Breaking the balance: ocular BDNF-injections induce visual asymmetry in pigeons

In pigeons, asymmetric photic stimulation around hatch induces functional visual asymmetries that are accompanied by left-right differences in tectal cell sizes. Different aspects of light-dependent neuronal differentiation are known to be mediated by the brain-derived neurotrophic factor (BDNF). Therefore, we investigated by means of single or triple BDNF- or saline-injections into the right eye of dark-incubated pigeon hatchlings if ocular BDNF enrichment mimics the effects of biased visual input. As adults, the birds were tested in a grit-grain discrimination task to estimate the degree and direction of visual lateralization followed by a morphometric analysis of retinal and tectal cells. The grit-grain discrimination task demonstrated that triple BDNF-injections enhanced visuoperceptual and visuomotor functioning of the left eye system. Morphometric analysis showed bilateral cell-type dependent effects within the optic tectum. While single-BDNF injections increased cell body sizes of calbindin-positive efferent neurons, triple-injections decreased cell sizes of parvalbumin-positive cells. Moreover, single BDNF-injections increased retinal cell sizes within the contralateral eye. Analysis of BDNF-induced intracellular signaling demonstrated enhanced downstream Ras activation for at least 24 h within both tectal halves whereas activity changes within the contralateral retina could not be detected. This points to primarily tectal effects of ocular BDNF. In sum, exogenous BDNF modulates the differentiation of retinotectal circuitries and dose-dependently shifts lateralized visuomotor processing towards the noninjected side. Since these effects are opposite to embryonic light stimulation, it is unlikely that the impact of light onto asymmetry formation is mediated by retinal BDNF.     

BDNF increases synapse density in dendrites of developing tectal neurons in vivo

Neuronal connections are established through a series of developmental events that involve close communication between pre- and postsynaptic neurons. In the visual system, BDNF modulates the development of neuronal connectivity by influencing presynaptic retinal ganglion cell (RGC) axons. Increasing BDNF levels in the optic tectum of Xenopus tadpoles significantly increases both axon arborization and synapse density per axon terminal within a few hours of treatment. Here, we have further explored the mechanisms by which BDNF shapes synaptic connectivity by imaging tectal neurons, the postsynaptic partners of RGCs. Individual neurons were co-labeled with DsRed2 and a GFP-tagged postsynaptic density protein (PSD95-GFP) to visualize dendritic morphology and postsynaptic specializations simultaneously in vivo. Immunoelectron microscopy confirmed that PSD95-GFP predominantly localized to ultrastructurally identified synapses. Time-lapse confocal microscopy of individual, double-labeled neurons revealed a coincident, activity-dependent mechanism of synaptogenesis and axon and dendritic arbor growth, which is differentially modulated by BDNF. Microinjection of BDNF into the optic tectum significantly increased synapse number in tectal neuron dendritic arbors within 24 hours, without significantly influencing arbor morphology. BDNF function-blocking antibodies had opposite effects. The BDNF-elicited increase in synapse number complements the previously observed increase in presynaptic sites on RGC axons. These results, together with the timescale of the response by tectal neurons, suggest that the effects of BDNF on dendritic synaptic connectivity are secondary to its effects on presynaptic RGCs. Thus, BDNF influences synaptic connectivity in multiple ways: it enhances axon arbor complexity expanding the synaptic territory of the axon, while simultaneously coordinating synapse formation and stabilization with individual postsynaptic cells.     

Neurotrophin-3 is required for appropriate establishment of thalamocortical connections

Ca2+-permeable AMPARs are inwardly rectifying due to block by intracellular polyamines. Neuronal activity regulates polyamine synthesis, yet whether this affects Ca2+-AMPAR-mediated synaptic transmission is unknown. We test whether 4 hr of increased visual stimulation regulates glutamatergic retino-tectal synapses in Xenopus tadpoles. Tectal neurons containing Ca2+-AMPARs form a gradient along the rostro-caudal developmental axis. These neurons had inwardly rectifying AMPAR-mediated EPSCs. Four hours of visual stimulation or addition of intracellular spermine increased rectification in immature neurons. Polyamine synthesis inhibitors blocked the effect of visual stimulation, suggesting that visual activity regulates AMPARs via the polyamine synthesis pathway. This modulation resulted in changes in the integrative properties of tectal neurons. Regulation of polyamine synthesis by physiological stimuli is a novel form of modulation of synaptic transmission important for understanding the short-term effects of enhanced sensory experience during development.     

