Rat Optic Nerve Research Articles

Evaluation of Fluoro-Jade C as a marker of degenerating neurons in the rat retina and optic nerve

Detection of neuronal death is an essential requirement for researchers investigating retinal degeneration. Fluoro-Jade C (FJC) is a novel, fluorescent dye that has been successfully used to label degenerating neurons in the brain, but its effectiveness in the eye has not been ascertained. In the current study, we determined the efficacy of FJC for detection of neuronal degeneration in the retina and optic nerve in various paradigms of injury. N-methyl-d-aspartate (NMDA) and kainic acid-induced excitotoxicity, optic nerve transection, and bilateral occlusion of the common carotid arteries (BCCAO) were performed using standard techniques. Rats were killed at various time points and the retinas with optic nerves attached were removed for tissue processing prior to labelling for FJC, for DNA fragmentation by TUNEL or for immunohistochemical analysis. Retinas from RCS rats of different ages were also analysed. After excitotoxicity-induced injury, cell bodies and dendrites within the ganglion cell and inner plexiform layers were specifically labelled by FJC within 6h, a time point comparable to the appearance of TUNEL-positive nuclei and to reductions in mRNA levels of retinal ganglion cell-specific proteins, but in advance of alterations in some immunohistochemical markers. The number of FJC-labelled cell bodies in the retina declined over time as cell loss proceeded, although dendritic staining remained prominent. Colocalisation of FJC with TUNEL and with immunohistochemical neuronal markers was achieved. FJC was successful at identifying somato-dendritic degeneration following ischemia induced by BCCAO, but surprisingly, not after optic nerve transection. FJC visualised photoreceptor degeneration in the RCS rat, albeit less effectively than with the TUNEL assay, and was also effective for imaging and quantifying degenerating axons in the optic nerve after multiple injuries. In addition to labelling degenerating neurons, however, FJC also bound non-specifically to astrocytes and to blood cells in unperfused rats. Since the ganglion cell layer is adjacent to astrocytes within the nerve fibre layer, caution is needed when using FJC as a quantitative tool for detecting ganglion cell death.

Immediate short-duration hypothermia provides long-term protection in an in vivo model of traumatic axonal injury

A prospective, multicenter, randomized trial did not demonstrate improved outcomes in severe traumatic brain injured patients treated with mild hypothermia [Clifton, G.L., Miller, E.R., Choi, S.C., Levin, H.S., McCauley, S., Smith, K.R., Jr., Muizelaar, J.P., Wagner, F.C., Jr., Marion, D.W., Luerssen, T.G., Chesnut, R.M., Schwartz, M., 2001. Lack of effect of induction of hypothermia after acute brain injury. N. Engl. J. Med. 344, 556–563.]. However, the mean time to target temperature was over 8 h and patient inclusion was based on Glasgow Coma Scale score so brain pathology was likely diverse. There remains significant interest in the benefits of hypothermia after traumatic brain injury (TBI) and, in particular, traumatic axonal injury (TAI), which is believed to significantly contribute to morbidity and mortality of TBI patients. The long-term beneficial effect of mild hypothermia on TAI has not been established. To address this issue, we developed an in vivo rat optic nerve stretch model of TAI. Adult male Sprague–Dawley rats underwent unilateral optic nerve stretch at 6, 7 or 8 mm piston displacement. The increased number of axonal swellings and bulbs immunopositive for non-phosphorylated neurofilament (SMI-32) seen four days after injury was statistically significant after 8 mm displacement. Ultrastructural analysis 2 weeks after 8 mm displacement revealed a 45.0% decrease ( p < 0.0001) in myelinated axonal density in the optic nerve core. There was loss of axons regardless of axon size. Immediate post-injury hypothermia (32 °C) for 3 h reduced axonal degeneration in the core ( p = 0.027). There was no differential protection based on axon size. These results support further clinical investigation of temporally optimized therapeutic hypothermia after traumatic brain injury.

