BDNF improves axon transportation and rescues visual function in a rodent model of acute elevation of intraocular pressure.

Optic neuropathies lead to blindness; the common pathology is the degeneration of axons of the retinal ganglion cells. In this study, we used a rat model of retinal ischemia-reperfusion and a one-time intravitreal brain-derived neurotrophic factor (BDNF) injection; then we examined axon transportation function, continuity, physical presence of axons in different part of the optic nerve, and the expression level of proteins involved in axon transportation. We found that in the disease model, axon transportation was the most severely affected, followed by axon continuity, then the number of axons in the distal and proximal optic nerve. BDNF treatment relieved all reductions and significantly restored function. The molecular changes were more minor, probably due to massive gliosis of the optic nerve, so interpretation of protein expression data should be done with some caution. The process in this acute model resembles a fast-forward of changes in the chronic model of glaucoma. Therefore, impairment in axon transportation appears to be a common early process underlying different optic neuropathies. This research on effective intervention can be used to develop interventions for all optic neuropathies targeting axon transportation.

Decreasing intraocular pressure significantly improves retinal vessel density, cytoarchitecture and visual function in rodent oxygen induced retinopathy.

We attempted to explore a noninvasive, easily applicable and economically affordable therapy for retinopathy of prematurity (ROP). Rat pups were raised in 80% oxygen from postnatal day 7 to P12, and returned to room air. Travoprost eye drops were administered twice a day for 7 days, to reduce intraocular pressure (IOP) by about 20%. Immunohistochemical staining was performed to visualize vessel endothelial cells, to analyze retinal neurons and cytoarchitecture. Behavioral experiments were carried out to test visual acuity and contrast sensitivity. At the end of the 7-day treatment, the number of vessels extending to the vitreous body was significantly reduced and retinal vessel density increased. This improvement was maintained to the end of the 12th week. In the central retina of the model group, the horizontal cells were completely wiped out, the outer plexiform layer was undetectable, and the rod bipolar cell dendrites sprouted into the outer nuclear layer. The treatment partially reverted these architectural changes. Most importantly, behavioral experiments revealed significantly improved visual acuity and contrast sensitivity in the treated group. Therefore, reducing IOP could potentially serve as a safe and economical measure to treat ROP.

Long-lasting impairments in rodent oxygen-induced retinopathy measured by retinal vessel density and visual function.

In mild or moderate retinopathy of prematurity (ROP), retinal vessels undergo obliteration, proliferation, and regression. Despite complete regression of vessel abnormalities, a variety of visual impairments have been reported. Rodent oxygen-induced retinopathy (OIR) is widely used as a model to study ROP. However, the long-term changes of OIR model remain unclear. The aim of this study is to examine long term changes of retinal vessel and visual function in a rodent OIR model resembling human mild or moderate ROP. In this study, after subjecting the animals to 80% oxygen (O2) for 5-7 d, the retinal vessel density at postnatal day 12 (P12) was approximately 30% lower than that in the age-matched control, but this difference was not significant between the groups. Vessel abnormalities, such as vessel tortuosity, neovascular tufts, and the number of vessels protruding into the vitreous, peaked between P17 and P20. Despite the regression of many abnormalities, vessel density in the OIR group was 36% and 32% lower than that in the control animals at 6 weeks and 4 months, respectively. The visual acuity and contrast sensitivity were impaired in the OIR group when measured at 2, 3 and 4 months. Therefore, the rodent OIR model exhibited long-lasting reduction in retinal vessel density and visual impairments, similar to those observed in mild or moderate human ROP. This study suggests that the rodent OIR model can be used to explore possible interventions for mild and moderate human ROP.

Different functional susceptibilities of mouse retinal ganglion cell subtypes to optic nerve crush injury.

