Evaluation of the osteogenic and chondrogenic differentiation capacities of equine adipose tissue-derived mesenchymal stem cells.

OBJECTIVE: To evaluate the proliferative behavior, telomere length, immunophenotype, and differentiation capacity of equine adipose tissue-derived mesenchymal stem cells (AT-MSCs). ANIMALS: 6 adult racing horses treated for articular Injury but otherwise healthy. PROCEDURES: AT-MSCs were Isolated from horses and expanded In Dulbecco modified Eagle medium enriched with fetal bovine serum and antimicrobials. Expression of cell surface antigens and telomere length were Investigated via flow cytometry Differentiation of MSCs Into chondrocytes, osteoblasts, and adipocytes was Induced In vitro by specific stimuli and was evaluated by analyzing marker genes with quantitative reverse transcriptase PCR assays and immunocytochemical and cytologie evaluations. RESULTS: Equine MSCs could be cultured up to the fifth passage before signs of senescence, apoptosis, and detachment Indicated cellular exhaustion. However, the AT-MSCs from 2 of 6 horses survived to later passages with Increased doubling rates and telomere lengths. The cells had a typical phenotype, with expression of CD14, CD73, CD90, CD105, CD140b, and CD164 antigens and a lack of CD34 and CD45 antigens. The cells also had a strong potential to differentiate Into osteoblasts, as characterized by Intense von Kossa and alizarin red staining as well as high Induction of osteopontin. Chondrogenic differentiation was detected via Alelan blue staining and expression of aggrecan and type II collagen Adipogenesis was Induced in AT-MSCs by supplementation of differentiation media with rabbit serum. CONCLUSIONS AND CLINICAL RELEVANCE: Equine AT-MSCs representa suitable cellular source for regenerative treatment of bone or cartilage defects, particularly when expanded In vitro for only a few passages.

[De-novo generation of vascularized tissue using different configurations of vascular pedicles in perforated and closed chambers]. / De-novo Generierung von vaskularisiertem Gewebe mittels unterschiedlicher Gefässstielkonfigurationen in perforierten und geschlossenen Wachstumskammern.

Growing three-dimensional tissue within a chamber requires vigorous angiogenesis initiated by, for example, an arteriovenous fistula or a ligated vascular pedicle. Growth may also be enhanced by contact with the external environment. In this study tissue growth in a rat model, vascularized via an arteriovenous loop (AV Loop) or ligated pedicle, was compared in chambers that were either closed or perforated. Chambers were harvested at 4 weeks and tissue volume and histology compared. In perforated chambers, more tissue were generated using the ligated pedicle (0.75 ml+/-0.04) than the AV Loop (0.59 ml+/-0.01). Perforated chambers generated larger volumes of tissue than closed chambers because they encouraged tissue ingrowth through the perforations. Both vessel configurations supported tissue growth but, interestingly, the ligated pedicle resulted in significantly more tissue in the perforated chambers.

ATP-induced non-neuronal cell permeabilization in the rat inner retina.

The P2X7 subtype holds a special position among P2X receptors because of its ability to act both as a classical, ligand-gated ion channel, and as a permeabilization pore that can induce cell death under prolonged activation by ATP. We have shown previously that, in rat retina, P2X7 receptors are located in the inner nuclear layer and ganglion cell layer (GCL). The present study was aimed at finding whether retinal P2X7 receptors can act as a mediator of cell permeabilization and, if so, at identifying the cellular target(s) of this effect. As an indicator of cell permeabilization, we used the fluorescent dye YO-PRO-1 (molecular weight, 375 Da), which enters cells only through large pores like those opened by prolonged or sustained stimulation of P2X(7) receptors and binds to DNA, providing a stable labeling of the activated cells. Different agonists for P2 receptors were tested for their ability to cause cell permeabilization in flat-mounted rat retinas. Among them, only high concentrations of ATP (500 microM) and BzATP (2',3'-O-(4-benzoyl-benzoyl)-ATP triethylammonium) (100 microM) were able to induce accumulation of YO-PRO-1 in the GCL and in the nerve fiber layer, suggesting that different cell types were responding to P2X7 stimulation. This effect was blocked by the P2 antagonists suramin and PPADS (pyridoxal-phosphate-6-azophenyl-2',4'-disulfonic acid) and by the P2X7-selective inhibitor Brilliant Blue G. To identify the retinal cell types affected by ATP-induced permeabilization, we used in vivo labeling techniques. Our data clearly reveal that prolonged stimulation of P2X7 receptors elicits permeabilization exclusively in microglial cells but not in neurons of the inner retina.