Connexin43 antisense oligodeoxynucleotide treatment down-regulates the inflammatory response in an in vitro interphase organotypic culture model of optic nerve ischaemia

Using a model of optic nerve ischaemia, this study investigated oxygen-glucose deprivation (OGD) on isolated rat optic nerve segments cultured in vitro. Thereafter, the effect of antisense oligodeoxynucleotides (ASODN) specific to the gap junction protein connexin43 (Cx43) was evaluated in this same model. Following exposure to OGD for 2 hours, optic nerves were maintained in interphase organotypic culture with and without exposure to Cx43 ASODN. Optic nerves were sectioned at 2 hours, 6 hours, and at days 1, 2, 3 and 6 following culture. Cell death was quantified using propidium iodide (PI) staining and specific markers for Cx43, capillaries (von Willebrand factor), astrocytes (glial fibrillary acidic protein), microglia and endothelial cells (isolectin B4) were used to evaluate these parameters in conjunction with digital light and confocal microscopy. In this model, up-regulation of Cx43 was seen at 2 hours following exposure of the optic nerve to OGD and peaked at day 3. Cx43 ASODN treatment dampened this up-regulation. Additionally, more PI labeled cells were found in the centre of control optic nerve segments than in treated nerves ( p < 0.01). Controls also showed evidence of capillary breakdown and increased numbers of astrocytes and activated microglia compared to Cx43 ASODN treated nerves ( p < 0.05). Thus, the application of Cx43 ASODN to post-ischaemic optic nerve segments significantly reduced the up-regulation of Cx43 and, subsequently, the spread of injury and a resultant inflammatory response. Cx43 up-regulation may play an important role in optic nerve injury, offering a potential avenue for treatment in optic neuropathy.

Dosage dependence of the effect of Ginkgo biloba on the rat retinal ganglion cell survival after optic nerve crush

To investigate the dosage dependence of the Ginkgo biloba effect on retinal ganglion cell survival in the rat optic nerve crush model. The study included 56 Sprague-Dawley rats, the right optic nerve of which was crushed in a standardized manner. Two hours after the crush and once daily during the follow-up, the animals received intragastral applications of saline (saline group; n=13), or of a G. biloba extract of 0.25% concentration (n=14; low-dosage group), 1% concentration (n=15; medium-dosage group), or 4% concentration (n=14; high-dosage group). At 23 days after the optic nerve crush, the retinal ganglion cells were retrogradely labelled by injecting 3% fluorogold into the superior colliculi of the brain. At 4 weeks after baseline, the animals were killed. Retinal flat mount photographs were assessed for number and density of the retinal ganglion cells. The mean survival rate defined as the ratio of retinal ganglion cell density in the right eye with optic nerve crash divided by the retinal ganglion cell density in left eye without optic nerve intervention increased significantly (P<0.001) from 58.4+/-9.0% in the saline group to 68.5+/-5.7% in the low-dosage group, to 73.7+/-6.4% in the medium-dosage group, and to 74.2+/-6.8% in the high-dosage group. Intragastral applications of a G. biloba extract applied after an experimental and standardized optic nerve crush in rats were associated with a higher survival rate of retinal ganglion cells in a dosage-dependent manner.

Reduced NGF level and TrkA protein and TrkA gene expression in the optic nerve of rats with experimentally induced glaucoma

Glaucoma (GL) is an optic neuropathy characterized by progressive loss of visual field due to retinal cell death and optic nerve (ON) degeneration, usually in response to abnormal elevated intraocular pressure (EIOP). It has previously demonstrated that cells of the ON express nerve growth factor (NGF) and NGF-receptors. Relatively little is known, however, about their role on the ON of the glaucomatous eye. The aim of the study was to elucidate this aspect. Using a rat model of GL we investigated the response of NGF and NGF-receptors in the ON of subjects with experimentally induced EIOP. Our results show that EIOP significantly impairs the presence of NGF and NGF-receptor proteins and TrkA gene expression in the ON of glaucomatous eye. These findings suggest that NGF and NGF-receptor might be important signals for the ON response in the EIOP.

Is brimonidine better at stabilizing visual field than timolol?