In optic neuropathies, the progressive deterioration of retinal ganglion cell (RGC) function leads to irreversible vision loss. Increasing experimental evidence suggests differing susceptibility for RGC functional subtypes. Here with multi-electrode array recordings, RGC functional loss was characterized at multiple time points in a mouse model of optic nerve crush. Firing rate, latency of response and receptive field size were analyzed for ON, OFF and ON-OFF RGCs separately. It was observed that responses and receptive fields of OFF cells were impaired earlier than ON cells after the injury. For the ON-OFF cells, the OFF component of response was also more susceptible to optic nerve injury than the ON component. Moreover, more ON transient cells survived than ON sustained cells post the crush, implying a diversified vulnerability for ON cells. Together, these data support the contention that RGCs' functional degeneration in optic nerve injury is subtype dependent, a fact that needs to be considered when developing treatments of glaucomatous retinal ganglion cell degeneration and other optic neuropathies.

Temporal properties of dual-peak responses of mouse retinal ganglion cells and effects of inhibitory pathways.

Dual-peak responses of retinal ganglion cells (RGCs) are observed in various species, previous researches suggested that both response peaks were involved in retinal information coding. In the present study, we investigated the temporal properties of the dual-peak responses recorded in mouse RGCs elicited by spatially homogeneous light flashes and the effect of the inhibitory inputs mediated by GABAergic and/or glycinergic pathways. We found that the two peaks in the dual-peak responses exhibited distinct temporal dynamics, similar to that of short-latency and long-latency single-peak responses respectively. Pharmacological studies demonstrated that the application of exogenous GABA or glycine greatly suppressed or even eliminated the second peak of the cells' firing activities, while little change was induced in the first peak. Co-application of glycine and GABA led to complete elimination of the second peak. Moreover, application of picrotoxin or strychnine induced dual-peak responses in some cells with transient responses by unmasking a second response phase. These results suggest that both GABAergic and glycinergic pathways are involved in the dual-peak responses of the mouse RGCs, and the two response peaks may arise from distinct pathways that would converge on the ganglion cells.

Overexpression of Wnt5a in mouse epidermis causes no psoriasis phenotype but an impairment of hair follicle anagen development.

Increased Wnt5a expression has been observed in psoriatic plaques. However, whether Wnt5a overexpression directly causes psoriasis is unknown. In this study, we generated transgenic (TG) mice with epidermal Wnt5a overexpression under the control of the human K14 promoter. The skin of Wnt5a TG mice was not psoriatic, but characterized with normal proliferation and homeostasis of epidermis. Instead, these TG mice displayed impaired hair follicle transition from telogen to anagen, most likely due to impaired canonical Wnt signalling. These results suggest that increased Wnt5a expression alone is inadequate to induce psoriasis in the skin and possible involvement of Wnt5a in hair follicle cycling.

Constitutive activation of ectodermal ß-catenin induces ectopic outgrowths at various positions in mouse embryo and affects abdominal ventral body wall closure.

Vertebrate limbs originate from the lateral plate mesoderm (LPM) and the overlying ectoderm. While normal limb formation in defined regions has been well studied, the question of whether other positions retain limb-forming potential has not been fully investigated in mice. By ectopically activating ß-catenin in the ectoderm with Msx2-cre, we observed that local tissue outgrowths were induced, which either progressed into limb-like structure within the inter-limb flank or formed extra tissues in other parts of the mouse embryo. In the presumptive abdominal region of severely affected embryos, ectopic limb formation was coupled with impaired abdominal ventral body wall (AVBW) closure, which indicates the existence of a potential counterbalance of limb formation and AVBW closure. At the molecular level, constitutive ß-catenin activation was sufficient to trigger, but insufficient to maintain the ectopic expression of a putative limb-inducing factor, Fgf8, in the ectoderm. These findings provide new insight into the mechanism of limb formation and AVBW closure, and the crosstalk between the Wnt/ß-catenin pathway and Fgf signal.

Drug addiction is associated with leukocyte telomere length.

