Oligodendrocytes make myelin and support axons metabolically with lactate. However, it is unknown how glucose utilization and glycolysis are adapted to the different axonal energy demands. Spiking axons release glutamate and oligodendrocytes express NMDA receptors of unknown function. Here we show that the stimulation of oligodendroglial NMDA receptors mobilizes glucose transporter GLUT1, leading to its incorporation into the myelin compartment in vivo. When myelinated optic nerves from conditional NMDA receptor mutants are challenged with transient oxygen-glucose deprivation, they show a reduced functional recovery when returned to oxygen-glucose but are indistinguishable from wild-type when provided with oxygen-lactate. Moreover, the functional integrity of isolated optic nerves, which are electrically silent, is extended by preincubation with NMDA, mimicking axonal activity, and shortened by NMDA receptor blockers. This reveals a novel aspect of neuronal energy metabolism in which activity-dependent glutamate release enhances oligodendroglial glucose uptake and glycolytic support of fast spiking axons.

In several neurodegenerative diseases and myelin disorders, the degeneration profiles of myelinated axons are compatible with underlying energy deficits. However, it is presently impossible to measure selectively axonal ATP levels in the electrically active nervous system. We combined transgenic expression of an ATP-sensor in neurons of mice with confocal FRET imaging and electrophysiological recordings of acutely isolated optic nerves. This allowed us to monitor dynamic changes and activity-dependent axonal ATP homeostasis at the cellular level and in real time. We find that changes in ATP levels correlate well with compound action potentials. However, this correlation is disrupted when metabolism of lactate is inhibited, suggesting that axonal glycolysis products are not sufficient to maintain mitochondrial energy metabolism of electrically active axons. The combined monitoring of cellular ATP and electrical activity is a novel tool to study neuronal and glial energy metabolism in normal physiology and in models of neurodegenerative disorders.DOI: http://dx.doi.org/10.7554/eLife.24241.001

Proteome and transcriptome analyses aim at comprehending the molecular profiles of the brain, its cell-types and subcellular compartments including myelin. Despite the relevance of the peripheral nervous system for normal sensory and motor capabilities, analogous approaches to peripheral nerves and peripheral myelin have fallen behind evolving technical standards. Here we assess the peripheral myelin proteome by gel-free, label-free mass-spectrometry for deep quantitative coverage. Integration with RNA-Sequencing-based developmental mRNA-abundance profiles and neuropathy disease genes illustrates the utility of this resource. Notably, the periaxin-deficient mouse model of the neuropathy Charcot-Marie-Tooth 4F displays a highly pathological myelin proteome profile, exemplified by the discovery of reduced levels of the monocarboxylate transporter MCT1/SLC16A1 as a novel facet of the neuropathology. This work provides the most comprehensive proteome resource thus far to approach development, function and pathology of peripheral myelin, and a straightforward, accurate and sensitive workflow to address myelin diversity in health and disease.

Myelination of axons facilitates rapid impulse propagation in the nervous system. The axon/myelin-unit becomes impaired in myelin-related disorders and upon normal aging. However, the molecular cause of many pathological features, including the frequently observed myelin outfoldings, remained unknown. Using label-free quantitative proteomics, we find that the presence of myelin outfoldings correlates with a loss of cytoskeletal septins in myelin. Regulated by phosphatidylinositol-(4,5)-bisphosphate (PI(4,5)P2)-levels, myelin septins (SEPT2/SEPT4/SEPT7/SEPT8) and the PI(4,5)P2-adaptor anillin form previously unrecognized filaments that extend longitudinally along myelinated axons. By confocal microscopy and immunogold-electron microscopy, these filaments are localized to the non-compacted adaxonal myelin compartment. Genetic disruption of these filaments in Sept8-mutant mice causes myelin outfoldings as a very specific neuropathology. Septin filaments thus serve an important function in scaffolding the axon/myelin-unit, evidently a late stage of myelin maturation. We propose that pathological or aging-associated diminishment of the septin/anillin-scaffold causes myelin outfoldings that impair the normal nerve conduction velocity.DOI: http://dx.doi.org/10.7554/eLife.17119.001