Abstract Purpose Alpha‐2 adrenergic agonists are neuroprotective in animal models of focal cerebral ischemia. Brimonidine protects the retinal ganglion cell (RGC) from secondary degeneration following partial crush injury to the rat optic nerve and in an ocular hypertensive rat model. The mechanism for this effect is up‐regulation of brain‐derived neurotrophic factor in RGC and the retina. Brimonidine activates the alpha‐2 adrenoceptor at 2 nM. Topical administration of brimonidine produces pharmacologic drug concentrations in the vitreous (100‐170 nM) in humans that provides a drug delivery route to the RGCs sufficient to bind and activate the alpha‐2 adrenoceptor. In this way, brimonidine could function to maintain the health of the optic nerve independent of its ability to reduce intraocular pressure (IOP) Methods The Low‐pressure Glaucoma Treatment Study (LoGTS) is a triple masked, randomized, multicenter clinical trial of the efficacy of monotherapy with brimonidine versus timolol eye drops to alter the course of low‐pressure (normal‐tension) glaucoma (untreated IOP &lt; 22 mmHg). The primary outcome was visual field progression analyzed using point‐wise linear regression analysis. This methodology required a minimum of four visual fields to obtain and compared the slope of decibel sensitivity of each test location to all previous examinations. Field progression was defined as a negative slope &gt; 1.0 decibel/year with a significance P£0.05 for the same three or more test locations on three consecutive examinations (i.e., over an 8 month interval). Results Previously published articles contains a detailed description of the study methods and baseline patient characteristics and baseline visual field and IOP asymmetry. The current presentation will discuss LoGTS outcomes. Commercial interest

Axonal transport deficit in the optic nerve of rats with inherited retinitis pigmentosa and experimentally induced glaucoma

Aim of this study was to investigate the retrograde axonal transport from optic nerve (ON) to retinal ganglion cell (RGC) in two animal models: in Royal College of Surgeons (RCS) rats, a rat model for retinal degeneration, and in a rat model for glaucoma induced by elevated intraocular pressure (IOP). To carry out this study, dextran tetramethylrhodamine (DTMR--an hydrophilic neurotracer dye) was injected into the ON; 24 hrs later, the retina was removed and the number of labeled RGCs of the experimental rats was counted and compared. The results of these studies showed that the number of fluorescent-labeled RGCs in RCS rats and in rats with elevated IOP was reduced compared to the number of labeled RGCs of their respective controls. Our findings suggest that RCS rats are characterized not only by loss of photoreceptor cells but also by functional deficits of RGCs.

Neuroprotective effects of recombinant human granulocyte colony-stimulating factor (G-CSF) in neurodegeneration after optic nerve crush in rats

The purpose of the present study was to investigate the effects of granulocyte colony-stimulating factor (G-CSF) on neurodegeneration of optic nerve (ON) and retinal ganglion cells (RGCs) in a rat model of ON crush. The ONs of adult male Wistar rats (150–180 g) were crushed by a standardized method. The control eyes received a sham operation. G-CSF (100 μg/kg/day in 0.2 ml phosphate-buffered saline) or phosphate-buffered saline (PBS control) was immediately administered after ON crush for 5 days by subcutaneous injection. Rats were euthanized at 1 or 2 weeks after the crush injury. RGC density was counted by retrograde labeling with FluoroGold application to the superior colliculus, and visual function was assessed by flash visual evoked potentials (FVEP). TUNEL assay, Western blot analysis and immunohistochemistry of p-AKT in the retina and ED1 (marker of macrophage/microglia) in the ON were conducted. 2 weeks after the insult, the RGC densities in the central and mid-peripheral retinas in ON-crushed, G-CSF-treated rats were significantly higher than that of the corresponding ON-crushed, PBS-treated rats (survival rate was 60% vs. 19.6% in the central retina; 46.5% vs. 23.9% in mid-peripheral retina, respectively; p < 0.001). FVEP measurements showed a significantly better preserved latency of the p1 wave in the ON-crushed, G-CSF-treated rats than the ON-crushed, PBS-treated rats (78 ± 9 ms in the sham operation group, 98 ± 16 ms in the G-CSF-treated group, and 174 ± 16 ms in the PBS-treated group; p < 0.001). TUNEL assays showed fewer apoptotic cells in the retinal sections in the ON-crushed, G-CSF-treated rats. p-AKT immunoreactivity was up-regulated in the retinas of the ON-crushed, G-CSF-treated rats at 1 and 2 weeks. In addition, the number of ED1-positive cells was attenuated at the lesion site of the optic nerve in the ON-crushed, G-CSF-treated group. From these results, we gather that administration of G-CSF is neuroprotective in the rat model of optic nerve crush, as demonstrated both structurally by RGC density and functionally by FVEP. G-CSF may work by being anti-apoptotic involving the p-AKT signaling pathway as well as by attenuation of the inflammatory responses at the injury site, as evidenced by less ED1-positive cell infiltration in the optic nerve.

