6 research outputs found
Real-Time Imaging of Rabbit Retina with Retinal Degeneration by Using Spectral-Domain Optical Coherence Tomography
Background: Recently, a transgenic rabbit with rhodopsin Pro 347 Leu mutation was generated as a model of retinitis pigmentosa (RP), which is characterized by a gradual loss of vision due to photoreceptor degeneration. The purpose of the current study is to noninvasively visualize and assess time-dependent changes in the retinal structures of a rabbit model of retinal degeneration by using speckle noise-reduced spectral-domain optical coherence tomography (SD-OCT). Methodology/Principal Findings: Wild type (WT) and RP rabbits (aged 4–20 weeks) were investigated using SD-OCT. The total retinal thickness in RP rabbits decreased with age. The thickness of the outer nuclear layer (ONL) and between the external limiting membrane and Bruch’s membrane (ELM–BM) were reduced in RP rabbits around the visual streak, compared to WT rabbits even at 4 weeks of age, and the differences increased with age. However, inner nuclear layer (INL) thickness in RP rabbits did not differ from that of WT during the observation period. The ganglion cell complex (GCC) thickness in RP rabbits increased near the optic nerve head but not around the visual streak in the later stages of the observation period. Hyper-reflective change was widely observed in the inner segments (IS) and outer segments (OS) of the photoreceptors in the OCT images of RP rabbits. Ultrastructural findings in RP retinas included the appearance of small rhodopsin-containing vesicles scattered in the extracellular space around the photoreceptors
Local connections of excitatory neurons to corticothalamic neurons in the rat barrel cortex.
Corticothalamic projection neurons in the cerebral cortex constitute an important component of the thalamocortical reciprocal circuit, an essential input/output organization for cortical information processing. However, the spatial organization of local excitatory connections to corticothalamic neurons is only partially understood. In the present study, we first developed an adenovirus vector expressing somatodendritic membrane-targeted green fluorescent protein. After injection of the adenovirus vector into the ventrobasal thalamic complex, a band of layer (L) 6 corticothalamic neurons in the rat barrel cortex were retrogradely labeled. In addition to their cell bodies, fine dendritic spines of corticothalamic neurons were well visualized without the labeling of their axon collaterals or thalamocortical axons. In cortical slices containing retrogradely labeled L6 corticothalamic neurons, we intracellularly stained single pyramidal/spiny neurons of L2-6. We examined the spatial distribution of contact sites between the local axon collaterals of each pyramidal neuron and the dendrites of corticothalamic neurons. We found that corticothalamic neurons received strong and focused connections from L4 neurons just above them, and that the most numerous nearby and distant sources of local excitatory connections to corticothalamic neurons were corticothalamic neurons themselves and L6 putative corticocortical neurons, respectively. These results suggest that L4 neurons may serve as an important source of local excitatory inputs in shaping the cortical modulation of thalamic activity
Branched chain amino acids attenuate major pathologies in mouse models of retinal degeneration and glaucoma
Retinal neuronal cell death underlies many incurable eye diseases such as retinitis pigmentosa (RP) and glaucoma, and causes adult blindness. We have shown that maintenance of ATP levels via inhibiting ATP consumption is a promising strategy for preventing neuronal cell death. Here, we show that branched chain amino acids (BCAAs) are able to increase ATP production by enhancing glycolysis. In cell culture, supplementation of the culture media with BCAAs, but not glucose alone, enhanced cellular ATP levels, which was canceled by a glycolysis inhibitor. Administration of BCAAs to RP mouse models, rd10 and rd12, significantly attenuated photoreceptor cell death morphologically and functionally, even when administration was started at later stages. Administration of BCAAs in a glaucoma mouse model also showed significant attenuation of retinal ganglion cell death. These results suggest that administration of BCAAs could contribute to a comprehensive therapeutic strategy for retinal neurodegenerative diseases such as RP and glaucoma
SD-OCT images of WT and Retinitis Pigmentosa (RP) rabbit retinas and histology of the visual streak in an RP rabbit.