Rat retinal dopaminergic neurons: differential maturation of somatodendritic and axonal compartments.

We examined developmental changes in dopaminergic (DA) neurons of rat pups between postnatal (P) days 3 and 21. DA cell bodies and dendrites grew progressively between P3-15. Voltage-sensitive sodium channels were present in axons at P11, but the ring-like DA axon terminals appeared only during the third postnatal week. The density of ring terminals increased markedly between P15 and P21. The vesicular monoamine transporter (VMAT2) was absent before P13 and became concentrated in DA ring terminals after P17. A steady increase in VMAT2-containing rings around AII amacrine cells occurred during the third postnatal week. The presynaptic membrane protein SNAP-25 colocalized with DA terminals, but several other presynaptic proteins tested, including synaptotagmin I, synapsin, bassoon, syntaxin, and synaptogyrin, appeared not to be associated with DA neurons. Our study shows that the somatodendritic compartment of DA neurons matures before the DA axon terminals do. Maturation of DA axons during the third postnatal week corresponds to the period of onset of visual function.

Identification of P2Y receptor subtypes in human muller glial cells by physiology, single cell RT-PCR, and immunohistochemistry.

PURPOSE: Retinal Müller glial cells are known to express metabotropic P2Y receptors. The present study was conducted to identify certain subtypes of P2Y receptors in human Müller cells. METHODS: The patch-clamp technique was used to measure increases of Ca(2+)-dependent K+ currents mediated by the activation of P2Y receptors in freshly isolated human Müller cells. Several P2 agonists were used. Subsequently, the cells were harvested into the patch pipette and a single cell RT-PCR was performed. Moreover, retinal tissue from organ donors was used for immunohistochemistry. RESULTS: The electrophysiological data were consistent with the expression of P2Y1, P2Y2, P2Y4, and P2Y6 receptor subtypes. RT-PCR revealed that mRNA for all these subtypes was present in Müller cells. However, the incidence of P2Y2 receptor mRNA was significantly lower than that of the other subtypes. Immunoreactivity for all four subtypes was found in retinal tissue, partly colocalized with immunoreactivity for vimentin. CONCLUSIONS: The presented data obtained by different techniques revealed that human Müller cells express P2Y1, P2Y2, P2Y4, and P2Y6 receptors. The specific roles of these receptor subtypes in retinal physiology and/or pathophysiology remain to be investigated in future studies.

Developmental plasticity of NMDA receptor function in the retina and the influence of light.

Despite the early expression of NMDA receptors (NMDARs) in the retina, not much is known about their regulation and involvement in plasticity processes during retinal development and synapse formation. Here we report that NMDAR function in the inner retina is developmentally regulated and controlled by ambient light condition. A prominent down-regulation after eye opening of NMDAR function was observed in rat retinal ganglion cells (RGCs), which was prevented by dark rearing the animals for 1 month but was again induced by subsequent light exposure. As shown by molecular analysis of single RGCs, alterations in the subunit composition of NMDAR did not account for the light-dependent regulation of NMDAR function. Immunocytochemistry showed no differences in the NMDAR protein expression pattern between normal and dark-reared animals. In conclusion, our data clearly demonstrate that NMDAR function is modulated during periods of retinal plasticity independent of structural alterations in its subunit composition and thus different from mechanisms observed in higher visual centers.

