FGF22 signaling regulates synapse formation during post‐injury remodeling of the spinal cord

The remodeling of axonal circuits after injury requires the formation of new synaptic contacts to enable functional recovery. Which molecular signals initiate such axonal and synaptic reorganisation in the adult central nervous system is currently unknown. Here, we identify FGF22 as a key regulator of circuit remodeling in the injured spinal cord. We show that FGF22 is produced by spinal relay neurons, while its main receptors FGFR1 and FGFR2 are expressed by cortical projection neurons. FGF22 deficiency or the targeted deletion of FGFR1 and FGFR2 in the hindlimb motor cortex limits the formation of new synapses between corticospinal collaterals and relay neurons, delays their molecular maturation, and impedes functional recovery in a mouse model of spinal cord injury. These results establish FGF22 as a synaptogenic mediator in the adult nervous system and a crucial regulator of synapse formation and maturation during post‐injury remodeling in the spinal cord.SynopsisFollowing spinal cord injury, transected projections form detour circuits that circumvent the lesion and contribute to functional recovery. The formation of new synaptic contacts is a crucial step of the process, but its molecular regulation is currently not understood. Members of the FGF family can promote synapse formation during nervous system development, suggesting that they might have a similar function in the injured adult CNS. Here, we show that:FGF22 and FGF22 receptors are expressed in the adult nervous system.FGF22 deficiency or deletion of FGF22 receptors restricts the formation and maturation of new synapses in the injured spinal cord.Genetic disruption of FGF22 signaling impedes spontaneous functional recovery following spinal cord injury.FGF22 is a synaptogenic mediator in the adult nervous system and promotes synaptic plasticity and circuit remodeling in a mouse model of spinal cord injury.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/111756/1/embj201490578-sup-0001-Suppl_Info.pdfhttp://deepblue.lib.umich.edu/bitstream/2027.42/111756/2/embj201490578.pdfhttp://deepblue.lib.umich.edu/bitstream/2027.42/111756/3/embj201490578.reviewer_comments.pd

Spontaneous activity promotes synapse formation in a cell-type-dependent manner in the developing retina

Spontaneous activity is thought to regulate synaptogenesis in many parts of the developing nervous system. In vivo evidence for this regulation, however, is scarce and comes almost exclusively from experiments in which normal activity was reduced or blocked completely. Thus, whether spontaneous activity itself promotes synaptogenesis or plays a purely permissive role, remains uncertain. In addition, how activity influences synapse dynamics to shape connectivity and whether its effects among neurons are uniform or cell type-dependent is unclear. In mice lacking the cone-rod homeobox gene (Crx), photoreceptors (PRs) fail to establish normal connections with bipolar cells (BCs). Here, we find that retinal ganglion cells (RGCs) in Crx−/− mice become rhythmically hyperactive around the time of eye-opening; as a result of increased spontaneous glutamate release from BCs. This elevated neurotransmission enhances synaptogenesis between BCs and RGCs, without altering the overall circuit architecture. Using live imaging, we discover that spontaneous activity selectively regulates the rate of synapse formation, not elimination, in this circuit. Reconstructions of the connectivity patterns of three BC types with a shared RGC target further revealed that neurotransmission specifically promotes the formation of multisynaptic appositions from one BC type without affecting the maintenance or elimination of connections from the other two. While hyperactivity in Crx−/− mice persists, synapse numbers do not increase beyond four weeks of age, suggesting closure of a critical period for synaptic refinement in the inner retina. Interestingly, despite their hyperactivity, RGC axons maintain normal eye-specific territories and cell type-specific layers in the dorsolateral geniculate nucleus (dLGN)

Review: ‘Gimme five’: future challenges in multiple sclerosis. ECTRIMS Lecture 2009

This article is based on the ECTRIMS lecture given at the 25th ECTRIMS meeting which was held in Düsseldorf, Germany, from 9 to 12 September 2009. Five challenges have been identified: (1) safeguarding the principles of medical ethics; (2) optimizing the risk/benefit ratio; (3) bridging the gap between multiple sclerosis and experimental autoimmune encephalitis; (4) promoting neuroprotection and repair; and (5) tailoring multiple sclerosis therapy to the individual patient. Each of these challenges will be discussed and placed in the context of current research into the pathogenesis and treatment of multiple sclerosis

An allosteric regulator of R7-RGS proteins influences light-evoked activity and glutamatergic waves in the inner retina

