problem in the world. In this study, the effects of nerve growth factor (NGF) and betamethasone on nerve regeneration after sciatic nerve crush injury were examined by footprint analysis, electron microscopic, histomorphometric, and biochemical methods. Fifty Wistar rats were divided into five groups as intact control, experimental control, NGF, betamethasone, and NGF+betamethasone combined treatment groups. After the injury, betamethasone was subcutaneously injected into the lesion area of the treatment groups three times during the first day. NGF was subcutaneously injected into the lesion area of treatment groups for 14 days. Footprint analysis was made on 7, 14, 21, 28, and 35 days and after 6 weeks, tissue samples were obtained from all groups. In the experimental control group, there were severe degenerative changes in most of the axons and myelin sheaths of the nerve fibers. Moreover, an increase of MDA levels and a decrease in SOD activities were found in this group. On the other hand, malondialdehyde (MDA) levels decreased, superoxide dismutase (SOD) activities increased and significant motor functional recovery were found in the combined treatment group. The number of axons, axon diameters, and myelin thickness were significantly greater in the combined treatment group when compared with experimental control and other treatment groups. It was thought that nerve regenerative effects of NGF and anti-inflammatory and/or anti-edematous effects of betamethasone could induce functional recovery in the combined treatment group. In conclusion, combined therapy of NGF and betamethasone may be an effective approach for the treatment of PNI.