Dopaminergic Modulation of Transient Neurite Outgrowth from Horizontal Cells of the Fish Retina is not Mediated by cAMP.

Horizontal cell dendrites invaginating the cone pedicles in the fish retina exhibit a marked light dependent plasticity in the morphology of their synaptic connections. Upon light adaptation of the retina, numerous spinules are formed which disappear during dark adaptation. This process is paralleled by a strengthening and weakening, respectively, of the horizontal cell's inhibitory output. The formation of spinules during light adaptation requires dopaminergic activity as it does not occur in dopamine-depleted retinas, but can be partially induced in depleted retinas by the exogenous administration of dopamine. Although horizontal cells do have D1 receptors the action of dopamine is not coupled to a stimulation of cAMP. An increase of intracellular cAMP either by injection of a cAMP analogue or by metabolic interference does not result in any spinule formation. The data suggest that dopamine must act through a cAMP independent intracellular mechanism.

Brain-derived neurotrophic factor regulates expression of vasoactive intestinal polypeptide in retinal amacrine cells.

Brain-derived neurotrophic-factor (BDNF) is expressed in the retina and controls the development of subtypes of amacrine cells. In the present study we investigated the effects of BDNF on amacrine cells expressing vasoactive intestinal polypeptide (VIP). Rats received three intraocular injections of BDNF on postnatal days (P) 16, 18, and 20. The animals were sacrificed on P22, P40, P60, P80, and P120, and VIP expression in their retinas was detected by immunohistochemistry (P22, P40) and by radioimmunoassay (RIA; P22, P40, P60, P80, P120) to assess the time course of BDNF effects on VIP. A significant increase in the density of VIP-positive amacrine cells was detected in BDNF-treated retinas, and VIP concentration was up-regulated by 150% both at P22 and at P40 with respect to untreated controls. VIP concentration then slowly declined in the treated retinas over a period of 3 months; however, a statistically significant increase of 50% was still detectable on P120. The impact of endogenous BDNF on the regulation of VIP expression in the retina was analyzed in mice homozygous for a targeted deletion of the BDNF gene locus (bdnf-/-). VIP immunohistochemistry revealed a marked reduction of VIP-positive amacrine cells and of VIP-immunopositive processes in the inner plexiform layer of the BDNF knockout mice. Mice lacking BDNF expressed only 5% of the VIP protein in their retinas compared with the retinas of wild-type mice as measured by RIA. Our data show that BDNF is a major regulator of VIP expression in retinal amacrine cells and exerts long-lasting effects on VIP content.

Double cone dystrophy and RPE degeneration in the retina of the zebrafish gnn mutant.

PURPOSE: To characterize morphologic alterations in the retina of the visual mutant zebrafish gantenbein (gnn) and to examine whether these alterations correlate with those present in human hereditary eye diseases. METHODS: The gnn mutant was isolated by behavioral and macroscopic screening. Retinas of gnn zebrafish larvae were examined at different developmental stages from 2 to 9 days postfertilization (dpf) by standard histologic staining techniques and by immunocytochemistry. Ultrastructural alterations were examined by electron microscopy. The genetic map position of the induced mutation was identified by mapping with two candidate primer pairs on single larvae. RESULTS: The gnn mutant exhibited shortened outer photoreceptor segments and altered RPE morphology. In the photoreceptor layer of the mutant, the total number of lectin-labeled cones was reduced in all developmental stages from 2 to 7 dpf, whereas the amount of rhodopsin-positive cells remained at the wild-type (WT) level. Labeling with zebrafish opsin antibodies revealed dystrophic red cones at 5 dpf, whereas the morphology of all other cone types was largely unaffected. Electron microscopy unveiled electron-dense deposits between the discs of the double cone outer segments. In addition, the onset of progressive RPE degeneration was observed at this stage of development. At later stages, all cone types and the RPE became degenerative. The morphology of distinct second-order neurons remained largely unaffected by the mutation. The gnn mutation was located approximately 4.3 cM from the simple sequence length polymorphism (SSLP) marker Z15453 on linkage group 16. CONCLUSIONS: In gnn mutant zebrafish, cones, and especially red cones, are dystrophic in early retinal development. Subsequent to this cone dystrophy, the RPE becomes dysfunctional and starts to degenerate in later stages of development. Thus, the early developmental morphology of gnn exhibits similarities to cone dystrophies most commonly seen in age-related macular degeneration (AMD) among humans, whereas the later stages of degeneration in gnn resemble RPE alterations in retinitis pigmentosa (RP) in humans. The gnn zebrafish mutant may therefore be a useful model for examining the possible interplay and connection between cone dystrophy and RPE degeneration.