Functional glutamate transport in rodent optic nerve axons and glia

Recent findings suggest that synaptic-type glutamate signaling operates between axons and their supporting glial cells. Glutamate reuptake will be a necessary component of such a system. Evidence for glutamate-mediated damage of oligodendroglia somata and processes in white matter suggests that glutamate regulation in white matter structures is also of clinical importance. The expression of glutamate transporters was examined in postnatal Day 14-17 (P14-17) mouse and in mature mouse and rat optic nerve using immuno-histochemistry and immuno-electron microscopy. EAAC1 was the major glutamate transporter detected in oligodendroglia cell membranes in both developing and mature optic nerve, while GLT-1 was the most heavily expressed transporter in the membranes of astrocytes. Both EAAC1 and GLAST were also seen in adult astrocytes, but there was little membrane expression of either at P14-17. GLAST, EAAC1, and GLT-1 were expressed in P14-17 axons with marked GLT-1 expression in the axolemma, while in mature axons EAAC1 was abundant at the node of Ranvier. Functional glutamate transport was probed in P14-17 mouse optic nerve revealing Na+-dependent, TBOA-blockable uptake of D-aspartate in astrocytes, axons, and oligodendrocytes. The data show that in addition to oligodendroglia and astrocytes, axons represent a potential source for extracellular glutamate in white matter during ischaemic conditions, and have the capacity for Na(+)-dependent glutamate uptake. The findings support the possibility of functional synaptic-type glutamate release from central axons, an event that will require axonal glutamate reuptake.

Correlation of axon size and myelin occupancy in rats prenatally exposed to methamphetamine

The abuse of methamphetamine (MA) and other psychostimulants is a social and medical problem. In particular, the use of these drugs by pregnant women results in an increased number of children exposed prenatally to psychostimulants. Our previous work has demonstrated that prenatal exposure to MA affects the normal development of the rat visual system due to alterations of biochemical mechanisms and oxidative stress. It was also demonstrated that prenatal exposure to MA affects the dopaminergic system of the rat retina and optic nerve (ON) myelination. The present work was conducted to evaluate the effects of prenatal exposure to MA on the development of the ON in terms of axon growth and the myelin sheath. Pregnant female rats were given 5 mg/kg/day MA, subcutaneously (s.c.), in 0.9% saline from gestational day (GD) 8 to 22. The pair-fed control group was injected s.c. with an isovolumetric dose of 0.9% saline. Qualitative analysis was performed using representative electron ultramicrographs. Quantitative analysis was performed at an electron microscopic level on ON cross sections; parameters measured included myelinated/unmyelinated ratio, outer axon mean area, inner axon mean area, myelin mean area, myelin occupancy and distribution of axons by size. The ON of prenatally MA-exposed rats presented a higher rate of deformed axons and slighter lamellar separation. At PND 21, the average outer axon area of MA-treated males was significantly reduced. The average inner axon area only showed a significant difference between MA and control males for axons with an area of less than 0.3 µm 2. The average myelin area of MA-treated males was significantly reduced, and in MA-treated females was only significantly reduced in axons with an area of less than 0.3 µm 2. The percentage of myelin occupancy was significantly affected in MA-treated males, and in MA-treated females in the group of axons with an area of more than 0.3 µm 2. At PND 14 no significant differences were found between MA and control groups. The spectrum of ON myelinated axon size of MA-treated animals was shifted to the left at PND 14 and PND 21 for both genders. These results are in agreement with previous animal studies of prenatal and perinatal exposure to drugs of abuse. Taken together, these data indicate that the ON is vulnerable to early exposure to MA which causes developmental changes and may interfere with the functioning of the visual system.

Ouabain-induced stimulation of sodium-hydrogen exchange in rat optic nerve astrocytes