Telomeres are protective chromosomal structures that play a key role in preserving genomic stability. Telomere length is known to be associated with ageing and age-related diseases. To study the impairment of telomeres induced by drug abuse, we conducted an association study in the Chinese Han population. Multivariate linear regression analyses were performed to evaluate the correlation of leukocyte telomere length (LTL) with addiction control status adjusted for age and gender. The results showed that drug abusers exhibited significantly shorter LTLs than controls (P = 1.32e-06). The time before relapse also presented an inverse correlation with LTL (P = 0.02). Drug abusers who had used heroin and diazepam displayed a shorter LTL than those taking other drugs (P = 0.018 and P = 0.009, respectively). Drug abusers who had ingested drugs via snuff exhibited longer LTLs than those using other methods (P = 0.02). These observations may offer a partial explanation for the effects of drug addiction on health.

Long-term rescue of rat retinal ganglion cells and visual function by AAV-mediated BDNF expression after acute elevation of intraocular pressure.

PURPOSE: To evaluate the ability of increased expression of brain-derived neurotrophic factor (BDNF) using adenoassociated viral (AAV) vector to prevent the loss of rat retinal ganglion cells (RGCs) and visual function after acute elevation of intraocular pressure (IOP). METHODS: AAV vectors (expressing BDNF or GFP) were injected into the vitreous 6 hours after a transient IOP elevation to 130 mm Hg for 45 minutes. Protective effects were evaluated by counting RGCs retrogradely labeled with fluorogold (FG) from the superior colliculus, measuring the amplitude and the latency of the P1 component of the visual evoked potential (VEP), and observing the visual acuity and contrast sensitivity in awake and behaving animals. RESULTS: RGC numbers decreased continuously to 9 weeks after the elevation of IOP. FG-positive RGC loss was significantly decreased in the retinas treated with AAV-BDNF at 3, 6, and 9 weeks after the insult, with corresponding improvements in VEP parameters. Supplementing BDNF protein once to compensate for the slow onset of AAV-mediated gene expression rescued a larger number of RGCs and the parameters of the VEP. Visual acuity and contrast sensitivity were significantly improved in all treated groups, with the largest improvement in the combined-therapy group, and were maintained for up to 70 weeks. The authors further demonstrated that BDNF rescued the RGCs by activating TrkB receptors through both autocrine and paracrine mechanisms. CONCLUSIONS: AAV-mediated BDNF expression in the rat retina achieved a sustained rescue of RGCs and visual function after an acute elevation of IOP.

Direct reprogramming of Sertoli cells into multipotent neural stem cells by defined factors.

Multipotent neural stem/progenitor cells hold great promise for cell therapy. The reprogramming of fibroblasts to induced pluripotent stem cells as well as mature neurons suggests a possibility to convert a terminally differentiated somatic cell into a multipotent state without first establishing pluripotency. Here, we demonstrate that Sertoli cells derived from mesoderm can be directly converted into a multipotent state that possesses neural stem/progenitor cell properties. The induced neural stem/progenitor cells (iNSCs) express multiple NSC-specific markers, exhibit a global gene-expression profile similar to normal NSCs, and are capable of self-renewal and differentiating into glia and electrophysiologically functional neurons. iNSC-derived neurons stain positive for tyrosine hydroxylase (TH), γ-aminobutyric acid, and choline acetyltransferase. In addition, iNSCs can survive and generate synapses following transplantation into the dentate gyrus. Generation of iNSCs may have important implications for disease modeling and regenerative medicine.

Direction-selective circuitry in rat retina develops independently of GABAergic, cholinergic and action potential activity.

The ON-OFF direction selective ganglion cells (DSGCs) in the mammalian retina code image motion by responding much more strongly to movement in one direction. They do so by receiving inhibitory inputs selectively from a particular sector of processes of the overlapping starburst amacrine cells, a type of retinal interneuron. The mechanisms of establishment and regulation of this selective connection are unknown. Here, we report that in the rat retina, the morphology, physiology of the ON-OFF DSGCs and the circuitry for coding motion directions develop normally with pharmacological blockade of GABAergic, cholinergic activity and/or action potentials for over two weeks from birth. With recent results demonstrating light independent formation of the retinal DS circuitry, our results strongly suggest the formation of the circuitry, i.e., the connections between the second and third order neurons in the visual system, can be genetically programmed, although emergence of direction selectivity in the visual cortex appears to require visual experience.