Expression of P2Y1, P2Y2, P2Y4, and P2Y6 receptor subtypes in the rat retina.

PURPOSE: To elucidate the expression pattern of different types of metabotropic P2Y receptors in the adult rat retina. METHODS: Qualitative RT-PCR was used to investigate the expression profile of different P2Y receptor subtypes (P2Y1, P2Y2, P2Y4, and P2Y6), and in situ hybridization studies were performed to show their cellular localization within the retina. Immunohistochemical staining was used to detect the corresponding P2Y proteins (P2Y1, P2Y2, and P2Y4) and their cellular localization. Southern blot analysis and sequencing verified the identity of the P2Y PCR products. RESULTS: RT-PCR revealed the presence of P2Y1, -2, -4, and -6 mRNA in the neural retina and the retinal pigment epithelium (RPE) and choroid. In situ hybridization showed labeling in the retinal ganglion cell layer for all four P2Y receptor subtypes, although the intensity varied. In addition, staining for P2Y1, -4, and -6 mRNA was shown in the inner nuclear layer, but was absent for the P2Y2 receptor subtype. Immunohistochemistry showed intense staining for P2Y1, -2, and -4 in the ganglion cell layer and the outer plexiform layer. There was also a specific subtype staining in the inner plexiform layer (P2Y2, -4), the inner (P2Y1, -4) and outer (P2Y1) nuclear layers and the inner segments of the photoreceptors (P2Y1, -2). discussion. The data suggest that extracellular nucleotides may play complex roles as autocrine-paracrine mediators and may have neuromodulatory effects in the retina through metabotropic P2Y receptors.

OPA1, the disease gene for autosomal dominant optic atrophy, is specifically expressed in ganglion cells and intrinsic neurons of the retina.

PURPOSE: Autosomal dominant optic atrophy is a hereditary disorder characterized by progressive loss of vision and caused by mutations in a dynamin-related gene, OPA1, which translates into a protein with a mitochondrial leader sequence. In this study the OPA1 gene and its protein were localized in the rat and mouse retina, and its rat orthologue, rOpa1, was identified. METHODS: The rOpa1 cDNA was isolated by using reverse transcribed cDNA from total RNA obtained from a rat retinal ganglion cell line. The spatial and temporal expression patterns of OPA1 and its gene product were investigated by RNA in situ hybridization and immunohistochemistry in mouse and rat retinas. To characterize further the OPA1-positive neurons, retinal ganglion cells were retrogradely labeled by an immunogold fluorescent tracer or double labeled with OPA1 and choline acetyltransferase or calbindin antibodies. RESULTS: Protein sequence alignment revealed a 96% identity between rat and human OPA1 mRNA. OPA1 expression was found as early as postnatal day 3 in the developing rodent retina. In the mature retina, the OPA1 gene and its protein were found not only in retinal ganglion cells, but also in starburst amacrine cells and horizontal cells, both of which are involved in lateral signal processing within the retina. However, OPA1 was absent from mitochondria rich nerve fibers and photoreceptor indicating a specific role for OPA1 in signal processing rather than in the requirement of mitochondrial energy supply in the retina. CONCLUSIONS: The data suggest an important and specific function of the OPA1 protein, not only in the optic nerve forming ganglion cells but also in the intrinsic signal processing of the inner retina.