Spatiotemporal specificity of neuronal activity directs the modification of receptive fields in the developing retinotectal system

The precise temporal relation between pre- and postsynaptic activity modulates the strength of synaptic connections. Despite its extensive characterization in vivo and in vitro, the degree to which spike timing-dependent plasticity can shape receptive field properties is unclear. We use in vivo patch-clamp recordings of tectal neurons in developing Xenopus tadpoles to control the precise timing of action potentials with respect to the arrival of a subset of visual inputs evoked by local light stimulation on the retina. The pattern of visual inputs onto a tectal neuron was tracked over time by rapid reverse correlation mapping of receptive fields. Spike timing-dependent potentiation or depression of a subset of synapses reliably shifts the spatial receptive fields toward or away from the trained subregion of visual space, respectively. These results demonstrate that natural patterns of activity evoked by sensory stimuli play an instructive role in the developing nervous system.     

Influence of the motor cortex on lateral geniculate responses evoked in the cat by contralateral superior colliculus stimulation

It is shown that in nembutal anesthetized cats, a single stimulation of motor cortex (MC) causes a response in lateral geniculate nucleus (LGN). The development of this response had a conditioning effect on the LGN response evoked by stimulation of the contralateral superior colliculus (SC), markedly inhibiting it. The degree of this inhibition depended on the time interval between the cortical conditioning stimulation and the tectal test stimulation. A single conditioning MC stimulation did not noticeably change the LGN responses evoked by a light stimulus, but markedly inhibited visual responses from deep SC layers (those regions which on stimulation gave rise to LGN responses). From the results, it is suggested that the MC monitors the execution of tectal influences on LGN function at the tectal level rather than the geniculate level, and it is precisely by this means that it regulates saccadic suppression of LGN function, in the realization of which, as presumed earlier, the SC takes part.A. I. Karaev Institute of Physiology, Azerbaijan Academy of Sciences, Baku. Translated from Neirofiziologiya, Vol. 24, No. 4, July–August 1992.     

Activity-dependent matching of excitatory and inhibitory inputs during refinement of visual receptive fields

The receptive field (RF) of single visual neurons undergoes progressive refinement during development. It remains largely unknown how the excitatory and inhibitory inputs on single developing neurons are refined in a coordinated manner to allow the formation of functionally correct circuits. Using whole-cell voltage-clamp recording from Xenopus tectal neurons, we found that RFs determined by excitatory and inhibitory inputs in more mature tectal neurons are spatially matched, with each spot stimulus evoking balanced synaptic excitation and inhibition. This emerges during development through a gradual reduction in the RF size and a transition from disparate to matched topography of excitatory and inhibitory inputs to the tectal neurons. Altering normal spiking activity of tectal neurons by either blocking or elevating GABA(A) receptor activity significantly impeded the developmental reduction and topographic matching of RFs. Thus, appropriate inhibitory activity is essential for the coordinated refinement of excitatory and inhibitory connections.     

Peripherally-derived BDNF promotes regeneration of ascending sensory neurons after spinal cord injury

Background

The blood brain barrier (BBB) and truncated trkB receptor on astrocytes prevent the penetration of brain derived neurotrophic factor (BDNF) applied into the peripheral (PNS) and central nervous system (CNS) thus restrict its application in the treatment of nervous diseases. As BDNF is anterogradely transported by axons, we propose that peripherally derived and/or applied BDNF may act on the regeneration of central axons of ascending sensory neurons.