In the outer retina, G protein-coupled receptor (GPCR) signaling mediates phototransduction and synaptic transmission between photoreceptors and ON bipolar cells. In contrast, the functions of modulatory GPCR signaling networks in the inner retina are less well understood. We addressed this question by determining the consequences of augmenting modulatory Gi/o signaling driven by endogenous transmitters. This was done by analyzing the effects of genetically ablating the R7 RGS-binding protein (R7BP), a membrane-targeting protein and positive allosteric modulator of R7-RGS (regulator of the G protein signaling 7) family that deactivates Gi/oα subunits. We found that R7BP is expressed highly in starburst amacrine cells and retinal ganglion cells (RGCs). As indicated by electroretinography and multielectrode array recordings of adult retina, ablation of R7BP preserved outer retina function, but altered the firing rate and latency of ON RGCs driven by rods and cones but not rods alone. In developing retina, R7BP ablation increased the burst duration of glutamatergic waves whereas cholinergic waves were unaffected. This effect on glutamatergic waves did not result in impaired segregation of RGC projections to eye-specific domains of the dorsal lateral geniculate nucleus. R7BP knockout mice exhibited normal spatial contrast sensitivity and visual acuity as assessed by optomotor reflexes. Taken together these findings indicate that R7BP-dependent regulation of R7-RGS proteins shapes specific aspects of light-evoked and spontaneous activity of RGCs in mature and developing retina

Remodeling of Axonal Connections Contributes to Recovery in an Animal Model of Multiple Sclerosis

In multiple sclerosis (MS), inflammation in the central nervous system (CNS) leads to damage of axons and myelin. Early during the clinical course, patients can compensate this damage, but little is known about the changes that underlie this improvement of neurological function. To study axonal changes that may contribute to recovery, we made use of an animal model of MS, which allows us to target inflammatory lesions to the corticospinal tract (CST), a major descending motor pathway. We demonstrate that axons remodel at multiple levels in response to a single neuroinflammatory lesion as follows: (a) surrounding the lesion, local interneurons show regenerative sprouting; (b) above the lesion, descending CST axons extend new collaterals that establish a “detour” circuit to the lumbar target area, whereas below the lesion, spared CST axons increase their terminal branching; and (c) in the motor cortex, the distribution of projection neurons is remodeled, and new neurons are recruited to the cortical motor pool. Behavioral tests directly show the importance of these changes for recovery. This paper provides evidence for a highly plastic response of the motor system to a single neuroinflammatory lesion. This framework will help to understand the endogenous repair capacity of the CNS and to develop therapeutic strategies to support it

Mechanisms of action of Methylthioadenosine: pathways implicated in neuroprotection in models of Multiple Sclerosis and other neurological diseases

From 5th European Workshop on Immune-Mediated Inflammatory Diseases (Sitges-Barcelona, Spain. 1-3 December 2010)Background Methylthioadenosine (MTA) has anti-oxidant and anti-proliferative properties and was shown to induce cell protection in hepatic cells. We previously demonstrated that exert immunomodulatory and neuroprotective effects in the animal model of Multiple Sclerosis (MS) and other neurological diseases like Parkinson disease, stroke and Epilepsy. Objective To study the mechanisms of action and different pathways implicated in the neuroprotective effect of MTA in neurological diseases. Methods RN22 (Schwnoma cell line) and PC12 (Pheochromocytoma cell line) were used to test the neuroprotective activity of MTA against stress in RN22 and to differentiate neurites in PC12. BV2 cells were used to test the effect of MTA in microglia. Organotypic cerebellum cultures were used to determine MTA effect in demyelination/remyelination. Luminex technology, western blot and ELISA were used in order to study the phosphorylated state of different pathways (AkT/PKB, ERK/MAPK, P38/SAPK or STAT3) and to determine the amount of different cytokines (IL-1β and TNF-α). Ros determination was also done by fluorescence determination. Results In vitro studies revealed that MTA protection against different stresses and its capacity to differentiate neurites implies pathways like ERK/MAPK, P38/SAPK or STAT3. MTA neuroprotective capacity is also related with its ability to reduce ROS production and oxidative stress. MTA was shown to protect against demyelination in cerebellum organotypic cultures treated with LPS or Lysolecithin. Conclusions MTA is neuroprotective in models of MS, Parkinson disease, stroke or Epilepsy. This neuroprotective effect depends on its capacity to protect against demyelination, its anti-oxidant effect and the activation of pathways related with protection against stress and production of neurite differentiation

Modeling-independent elucidation of inactivation pathways in recombinant and native A-type Kv channels.