Retinal colocalization and in vitro interaction of the glutamate transporter EAAT3 and the serum- and glucocorticoid-inducible kinase SGK1 [correction].

PURPOSE: The serum- and glucocorticoid-inducible kinase SGK1 regulates several epithelial channels and transporters, the related protein kinase B (PKB) regulates glucose transport. SGK1 is expressed in the brain and could thus regulate glial and/or neuronal transport processes. The present study explores whether SGK1 is expressed in the retina and whether it regulates EAAT3, a Na(+)-coupled glutamate transporter. EAAT3 is expressed in retinal ganglion cells and accomplishes the clearance of glutamate from synaptic clefts. METHODS: Immunohistochemistry was performed to test for retinal SGK1 expression. For functional analysis, cRNA encoding EAAT3 was injected into Xenopus oocytes with or without additional injection of wild-type SGK1, constitutively active (S422D)SGK1, inactive (K127N)SGK1, and/or constitutively active (T308D,S473D)PKB. Glutamate induced current (I(GLU)) was taken as a measure for transport. RESULTS: SGK1 is indeed expressed in several retinal cells including retinal ganglion cells where it is colocalized with EAAT3. In EAAT3-expressing Xenopus oocytes, glutamate-induced current was stimulated by coexpression of wild-type SGK1, constitutively active (S422D)SGK1, and constitutively active (T308D,S473D)PKB, but not by inactive (K127N)SGK1. CONCLUSIONS: SGK1 and EAAT3 are coexpressed in retinal neurons, and SGK1 serves to stimulate EAAT3. This function is shared by protein kinase B (PKB). The experiments reveal a novel mechanism regulating EAAT3, which may be essential for the function of the retinal ganglion cells.

Isolation of the mouse nyctalopin gene nyx and expression studies in mouse and rat retina.

PURPOSE: It has been shown recently that mutations in NYX (nyctalopin on chromosome X), encoding a novel protein associated with the leucine-rich repeat (LRR) protein superfamily, are responsible for the complete form of X-linked congenital stationary night blindness (CSNB1). This study describes the isolation and molecular characterization of the mouse orthologue Nyx and its expression pattern in the retina. METHODS: Nyx was isolated by conventional DNA library screening and polymerase chain reaction (PCR)-based approaches. Gene expression in different mouse tissues was studied by RT-PCR. Subsequently, the expression pattern of Nyx and its gene product in mouse and rat retinas was investigated by RNA in situ hybridization and immunohistochemistry with Nyx-specific antibodies. RESULTS: The Nyx gene encodes a protein of 476 amino acids that contain 11 consecutive LRR motifs flanked by amino- and carboxyl-terminal cysteine-rich LRRs. At the amino acid level, Nyx is highly homologous to its human orthologue (86% identity). The gene is expressed in the eye but also, at lower levels, in brain, lung, spleen, and testis. Nyx expression was found during all stages of postnatal retinal development and was confined to cells of the inner nuclear layer and the ganglion cell layer in adult mouse and rat retinas. CONCLUSIONS: These data suggest an important function of the Nyx protein in the inner retina and provide evidence that CSNB1 is based on a defect in the inner retinal circuitry.

N-methyl-D-aspartate receptor subunit NR3A in the retina: developmental expression, cellular localization, and functional aspects.