<p>(A) A fundus infrared image of a WT rabbit retina, including optic nerve head (ONH) and visual streak. The area between dotted lines is the visual streak. (B) A vertical SD-OCT image along the green arrow in panel A, which passes through the center of the ONH. On this vertical image, the scleral ring was regarded as the lower margin of the ONH. (C) A magnified OCT image of the area enclosed by the blue square in panel B, which includes the visual streak. (D) A fundus infrared image of a RP rabbit retina, including the ONH and visual streak. (E) A vertical SD-OCT image of a 20-week-old RP rabbit along the green arrow in panel D. (F) A magnified OCT image of the area enclosed by the blue square in panel E. The 2.2 mm width of this OCT section was vertically cut between 1.8 mm and 4.0 mm ventral to the inferior edge of the ONH. A dotted arrow indicates the region of the visual streak. (G) Hematoxylin-Eosin staining of a retinal section corresponding to the area in the OCT image in F. Scale Bar = 200 µm (B, E), 100 µm (C, F), and 50 µm (G). RNFL, retinal nerve fiber layer; GCL, ganglion cell layer; IPL, inner plexiform layer; GCC, ganglion cell complex; INL, inner nuclear layer; OPL, outer plexiform layer; ONL, outer nuclear layer; ELM, external limiting membrane; IS, inner segments of photoreceptors; OS, outer segments of photoreceptors; IS/OS, junctions between IS and OS; RPE, retinal pigment epithelium; and BM, Bruch's membrane.</p
Time-dependent changes in morphological features of the retina and visual function in the RP rabbits.
<p>(A) SD-OCT images beneath the visual streak in an RP rabbit at 4, 6, 10, and 20 weeks and in a 20-week-old WT rabbit. The total retinal and ONL thickness in the RP rabbits decreased with age. The IS and OS were highly reflective in the RP rabbits compared with the WT rabbits. ONL, outer nuclear layer; and OS, outer segments of photoreceptors. (B) Hematoxylin-eosin staining of retinas in 4-, 6-, 10-, and 20-week-old RP and 20-week-old WT rabbits. The ONL in RP rabbits thinned with age. In 20-week-old RP rabbits, only 1–2 layers of nuclei were detected in the ONL. (C) Representative scotopic electroretinograms of 4-, 6-, 10-, and 20-week-old RP and 20-week-old WT rabbits. (D) The a-wave amplitude of the mixed rod and cone response. The amplitude was smaller in the RP rabbits than in the WT rabbits. The differences between the WT and RP rabbits were significant at all study points between 4 and 20 weeks. *<i>P</i><0.05, ***<i>P</i><0.001 (unpaired <i>t</i>-test). Scale Bar = 100 µm in A, and 50 µm in B. ONL, outer nuclear layer; ELM, external limiting membrane; IS/OS, junctions between inner segment (IS) and outer segment (OS); RPE, retinal pigment epithelium; and BM, Bruch's membrane.</p
Ultrastructure of photoreceptors in WT and RP rabbits.
<p>(A, B) Ultrastructure of photoreceptors in 4-week-old WT rabbits. The inner (IS) and outer segments (OS) of the photoreceptors were regular and dense. There are no vesicles in the extracellular spaces (*). (C–E) Ultrastructure of the photoreceptors in the 4-week-old RP rabbits. The IS and OS were less organized than those in the WT rabbits. In the magnified image (D), the RP rabbit retina showing many small vesicles (arrowheads) accumulated in the extracellular spaces (indicated with * in panel C). The vesicles appeared to be cleaved from the IS into the extracellular space around the photoreceptors (arrows in panel E). (F) Ultrastructural immunohistochemistry by using an anti-rhodopsin antibody. The small vesicles (disintermediated arrowheads) in the extracellular spaces around the photoreceptors exhibit black dots indicating the presence of rhodopsin.</p