Myelin membranes are dominated by lipids while the complexity of their protein composition has long been considered to be low. However, numerous additional myelin proteins have been identified since. Here we revisit the proteome of myelin biochemically purified from the brains of healthy c56Bl/6N-mice utilizing complementary proteomic approaches for deep qualitative and quantitative coverage. By gel-free, label-free mass spectrometry, the most abundant myelin proteins PLP, MBP, CNP, and MOG constitute 38, 30, 5, and 1% of the total myelin protein, respectively. The relative abundance of myelin proteins displays a dynamic range of over four orders of magnitude, implying that PLP and MBP have overshadowed less abundant myelin constituents in initial gel-based approaches. By comparisons with published datasets we evaluate to which degree the CNS myelin proteome correlates with the mRNA and protein abundance profiles of myelin and oligodendrocytes. Notably, the myelin proteome displays only minor changes if assessed after a post-mortem delay of 6 h. These data provide the most comprehensive proteome resource of CNS myelin so far and a basis for addressing proteomic heterogeneity of myelin in mouse models and human patients with white matter disorders.

Deficiency of the major constituent of central nervous system (CNS) myelin, proteolipid protein (PLP), causes axonal pathology in spastic paraplegia type-2 patients and in Plp1(null) -mice but is compatible with almost normal myelination. These observations led us to speculate that PLP's role in myelination may be partly compensated for by other tetraspan proteins. Here, we demonstrate that the abundance of the structurally related tetraspanin-2 (TSPAN2) is highly increased in CNS myelin of Plp1(null) -mice. Unexpectedly, Tspan2(null) -mutant mice generated by homologous recombination in embryonic stem cells displayed low-grade activation of astrocytes and microglia in white matter tracts while they were fully myelinated and showed no signs of axonal degeneration. To determine overlapping functions of TSPAN2 and PLP, Tspan2(null) *Plp1(null) double-mutant mice were generated. Strikingly, the activation of astrocytes and microglia was strongly enhanced in Tspan2(null) *Plp1(null) double-mutants compared with either single-mutant, but the levels of dysmyelination and axonal degeneration were not increased. In this model, glial activation is thus unlikely to be caused by axonal pathology, and vice versa does not potentiate axonal degeneration. Our results support the concept that multiple myelin proteins have distinct roles in the long-term preservation of a healthy CNS, rather than in myelination per se.

In neuroinflammatory disorders such as multiple sclerosis, the physiological function of the blood-brain barrier (BBB) is perturbed, particularly in demyelinating lesions and supposedly secondary to acute demyelinating pathology. Using the toxic non-inflammatory cuprizone model of demyelination, we demonstrate, however, that the onset of persistent BBB impairment precedes demyelination. In addition to a direct effect of cuprizone on endothelial cells, a plethora of inflammatory mediators, which are mainly of astroglial origin during the initial disease phase, likely contribute to the destabilization of endothelial barrier function in vivo. Our study reveals that, at different time points of pathology and in different CNS regions, the level of gliosis correlates with the extent of BBB hyperpermeability and edema. Furthermore, in mutant mice with abolished type 3 CXC chemokine receptor (CXCR3) signaling, inflammatory responses are dampened and BBB dysfunction ameliorated. Together, these data have implications for understanding the role of BBB permeability in the pathogenesis of demyelinating disease.

In contrast to acute peripheral nerve injury, the molecular response of Schwann cells in chronic neuropathies remains poorly understood. Onion bulb structures are a pathological hallmark of demyelinating neuropathies, but the nature of these formations is unknown. Here, we show that Schwann cells induce the expression of Neuregulin-1 type I (NRG1-I), a paracrine growth factor, in various chronic demyelinating diseases. Genetic disruption of Schwann cell-derived NRG1 signalling in a mouse model of Charcot-Marie-Tooth Disease 1A (CMT1A), suppresses hypermyelination and the formation of onion bulbs. Transgenic overexpression of NRG1-I in Schwann cells on a wildtype background is sufficient to mediate an interaction between Schwann cells via an ErbB2 receptor-MEK/ERK signaling axis, which causes onion bulb formations and results in a peripheral neuropathy reminiscent of CMT1A. We suggest that diseased Schwann cells mount a regeneration program that is beneficial in acute nerve injury, but that overstimulation of Schwann cells in chronic neuropathies is detrimental.