Methodology/Principal Findings

The present study aimed to test the hypothesis by using conditioning lesion of the sciatic nerve as a model to increase the expression of endogenous BDNF in sensory neurons and by injecting exogenous BDNF into the peripheral nerve or tissues. Here we showed that most of regenerating sensory neurons expressed BDNF and p-CREB but not p75NTR. Conditioning-lesion induced regeneration of ascending sensory neuron and the increase in the number of p-Erk positive and GAP-43 positive neurons was blocked by the injection of the BDNF antiserum in the periphery. Enhanced neurite outgrowth of dorsal root ganglia (DRG) neurons in vitro by conditioning lesion was also inhibited by the neutralization with the BDNF antiserum. The delivery of exogenous BDNF into the sciatic nerve or the footpad significantly increased the number of regenerating DRG neurons and regenerating sensory axons in the injured spinal cord. In a contusion injury model, an injection of BDNF into the footpad promoted recovery of motor functions.

Conclusions/Significance

Our data suggest that endogenous BDNF in DRG and spinal cord is required for the enhanced regeneration of ascending sensory neurons after conditioning lesion of sciatic nerve and peripherally applied BDNF may have therapeutic effects on the spinal cord injury.     

Neurotrophins, but not depolarization, regulate substance P expression in the developing optic tectum.

Neurotransmitter expression can be regulated by both activity and neurotrophins in a number of in vitro systems. We examined whether either of these factors was likely to play a role in the in vivo optic nerve-dependent regulation of a substance P-like immunoreactive (SP-ir) population of cells in the developing optic tectum of the frog. In contrast to our previous results with the adult system, blocking tectal cell responses to glutamate release by retinal ganglion cells with 6-cyano-7-nitroquinoxaline-2,3 dione (CNQX) did not affect the percent of SP-ir cells in the developing tectum. Treatment with d-(-)-2-amino-5-phosphonovaleric acid (d-AP-5) was also ineffective in this regard, although both it and CNQX treatment disrupted visual map topography. Chronic treatment with brain-derived neurotrophic factor (BDNF) and neurotrophin-4/5 (NT-4/5) produced increases in SP-ir cells in the treated lobes of normal animals, which were significant in the case of NT-4/5. Both substances also prevented the decrease of SP cells that would otherwise occur in the deafferented lobe of unilaterally optic nerve-transected tadpoles. These changes in the percent of SP-ir cells occurred without any detectable changes in the overall number of tectal cells. NGF had no effect on SP expression. Nor did it affect topographic map formation, which was disrupted by treatment with either BDNF or NT-4/5. Our results demonstrate that different mechanisms regulate SP expression in the developing and adult tectum. They indicate that neurotrophin levels in the developing optic tectum may selectively regulate a specific neuropeptide-expressing population of cells.     

用脑光学成像精确测定猫初级视皮层视野拓扑投射关系

利用基于脑内源信号的光学成像和二维互相关分析的方法,对猫初级视皮层17区的视野拓扑离心度(即视网膜-皮层拓扑关系)进行了精确测量。当采用在同一屏幕内处于上下视野的、方位互相垂直的两个相邻光栅刺激时,皮层中一部分区域的绝大部分细胞因同时兴奋而导致方位功能图模糊不清。将这种方位功能图和用单一方位(水平或垂直)全屏光栅刺激所得到的功能图进行比较,通过计算每一像素的互相关系数,从而获得皮层的精确视野拓扑离心度。同时用电生理的方法测量了同一视皮层内的单细胞的感受野位置,证明这种方法得到的视野离心度和光学记录方法得到的相同。因此,本研究为大面积地确定视皮层细胞感受野在视野中的位置提供了一种快速和较准确的方法。     