Sodium-dependent transporters are inhibited indirectly by the Na-K-ATPase inhibitor ouabain. Here we report stimulation of sodium-hydrogen exchange (NHE) in ouabain-treated cells. BCECF was used to measure cytoplasmic pH in cultured rat optic nerve astrocytes. Ammonium chloride was applied to acid load the cells. On removal of ammonium chloride, cytoplasmic pH fell abruptly, then gradually recovered toward baseline. Ouabain (1 microM) did not change cell sodium content, but the rate of pH recovery increased by 68%. Ouabain speeded pH recovery both in the presence and absence of bicarbonate. In bicarbonate-free medium, dimethylamiloride, an NHE inhibitor, eliminated the effect of 1 microM ouabain on pH recovery. Western blot analysis showed an NHE1 immunoreactive band but not NHE2, NHE3, or NHE4. Immunoprecipitation studies showed phosphorylation of NHE1 in cells treated with 1 microM ouabain. Ouabain evoked an increase of cAMP, and the effect of 1 microM ouabain on pH recovery was abolished by H-89, a protein kinase A inhibitor. 8-Bromoadenosine-cAMP increased the pH recovery rate, and this recovery was not further increased by ouabain. Although 1 microM ouabain did not alter cytoplasmic calcium concentration, it stimulated calcium entry after store depletion, a response abolished by 2-APB. Ouabain-induced stimulation of pH recovery was suppressed by inhibitors of capacitative calcium entry, SKF-96365, and 2-APB, as well as the cytoplasmic calcium chelator BAPTA. The cAMP increase in ouabain-treated cells was abolished by BAPTA and 2-APB. Taken together, the results are consistent with increased capacitative calcium entry and subsequent cAMP-PKA-dependent stimulation of NHE1 in ouabain-treated cells.

Engrafted Chicken Neural Tube–Derived Stem Cells Support the Innate Propensity for Axonal Regeneration within the Rat Optic Nerve

Injury to the adult optic nerve, caused mechanically or by diseases, is still not reparable because the retinal ganglion cells (RGCs) are not allowed to regrow their axons and die retrogradely, although they possess the intrinsic propensity to regenerate axons in experimental conditions. In vitro propagated embryonic stem cells derived from the early chicken neural tube (NTSCs) were used to examine whether transplanted NTSCs produce growth-promoting factors and pave the microenvironment, thus facilitating axonal regeneration within the rat optic nerve. NTSCs survived within the site where the optic nerve had been cut and continued to be nestin-positive, thus preserving their undifferentiated cell phenotype. Transplanted NTSCs activated the matrix metalloproteases (MMP)-2 and -14 in glial fibrillary acidic protein (GFAP)-positive optic nerve astrocytes. MMP2 production correlated with immunohistochemically visible degradation of inhibitory chondroitin sulfate proteoglycans (CSPGs). In addition, NTSCs produced a panoply of neurite-promoting factors including oncomodulin, ciliary neurotrophic factor, brain-derived neurotrophic factor and crystallins beta and gamma. Cut axons intermingled with NTSCs and passed through the zone of injury to enter the distal optic nerve over long distances, arriving at the thalamus and midbrain. This study showed evidence that paving of the distal optic nerve microenvironment with proteolytically active MMPs and providing stem-cell-derived growth factors is a suitable method for facilitating regenerative repair of the optic nerve. Understanding the molecular mechanisms of this repair has fundamental implications for development of NTSC-based subsidiary therapy after neural injuries.

Transient 5‐(4‐phenylbutoxy)psoralen (PAP‐1) treatment dissociates developing pathologies in autoimmune optic neuritis into two distinct pathology profiles

Discovery of treatments to protect axonal function of neurons and prevent permanent disability associated with progressive multiple sclerosis (MS) has faced the uphill challenge of assessing relatively small changes in accumulated axon damage within a background environment that is disorganized by CNS inflammation. We hypothesized that transient immunosuppression after initiation of MS-like autoimmune mechanisms would disassociate development of MS-like myelinated axon pathology from development of CNS inflammation in a rat model of autoimmune optic neuritis (AON). A rat model of myelin oligodendrocyte glycoprotein peptide-induced AON was transiently treated (on days 3-7 after antigen exposure) with 5-(4-phenylbutoxy)psoralen (PAP-1), an immunomodulatory drug previously shown specifically to suppress proliferation of effector memory T-cells and immunoglobulin class-switched B-cells. Thirteen days after antigen exposure, optic nerves were harvested for quantitative assessment of 12 MS-associated pathologies using microfluorimetry. With one exception, the immunoreactivities (-ir) for eight markers of MS-like neuroinflammation and immune infiltration were significantly reduced (P < 0.05) by transient PAP-1 treatment, often to levels significantly below those detected in normal control rat optic nerves. With one exception, four immunoreactive markers of MS-like myelinated axon pathology were detected at levels indicating increased axon/myelin pathology compared with vehicle-treated rats with AON (P < 0.05). These data suggest the conclusion that early causative mechanisms in CNS autoimmunity initiate signaling mechanisms that diverge into two separate pathways, one that is strongly associated with inflammatory responses and one that is associated predominantly with disturbed axon-myelin interactions and impaired fast axonal transport.