A-type voltage-gated K(+) (Kv) channels self-regulate their activity by inactivating directly from the open state (open-state inactivation [OSI]) or by inactivating before they open (closed-state inactivation [CSI]). To determine the inactivation pathways, it is often necessary to apply several pulse protocols, pore blockers, single-channel recording, and kinetic modeling. However, intrinsic hurdles may preclude the standardized application of these methods. Here, we implemented a simple method inspired by earlier studies of Na(+) channels to analyze macroscopic inactivation and conclusively deduce the pathways of inactivation of recombinant and native A-type Kv channels. We investigated two distinct A-type Kv channels expressed heterologously (Kv3.4 and Kv4.2 with accessory subunits) and their native counterparts in dorsal root ganglion and cerebellar granule neurons. This approach applies two conventional pulse protocols to examine inactivation induced by (a) a simple step (single-pulse inactivation) and (b) a conditioning step (double-pulse inactivation). Consistent with OSI, the rate of Kv3.4 inactivation (i.e., the negative first derivative of double-pulse inactivation) precisely superimposes on the profile of the Kv3.4 current evoked by a single pulse because the channels must open to inactivate. In contrast, the rate of Kv4.2 inactivation is asynchronous, already changing at earlier times relative to the profile of the Kv4.2 current evoked by a single pulse. Thus, Kv4.2 inactivation occurs uncoupled from channel opening, indicating CSI. Furthermore, the inactivation time constant versus voltage relation of Kv3.4 decreases monotonically with depolarization and levels off, whereas that of Kv4.2 exhibits a J-shape profile. We also manipulated the inactivation phenotype by changing the subunit composition and show how CSI and CSI combined with OSI might affect spiking properties in a full computational model of the hippocampal CA1 neuron. This work unambiguously elucidates contrasting inactivation pathways in neuronal A-type Kv channels and demonstrates how distinct pathways might impact neurophysiological activity

An assay to image neuronal microtubule dynamics in mice

Microtubule dynamics in neurons play critical roles in physiology, injury and disease and determine microtubule orientation, the cell biological correlate of neurite polarization. Several microtubule binding proteins, including end-binding protein 3 (EB3), specifically bind to the growing plus tip of microtubules. In the past, fluorescently tagged end-binding proteins have revealed microtubule dynamics in vitro and in non-mammalian model organisms. Here, we devise an imaging assay based on transgenic mice expressing yellow fluorescent protein-tagged EB3 to study microtubules in intact mammalian neurites. Our approach allows measurement of microtubule dynamics in vivo and ex vivo in peripheral nervous system and central nervous system neurites under physiological conditions and after exposure to microtubule-modifying drugs. We find an increase in dynamic microtubules after injury and in neurodegenerative disease states, before axons show morphological indications of degeneration or regrowth. Thus increased microtubule dynamics might serve as a general indicator of neurite remodelling in health and disease

Induction of Cell Stress in Neurons from Transgenic Mice Expressing Yellow Fluorescent Protein: Implications for Neurodegeneration Research

Peer reviewedPublisher PD

Adaptive Movement Compensation for In Vivo Imaging of Fast Cellular Dynamics within a Moving Tissue

In vivo non-linear optical microscopy has been essential to advance our knowledge of how intact biological systems work. It has been particularly enabling to decipher fast spatiotemporal cellular dynamics in neural networks. The power of the technique stems from its optical sectioning capability that in turn also limits its application to essentially immobile tissue. Only tissue not affected by movement or in which movement can be physically constrained can be imaged fast enough to conduct functional studies at high temporal resolution. Here, we show dynamic two-photon Ca2+ imaging in the spinal cord of a living rat at millisecond time scale, free of motion artifacts using an optical stabilization system. We describe a fast, non-contact adaptive movement compensation approach, applicable to rough and weakly reflective surfaces, allowing real-time functional imaging from intrinsically moving tissue in live animals. The strategy involves enslaving the position of the microscope objective to that of the tissue surface in real-time through optical monitoring and a closed feedback loop. The performance of the system allows for efficient image locking even in conditions of random or irregular movements