Responses of medullary neurons to moving visual stimuli in the common toad

The concept of coded 'command releasing systems' proposes that visually specialized descending tectal (and pretectal) neurons converge on motor pattern generating medullary circuits and release--in goal-specific combination--specific action patterns. Extracellular recordings from medullary neurons of the medial reticular formation of the awake immobilized toad in response to moving visual stimuli revealed the following main results. (i) Properties of medullary neurons were distinguished by location, shape, and size of visual receptive fields (ranging from relatively small to wide), by trigger features of various moving configural stimulus objects (including prey- and predator-selective properties), by tactile sensitivity, and by firing pattern characteristics (sluggish, tonic, warming-up, and cyclic). (ii) Visual receptive fields of medullary neurons and their responses to moving configural objects suggest converging inputs of tectal (and pretectal) descending neurons. (iii) In contrast to tectal monocular 'small-field' neurons, the excitatory visual receptive fields of comparable medullary neurons were larger, ellipsoidally shaped, mostly oriented horizontally, and not topographically mapped in an obvious fashion. Furthermore, configural feature discrimination was sharper. (iv) The observation of multiple properties in most medullary neurons (partly showing combined visual and cutaneous sensitivities) suggests integration of various inputs by these cells, and this is in principle consistent with the concept of command releasing systems. (v) There is evidence for reciprocal tectal/medullary excitatory pathways suitable for premotor warming-up. (vi) Cyclic bursting of many neurons, spontaneously or as a post-stimulus sustaining event, points to a medullary premotor/motor property.     

Neuronal NADPH oxidase 2 regulates growth cone guidance downstream of slit2/robo2

NADPH oxidases (Nox) are membrane‐bound multi‐subunit protein complexes producing reactive oxygen species (ROS) that regulate many cellular processes. Emerging evidence suggests that Nox‐derived ROS also control neuronal development and axonal outgrowth. However, whether Nox act downstream of receptors for axonal growth and guidance cues is presently unknown. To answer this question, we cultured retinal ganglion cells (RGCs) derived from zebrafish embryos and exposed these neurons to netrin‐1, slit2, and brain‐derived neurotrophic factor (BDNF). To test the role of Nox in cue‐mediated growth and guidance, we either pharmacologically inhibited Nox or investigated neurons from mutant fish that are deficient in Nox2. We found that slit2‐mediated growth cone collapse, and axonal retraction were eliminated by Nox inhibition. Though we did not see an effect of either BDNF or netrin‐1 on growth rates, growth in the presence of netrin‐1 was reduced by Nox inhibition. Furthermore, attractive and repulsive growth cone turning in response to gradients of BDNF, netrin‐1, and slit2, respectively, were eliminated when Nox was inhibited in vitro. ROS biosensor imaging showed that slit2 treatment increased growth cone hydrogen peroxide levels via mechanisms involving Nox2 activation. We also investigated the possible relationship between Nox2 and slit2/Robo2 signaling in vivo. astray/nox2 double heterozygote larvae exhibited decreased area of tectal innervation as compared to individual heterozygotes, suggesting both Nox2 and Robo2 are required for establishment of retinotectal connections. Our results provide evidence that Nox2 acts downstream of slit2/Robo2 by mediating growth and guidance of developing zebrafish RGC neurons.     

Electrical Stimulation Promotes BDNF Expression in Spinal Cord Neurons Through Ca<Superscript>2+</Superscript>- and Erk-Dependent Signaling Pathways

Brief electrical stimulation has been shown to be effective in promoting neuronal regeneration following peripheral nerve injury. These effects are thought to be mediated largely by the upregulation of the expression of brain-derived neurotrophic factor (BDNF) in spinal cord neurons. However, the molecular mechanisms by which electrical stimulation can promote BDNF expression are not known. The mechanism involved in BDNF expression after electrical stimulation was explored in this study. Immunohistochemistry and Western blotting were used to test BDNF expression. Confocal microscopy was utilized to study intracellular Ca²⁺ volume. Immunohistochemistry and Western blotting confirmed that brief electrical stimulation increased BDNF expression in spinal cord neurons both in vivo and in vitro. Treatment of cultured neurons with nifedipine, an inhibitor of voltage-gated calcium channels, significantly reduced the BDNF increase produced by electrical stimulation, and an inhibitor of Erk completely abolished the effect of electrical stimulation. Levels of BDNF expression in the presence of the Erk inhibitor were lower that in unstimulated and untreated controls, indicating that Erk activation is required to maintain baseline levels of BDNF. Confocal microscopy using a Ca²⁺-sensitive fluorochrome revealed that electrical stimulation is accompanied by an increase in intracellular Ca²⁺ levels; the increase was partly blocked by nifedipine. These findings argue that electrical stimulation increases BDNF expression in spinal cord neurons by activating a Ca²⁺- and Erk-dependent signaling pathways.     