Inhibition of capacitative calcium entry suppresses ouabain‐induced stimulation of sodium‐hydrogen exchange in rat optic nerve astrocytes

Objective: Earlier we showed that 1 μM ouabain increased capacitative calcium entry (CCE) but did not increase cell sodium1. Here we studied cytoplasmic pH recovery after an acid load in ouabain‐treated cells and tested whether pH response were linked to altered calcium entry.Methods: Ammonium chloride (20mM) was used to acid load the cells. BCECF and FURA‐2 were used to measure intracellular pH and calcium.Results: Ammonium chloride removal causes the pHi to fall abruptly, followed by gradual recovery. lμM ouabain increased the rate of pHi recovery both in the presence and in the absence of bicarbonate in the bathing medium. NHE inhibitor DMA reduced the rate of pHi recovery. Ouabain did not detectably alter cytoplasmic Ca2+ (Ca2+i) but CCE inhibitors SKF96365 and 2APB reduced both the basal and ouabain‐stimulated rates of pHi recovery. Calcium chelator BAPTA‐AM also inhibited basal and ouabain‐stimulated pHi recovery. Western blot showed presence of NHE1 but not NHE2, NHE3 or NHE4.Conclusion: Ouabain‐induced stimulation of pHi recovery was due, at least in part, to stimulation of NHE1. Although lμM ouabain failed to alter Ca2+i, the findings suggest the NHE response may be dependent on CCE.Funding: NIH Grant EY014069

Transport of multiple nicotinic acetylcholine receptors in the rat optic nerve: high densities of receptors containing α6 and β3 subunits

Neuronal nicotinic acetylcholine receptors (nAChRs) are abundant in the rat retina and at least seven heteromeric subtypes have been detected. Axons of retinal ganglion cells form the optic nerve and innervate areas of the brain important for visual processing, including the lateral geniculate nucleus, the superior colliculus, and the pretectal nucleus. Development of eye-specific layers in these projection areas are dependent upon retinal waves which are initially mediated by nAChRs [Feller et al., Science 272 (1996), 1182; Penn et al., Science 279 (1998), 2108; Bansal et al., J. Neurosci. 20 (2000), 7672]. Unilateral eye-enucleation studies in the rat indicate that nAChRs are on the terminals of optic nerve axons, where they may mediate influences of acetylcholine on visual pathways. In this study, we use radioligand binding and immunoprecipitation with subunit-selective antibodies to investigate the subunit composition of nAChRs in the rat optic nerve. We found multiple nAChR subtypes in the optic nerve, all of which contain the beta2 subunit. Most of these receptors are mixed heteromeric subtypes, composed of at least three different subunits. Included among these subtypes is the highest percentage and density of alpha6- and beta3-containing nAChRs of any area of the rat CNS that has been reported.

The serine racemase mRNA is expressed in both neurons and glial cells of the rat retina

D-Serine, an endogenous and obligatory coagonist for the glycine site of the N-methyl-D-aspartate receptor in mammals, is synthesized from L-serine by serine racemase. Serine racemase and D-serine have long been believed to occur predominantly in astrocytes, according to immunohistochemical studies. Recent studies have demonstrated, however, that both the mRNA and protein levels of serine racemase are considerably higher in neurons than in astrocytes in primary cultures of the rat brain and that the mRNA level of serine racemase predominates in neurons of the adult rat brain. Here we report the application of in situ hybridization based on tyramide signal amplification for the detection of serine racemase mRNA in sections of the adult rat retina and optic nerve head. The localization of serine racemase mRNA could be demonstrated in ganglion cells, amacrine cells, bipolar cells, horizontal cells, and Müller cells of the retina as well as in the astrocytes of the optic nerve head and the lamina cribrosa. This is the first study to demonstrate the exact localization of serine racemase mRNA at the cellular or tissue level in the retina and the optic nerve head. These results suggest that both the neuron- and glia-derived D-serine could modulate neurotransmission via the glycine site of the N-methyl-D-aspartate receptors in the retina.