PURPOSE: Recently, a novel N-methyl-D-aspartate receptor (NMDAR) subunit, NR3A, has been discovered in the brain and shown to decrease NMDAR activity by modulating the calcium permeability of the receptor channel. The insertion of NR3A within the NMDAR complex may thus alter NMDAR properties and play a crucial role during processes of neuronal development and degeneration. The present study is the first to investigate the expression and cellular localization of NR3A on the protein level in the retina and to elucidate its putative functional roles within the retinal circuitry. METHODS: The expression of NR3A in the retina was analyzed by reverse transcription-polymerase chain reaction (RT-PCR), immunohistochemistry, and Western blot analysis. Functional aspects of NR3A in the retina were addressed by measuring the NMDA-induced increase in intracellular calcium, [Ca(2+)](i), in retinal cells prepared from wild-type (NR3A(+/+)) and NR3A knockout (NR3A(+/-), and NR3A(-/-)) mice. RESULTS: NR3A protein expression was initially observed in the first postnatal week and was predominantly localized to cell bodies in the ganglion cell layer. In older animals, two bands of NR3A immunoreactivity were additionally observed in the inner plexiform layer. NMDA-evoked [Ca(2+)](i) responses were found to be significantly greater in retinal cells in NR3A(-/-) mice than in wild-type retinas. CONCLUSIONS: The data indicate that NR3A is specifically expressed in the inner retina and may modulate NMDAR-mediated calcium influx and thus [Ca(2+)](i) levels in retinal ganglion cells and amacrine cells.

A mouse model for Sorsby fundus dystrophy.

PURPOSE: Sorsby fundus dystrophy (SFD) is a rare, late-onset macular dystrophy caused by mutations in the tissue inhibitor of metalloproteinases-3 (TIMP3) gene. The known mutations introduce potentially unpaired cysteine residues in the C terminus of the protein and result in the formation of higher-molecular-weight protein complexes of as yet unknown composition and functional consequences in the pathologic course of SFD. To facilitate in vivo investigation of mutant TIMP3, the authors generated a knock-in mouse carrying a disease-related Ser156Cys mutation in the orthologous murine Timp3 gene. METHODS: Site-directed mutagenesis and homologous recombination in embryonic stem (ES) cells was used to generate mutant ES cells carrying the Timp3(S156C) allele. Chimeric animals were obtained, of which two displayed germline transmission of the mutated allele. Molecular genetic, biochemical, electron microscopic, and electrodiagnostic techniques were used for characterization. RESULTS: At 8 months of age, knock-in mice showed abnormalities in the inner aspect of Bruch's membrane and in the organization of the adjacent basal microvilli of the retinal pigment epithelium (RPE). Changes resembling those in the mutant animals were also present to some extent in normal littermates, but only at an advanced age of 30 months. Long-term electrodiagnostic recordings indicated normal retinal function throughout life. The biochemical characteristics of the mutant protein appear similar in humans and knock-in mice, suggesting common molecular pathways in the two species. The localization of the mutant protein in the eye is normal, although there is evidence of increased Timp3 levels in Bruch's membrane of mutant animals. CONCLUSIONS: The knock-in mice display early features of age-related changes in Bruch's membrane and the RPE that may represent the primary clinical manifestations of SFD. In addition, our immunolabeling studies and biochemical data support a model proposing that site-specific excess rather than absence or deficiency of functional Timp3 may be the primary consequence of the known Timp3 mutations.

Distribution of metabotropic P2Y receptors in the rat retina: a single-cell RT-PCR study.

P2Y receptors are metabotropic G-protein linked purinergic receptors, which are especially widespread in the central nervous system. The purpose of the present study was to determine the distribution patterns of P2Y receptors in distinct retinal cell types in the adult retina. Retinal ganglion cells (RGC), bipolar cells (BPC) and Muller cells (MC) of adult pigmented rats were analyzed for their expression of P2Y-receptor subtypes P2Y1, P2Y2, P2Y4, and P2Y6 by single-cell reverse transcription polymerase chain reaction (SC-RT-PCR). SC-RT-PCR resulted in a positive amplification signal for all P2Y-receptor subtype mRNAs in all cell types examined. However, subtype distribution differed among the different cell types. The percentage of cells expressing a distinct P2Y subtype was: (a) for RGCs: 80% with P2Y1, 100% with P2Y2, 30% with P2Y4 and 50% with P2Y6, (b) for BPCs: 60% with P2Y1, 40% with P2Y2, 20% with P2Y4 and 80% with P2Y6, and (c) for MCs: 60% with P2Y1, 80% with P2Y2, 60% with P2Y4 and 100% with P2Y6. Our data show that different subtypes of P2Y receptors (P2Y1, P2Y2, P2Y4 and P2Y6) are expressed in various retinal cells and indicate that extracellular purines and pyrimidines act on RGCs, BPCs and MCs via different P2Y receptors.