Motion processing with wide-field neurons in the retino-tecto-rotundal pathway

The retino-tecto-rotundal pathway is the main visual pathway in non-mammalian vertebrates and has been found to be highly involved in visual processing. Despite the extensive receptive fields of tectal and rotundal wide-field neurons, pattern discrimination tasks suggest a system with high spatial resolution. In this paper, we address the problem of how global processing performed by motion-sensitive wide-field neurons can be brought into agreement with the concept of a local analysis of visual stimuli. As a solution to this problem, we propose a firing-rate model of the retino-tecto-rotundal pathway which describes how spatiotemporal information can be organized and retained by tectal and rotundal wide-field neurons while processing Fourier-based motion in absence of periodic receptive-field structures. The model incorporates anatomical and electrophysiological experimental data on tectal and rotundal neurons, and the basic response characteristics of tectal and rotundal neurons to moving stimuli are captured by the model cells. We show that local velocity estimates may be derived from rotundal-cell responses via superposition in a subsequent processing step. Experimentally testable predictions which are both specific and characteristic to the model are provided. Thus, a conclusive explanation can be given of how the retino-tecto-rotundal pathway enables the animal to detect and localize moving objects or to estimate its self-motion parameters.     

Binocular depth perception mechanisms in tongue-projecting salamanders

Tongue-projecting salamanders (Bolitoglossini) combine extreme speed and high precision in prey capture. They possess all requirements for stereoscopic depth perception: frontally oriented eyes, a substantial amount of direct ipsilateral projection in addition to the contralateral one, and binocularly driven neurons. Extracellular recordings were made from retinal afferents in the tectum as well as from the somata of tectal neurons. RF-sizes of afferents and tectal neurons were determined, and the response properties of tectal neurons were tested under monocular and binocular conditions with stimuli of different size and velocity. While RF-sizes and response properties of binocular neurons during binocular and contralateral stimulation were similar, ipsilaterally stimulated neurons exhibited much smaller RFs, lower spike rates and different size preferences.Furthermore, the contralateral retinotectal projection from one eye and the ipsilateral from the other are in register. While retinal afferents are distributed linearly over the tectal surface, most tectal neurons are activated by a retinal area corresponding to the frontal visual field; this results in a magnification of this region. The two monocular receptive fields of binocular neurons exhibit zero disparities (horopter) at distances that coincide with the maximum reach of the tongue. We hypothesize that bolitoglossine salamanders (as well as amphibians in general) make use of two kinds of disparities: (1) between the maps in the left and right tectal hemisphere, coding for the lateral eccentricity of an object, and (2) between the ipsilateral and contralateral retinotectal map, coding for the distance. The presence of substantial direct ipsilateral afferents in bolitoglossine salamanders appears to be the basis for a fast computation of object distance, which is characteristic of these animals.Abbreviations Ax/Ay coordinates of a recorded afference - Nx/Ny coordinates of a recorded neuron - RF receptive field - RFc contralateral receptive field - RFi ipsilateral receptive field - RFx/RFy coordinates of a receptive field center - RGC retinal ganglion cell     

Brain-derived neurotrophic factor activates ERK5 in cortical neurons via a Rap1-MEKK2 signaling cascade

The extracellular signal-regulated kinase 5 (ERK5) is activated in neurons of the central nervous system by neurotrophins including brain-derived neurotrophic factor (BDNF). Although MEK5 is known to mediate BDNF stimulation of ERK5 in central nervous system neurons, other upstream signaling components have not been identified. Here, we report that BDNF induces a sustained activation of ERK5 in rat cortical neurons and activates Rap1, a small GTPase, as well as MEKK2, a MEK5 kinase. Our data indicate that activation of Rap1 or MEKK2 is sufficient to stimulate ERK5, whereas inhibition of either Rap1 or MEKK2 attenuates BDNF activation of ERK5. Furthermore, BDNF stimulation of MEKK2 is regulated by Rap1. Our evidence also indicates that Ras and MEKK3, a MEK5 kinase in non-neuronal cells, do not play a significant role in BDNF activation of ERK5. This study identifies Rap1 and MEKK2 as critical upstream signaling molecules mediating BDNF stimulation of ERK5 in central nervous system neurons.     