Rat retinal morphological changes after optic nerve crush

目的 探讨大鼠视神经夹伤后视网膜病变的时序性变化.方法 建立大鼠视神经夹伤模型.分别于伤后1、3、7、15、30 d光镜下计数视网膜神经节细胞(RGCs)数目的变化.结果 大鼠视神经夹伤诱导RGCs较正常眼明显减少.结论 应用40 g力视神经夹建立的定量视神经损伤动物模型是可行的,并可应用于视神经损伤修复的研究中。

Axon tracing in the adult rat optic nerve and tract after intravitreal injection of MnDPDP using a semiautomatic segmentation technique

To develop and validate an objective technique for 3D segmentation of manganese-enhanced MR images of the optic nerve/tract (ON) in adult rats to improve contrast-to-noise (CNR) calculations and use the technique to ascertain if manganese dipyridoxyl diphosphate (MnDPDP) gives sufficient Mn(2+) enhancement compared to MnCl(2) when used for functional imaging of the visual pathway. Intravitreous injection of the manganese-releasing MR contrast agent MnDPDP (30 nmol Mn(2+)) was performed to trace the ON in adult rats (n = 4). A positive control group of rats (n = 5) received a standard preparation of MnCl(2) (200 nmol Mn(2+)), while gadodiamide (1500 nmol Gd(3+)) was administered in negative control rats (n = 2). An objective technique for 3D segmentation of the enhanced ON was developed. CNR profiles along the ON were calculated by resampling the 3D image-volume in 2D planes perpendicular to the Mn(2+) enhanced ON in 0.2 mm steps, 4 mm along the segmented ON measured from the lamina cribrosa. The ON was successfully segmented and CNR calculated accurately within 2 minutes in a representative 3D MR image volume. Intravitreal MnDPDP injection resulted in significant MRI contrast enhancement of the retina and ON after 12-24 hours similar to that of MnCl(2) injection. 3D semiautomated image segmentation and the use of MnDPDP can improve in vivo axon tracing based on MRI. Mn(2+) was found to be released from MnDPDP after intravitreal injection in sufficient amounts to obtain functional tracing of the adult rat primary visual pathway.

Protective effect of modified and wild type acidic fibroblast growth factor after rat optic nerve crush

目的 观察改构体酸性成纤维细胞生长因子(MaFGF)和野生型酸性成纤维细胞生长因子(aFGF)对视神经夹挫伤后的保护作用.方法 健康Wistar大鼠42只随机分为7组:生理盐水组,aFGF 0.25、1.0、4.0μg/mL组,MaFGF0.25、1.0、4.0μg/mL组.均制成右眼视神经夹挫伤模型,于伤后1 h及4周分别记录左右眼的闪光视觉诱发电位(F-VEP),分5次向右眼球后注入各组液体0.1 mL.取眼球及部分视神经,光镜下观察形态学改变,计数视网膜神经节细胞(RGCs).结果 伤后1 h,MaFGF和aFGF各组F-VEP潜伏期均延长,波幅均降低,与正常对照组相比差异均有统计学意义(P＜0.01).伤后4周,较生理盐水组F-VEP更接近正常值(P＜0.05),RGCs计数均明显增多(P＜0.01).结论 MaFGF和aFGF对轴索的退变和RGCs有明显的保护作用,且这种保护作用不依赖于其有丝分裂活性。

Evoked membrane potential change in rat optic nerve fiber: Computer simulation

The optic nerve is a key component regarding research on visual prosthesis. Previous pharmacological and electrical studies has pinned down the main features of the mechanisms underlying the nerve impulse in the rat optic nerve, and this work proposed a mathematical model to simulate these phenomena. The main active nodal channels: fast Na+, persistent Na+, slow K+ and a fast repolarizing K+ (A-current) were added on a double layer representation of the axon. A simplified representation of K+ accumulation and clearance in the vicinity of the Ranvier node was integrated in this model. The model was able to generate the following features. In the presence of 4-aminopyridine (4-AP), spike duration increased and a depolarizing afterpotential (DAP) appeared. In the presence of 4-AP and tetraethylammonium (TEA), the DAP was followed by a hyperpolarizing afterpotential (AHP) and the amplitude of this AHP increased with the frequency of the stimulation. In normal conditions (no drugs): DAP and AHP were absent after a single action potential (AP) and a short train of AP; there was a relative refractoriness in amplitude lasting for 30 ms after an AP; an early AHP was revealed by a continuous depolarizing current; and there was a partial spike adaptation for a long current step stimulus. The model successfully reproduced previous experiments results including long-lasting stimulation experiment, which is known to modify nerve physiological parameter values and is a key issue for visual prosthesis research.