Tracer coupling of intrinsically photosensitive retinal ganglion cells to amacrine cells in the mouse retina.

Intrinsically photosensitive retinal ganglion cells (ipRGCs) are a subtype of ganglion cell in the mammalian retina that expresses the photopigment melanopsin and drives non-image-forming visual functions. Three morphological subtypes of ipRGCs (M1, M2, and M3) have been described based on their dendritic stratifications in the inner plexiform layer (IPL), but the question of their potential interactions via electrical coupling remains unsettled. In this study, we have addressed this question in the mouse retina by, injecting the tracer Neurobiotin into ipRGCs that had been genetically labelled with the fluorescent protein, tdTomato. We confirmed the presence of the M1-M3 subtypes of ipRGCs based on their distinct dendritic stratifications. All three subtypes were tracer coupled to putative amacrine cells situated within the ganglion cell layer (GCL) but not the inner nuclear layer (INL). The cells tracer coupled to the M1 and M2 cells were shown to be widefield GABA-immunoreactive amacrine cells. We found no evidence of homologous tracer coupling of ipRGCs or heterologous coupling to other types of ganglion cells.

Properties of mouse retinal ganglion cell dendritic growth during postnatal development.

The property of dendritic growth dynamics during development is a subject of intense interest. Here, we investigated the dendritic motility of retinal ganglion cells (RGCs) during different developmental stages, using ex vivo mouse retina explant culture, Semliki Forest Virus transfection and time-lapse observations. The results illustrated that during development, the dendritic motility underwent a change from rapid growth to a relatively stable state, i.e., at P0 (day of birth), RGC dendrites were in a highly active state, whereas at postnatal 13 (P13) they were more stable, and at P3 and P8, the RGCs were in an intermediate state. At any given developmental stage, RGCs of different types displayed the same dendritic growth rate and extent. Since the mouse is the most popular mammalian model for genetic manipulation, this study provided a methodological foundation for further exploring the regulatory mechanisms of dendritic development.

Changing dendritic field size of mouse retinal ganglion cells in early postnatal development.

During early postnatal development, dendrites of retinal ganglion cells (RGCs) extend and branch in the inner plexiform layer to establish the adult level of stratification, pattern of branching, and coverage. Many studies have described the branching patterns, transient features, and regulatory factors of stratification of the RGCs. The rate of RGC dendritic field (DF) expansion relative to the growing retina has not been systematically investigated. In this study, we used two methods to examine the relative expansion of RGC DFs. First, we measured the size of RGC DFs and the diameters of the eyeballs at several postnatal stages. We compared the measurements with the RGC DF sizes calculated from difference of the eyeball sizes based on a linear expansion assumption. Second, we used the number of cholinergic amacrine cells (SACs) circumscribed by the DFs of RGCs at corresponding time points as an internal ruler to assess the size of DFs. We found most RGCs exhibit a phase of faster expansion relative to the retina between postnatal day 8 (P8) and P13, followed by a phase of retraction between P13 and adulthood. The morphological alpha cells showed the faster growing phase but not the retraction phase, whereas the morphological ON-OFF direction selective ganglion cells expanded in the same pace as the growing retina. These findings indicate different RGCs show different modes of growth, whereas most subtypes exhibit a fast expansion followed by a retraction phase to reach the adult size.

Change detection by thalamic reticular neurons.

The thalamic reticular nucleus (TRN) is thought to function in the attentional searchlight. We analyzed the detection of deviant acoustic stimuli by TRN neurons and the consequences of deviance detection on the TRN target, the medial geniculate body (MGB) of the rat. TRN neurons responded more strongly to pure-tone stimuli presented as deviant stimuli (low appearance probability) than those presented as standard stimuli (high probability) (deviance-detection index = 0.321). MGB neurons also showed deviance detection in this procedure, albeit to a smaller extent (deviance-detection index = 0.154). TRN neuron deviance detection either enhanced (14 neurons) or suppressed (27 neurons) MGB neuronal responses to a probe stimulus. Both effects were neutralized by inactivation of the auditory TRN. Deviance modulation effects were cross-modal. Deviance detection probably causes TRN neurons to transiently deactivate surrounding TRN neurons in response to a fresh stimulus, altering auditory thalamus responses and inducing attention shift.