Responses of tectal neurons to contrasting stimuli: an electrophysiological study in the barn owl

The saliency of visual objects is based on the center to background contrast. Particularly objects differing in one feature from the background may be perceived as more salient. It is not clear to what extent this so called "pop-out" effect observed in humans and primates governs saliency perception in non-primates as well. In this study we searched for neural-correlates of pop-out perception in neurons located in the optic tectum of the barn owl. We measured the responses of tectal neurons to stimuli appearing within the visual receptive field, embedded in a large array of additional stimuli (the background). Responses were compared between contrasting and uniform conditions. In a contrasting condition the center was different from the background while in the uniform condition it was identical to the background. Most tectal neurons responded better to stimuli in the contrsating condition compared to the uniform condition when the contrast between center and background was the direction of motion but not when it was the orientation of a bar. Tectal neurons also preferred contrasting over uniform stimuli when the center was looming and the background receding but not when the center was receding and the background looming. Therefore, our results do not support the hypothesis that tectal neurons are sensitive to pop-out per-se. The specific sensitivity to the motion contrasting stimulus is consistent with the idea that object motion and not large field motion (e.g., self-induced motion) is coded in the neural responses of tectal neurons.     

Spike timing-dependent LTP/LTD mediates visual experience-dependent plasticity in a developing retinotectal system

Sensory experience plays an instructive role in the development of the nervous system. Here we showed that visual experience can induce persistent modification of developing retinotectal circuits via spike timing-dependent plasticity (STDP). Pairing light stimuli with spiking of the tectal cell induced persistent enhancement or reduction of light-evoked responses, with a dependence on the relative timing between light stimulus and postsynaptic spiking similar to that for STDP. Using precisely timed sequential three-bar stimulation to mimic a moving bar, we showed that spike timing-dependent LTP/LTD can account for the asymmetric modification of the tectal cell receptive field induced by moving bar. Furthermore, selective inhibition of signaling mediated by brain-derived neurotrophic factor and nitric oxide, which are respectively required for light-induced LTP and LTD, interfered with moving bar-induced temporally specific changes in the tectal cell responses. Together, these findings suggest that STDP can mediate sensory experience-dependent circuit refinement in the developing nervous system.     

The excitoprotective effect of N-methyl-D-aspartate receptors is mediated by a brain-derived neurotrophic factor autocrine loop in cultured hippocampal neurons

The neuroprotective effect and molecular mechanisms underlying preconditioning with N-methyl-D-aspartate (NMDA) in cultured hippocampal neurons have not been described. Pre-incubation with subtoxic concentrations of the endogenous neurotransmitter glutamate protects vulnerable neurons against NMDA receptor-mediated excitotoxicity. As a result of physiological preconditioning, NMDA significantly antagonizes the neurotoxicity resulting from subsequent exposure to an excitotoxic concentration of glutamate. The protective effect of glutamate or NMDA is time- and concentration-dependent, suggesting that sufficient agonist and time are required to establish an intracellular neuroprotective state. In these cells, the TrkB ligand, brain-derived neurotrophic factor (BDNF) attenuates glutamate toxicity. Therefore, we tested the hypothesis that NMDA protects neurons via a BDNF-dependent mechanism. Exposure of hippocampal cultures to a neuroprotective concentration of NMDA (50 microM) evoked the release of BDNF within 2 min without attendant changes in BDNF protein or gene expression. The accumulated increase of BDNF in the medium is followed by an increase in the phosphorylation (activation) of TrkB receptors and a later increase in exon 4-specific BDNF mRNA. The neuroprotective effect of NMDA was attenuated by pre-incubation with a BDNF-blocking antibody and TrkB-IgG, a fusion protein known to inhibit the activity of extracellular BDNF, suggesting that BDNF plays a major role in NMDA-mediated survival. These results demonstrate that low level stimulation of NMDA receptors protect neurons against glutamate excitotoxicity via a BDNF autocrine loop in hippocampal neurons and suggest that activation of neurotrophin signaling pathways plays a key role in the neuroprotection of NMDA.