Biostability of micro-photodiode arrays for subretinal implantation.

Micro-photodiode arrays based on semiconductor chip technology are being developed to replace degenerated photoreceptor cells in the retina. Electric current is generated in tiny micro-photodiodes and delivered to the adjacent tissue by micro-electrodes. One of the main requirements of a sub-retinal implantable device is long-term stability versus corrosion in vivo (biostability). Biostability of micro-photodiode arrays (MPDA) was investigated in vitro and in vivo. No significant damage was found on chips immersed for up to 21 months in saline solution. Under in vivo conditions, however, the silicon oxide passivation layer of the chip was dissolved within a period of about 6-12 months. Subsequently, the underlying silicon was corroded. In contrast, stimulation electrodes consisting of titanium nitride were well preserved both in vitro and in vivo. The deterioration of the electrical properties of the micro-photodiodes correlated with the morphological damage observed. Strategies aiming at the development of an improved biostable encapsulation of neurotechnological implants have to be investigated and will be discussed briefly.

BDNF regulates NMDA receptor activity in developing retinal ganglion cells.

Brain-derived neurotrophic factor (BDNF) modulates glutamate receptors of the NMDA type in many areas of the brain. We assessed whether BDNF exerts an effect on NMDA receptor properties in retinal ganglion cells during early postnatal development. Electrophysiological responses to the glutamate agonist NMDA (500 microM-2 mM) in retinal slices of wildtype and BDNF deficient mice (bdnf-/-) were recorded using the whole-cell patch-clamp technique. Retinal ganglion cells of bdnf-/- mice displayed significantly smaller NMDA currents than those of age-matched wildtype mice. Remarkably, NMDA receptor activity was restored by incubating retinal slices of bdnf-/- mice in BDNF (50 ng/ml) for 1-3 h. We suggest that BDNF plays a role in the activation of functional NMDA receptors in early ganglion cell development.

Age-dependent decrease of retinal kynurenate and kynurenine aminotransferases in DBA/2J mice, a model of ocular hypertension.

The study examines age-dependent changes of kynurenic acid (KYNA) content and kynurenine aminotransferases (KAT I and KAT II) celluar expression in the retinas of DBA/2J mice. Retinas were obtained from DBA/2J mice of different ages (3, 6 and 11 months). C57BL6 mice were used as controls. As measured with HPLC, KYNA content decreased (p < 0.01) in the retinas of 6-month-old DBA/2J mice and continued to decrease (p < 0.0074) in the retinas of 11-month-old animals compared to the controls. Immunohistochemistry showed that expression of both KAT I and KAT II decreased markedly in the retinas of 11-month-old DBA/2J mice compared to controls. The impairment in KYNA biosynthesis in the retinas of DBA/2J mice may be one of the mechanisms of retinal neurodegeneration related to ocular hypertension.

Ontogenic changes of kynurenine aminotransferase I activity and its expression in the chicken retina.

Kynurenine aminotransferases are key enzymes for the synthesis of kynurenic acid (KYNA), an endogenous glutamate receptor antagonist. The study described here examined ontogenic changes of kynurenine aminotransferase I (KAT I) activity and its expression in the chicken retina. KAT I activity measured on embryonic day 16 (E16) was significantly higher than at all other stages (E12, P0 and P7). Double labeling with antibodies against glutamine synthetase showed that on P7 KAT I was expressed in Müller cell endfeet and their processes in the inner retina. Since KAT I activity is high in the late embryonic stages, it is conceivable that it plays a neuromodulatory role in the retina during the late phase of embryogenesis.