Physiological properties of direction-selective ganglion cells in early postnatal and adult mouse retina.

Selective responses of retinal ganglion cells (RGCs) to the direction of motion have been recorded extracellularly from the rabbit and the mouse retina at eye opening. Recently, it has been shown that the development of this circuitry is light independent. Using whole-cell patch clamp recording, we report here that mouse early postnatal direction-selective ganglion cells (DSGCs) showed lower membrane excitability, lower reliability of synaptic transmission and much slower kinetics of light responses compared with adult DSGCs. However, the degree of direction selectivity of early postnatal DSGCs measured by the direction-selective index and the width of the directional tuning curve was almost identical to that of adult DSGCs. The DSGCs exhibited a clear selectivity for the direction of motion at the onset of light sensitivity. Furthermore, the degree of direction selectivity was not affected by rearing in complete darkness from birth to postnatal day 11 or 30. The formation of the retinal neurocircuitry for coding motion direction is completely independent of light.

Slow recovery from excitation of thalamic reticular nucleus neurons.

Responses to repeated auditory stimuli were examined in 103 neurons in the auditory region of the thalamic reticular nucleus (TRN) and in 20 medial geniculate (MGB) neurons of anesthetized rats. A further six TRN neurons were recorded from awake rats. The TRN neurons showed strong responses to the first trial and weak responses to the subsequent trials of repeated auditory stimuli and electrical stimulation of the MGB and auditory cortex when the interstimulus interval (ISI) was short (<3 s). They responded to the second trial when the interstimulus interval was lengthened to >or=3 s. These responses contrasted to those of MGB neurons, which responded to repeated auditory stimuli of different ISIs. The TRN neurons showed a significant increase in the onset auditory response from 9.5 to 76.5 Hz when the ISI was increased from 200 ms to 10 s (P<0.001, ANOVA). The duration of the auditory-evoked oscillation was longer when the ISI was lengthened. The slow recovery of the TRN neurons after oscillation of burst firings to fast repetitive stimulus was a reflection of a different role than that of the thalamocortical relay neurons. Supposedly the TRN is involved in the process of attention such as attention shift; the slow recovery of TRN neurons probably limits the frequent change of the attention in a fast rhythm.

Alpha cells take off first.

Using a transgenic mouse line in which GFP is expressed in a single population of retinal ganglion cells (RGCs), Huberman and colleagues report in this issue of Neuron that the axon terminals of RGCs exhibit an orderly pattern of distribution in the higher visual centers. This pattern undergoes a developmental refinement, and synchronous activity in the retina regulates columnar but not laminar formation.

ON direction-selective ganglion cells in the mouse retina.

Two types of ganglion cells (RGCs) compute motion direction in the retina: the ON-OFF direction-selective ganglion cells (DSGCs) and the ON DSGCs. The ON DSGCs are much less studied mostly due to the low encounter rate. In this study, we investigated the physiology, dendritic morphology and synaptic inputs of the ON DSGCs in the mouse retina. When a visual stimulus moved back and forth in the preferred-null axis, we found that the ON DSGCs exhibited a larger EPSC when the visual stimulus moved in the preferred direction and a larger IPSC in the opposite, or null direction, similar to what has been found in ON-OFF DSGCs. This similar synaptic input pattern is in contrast to other well-known differences, namely: profile of velocity sensitivity, distribution of preferred directions, and different central projection of the axons. Immunohistochemical staining showed that the dendrites of ON DSGCs exhibited tight cofasciculation with the cholinergic plexus. These findings suggest that cholinergic amacrine cells may play an important role in generating direction selectivity in the ON DSGCs, and that the mechanism for coding motion direction is probably similar for the two types of DSGCs in the retina.