Platelet-derived growth factor receptor (PDGFRA)/NG2-expressing glia are distributed throughout the adult CNS. They are descended from oligodendrocyte precursors (OLPs) in the perinatal CNS, but it is not clear whether they continue to generate myelinating oligodendrocytes or other differentiated cells during normal adult life. We followed the fates of adult OLPs in Pdgfra-creER T2 /Rosa26-YFP double-transgenic mice and found that they generated many myelinating oligodendrocytes during adulthood; >20% of all oligodendrocytes in the adult mouse corpus callosum were generated after 7 weeks of age, raising questions about the function of the late-myelinating axons. OLPs also produced some myelinating cells in the cortex, but the majority of adult-born cortical cells did not appear to myelinate. We found no evidence for astrocyte production in gray or white matter. However, small numbers of projection neurons were generated in the forebrain, especially in the piriform cortex, which is the main target of the olfactory bulb.Oligodendrocytes, the myelin-forming cells of the CNS, are mostly generated during the first few postnatal weeks in rodents, peaking in the second week (postnatal day 7-14, P7-P14). They differentiate from proliferative, migratory OLPs that originate in the ventricular zones of the developing spinal cord and brain. OLPs express a characteristic set of markers, including PDGFRA and the NG2 proteoglycan, allowing OLP development to be followed in situ. Both PDGFRA and NG2 are rapidly downregulated when OLPs differentiate into oligodendrocytes 1-3 , unlike other lineage markers (for example, transcription factors SOX10 and OLIG2), which are expressed in both OLPs and oligodendrocytes. By the time of birth, OLPs are more or less evenly distributed throughout the brain, both in gray matter and developing white matter, and remain so during the early postnatal period, when oligodendrocyte production is in full swing. The size of the OLP population remains relatively stable throughout this time, presumably because, although OLPs continue to proliferate, half of the daughter cells either differentiate or die. To determine the behavior and fates of adult OLPs in vivo, we generated a transgenic mouse line that expresses a tamoxifen-inducible form of the Cre recombinase (CreER T2 ) under PDGFRA transcriptional control in a phage artificial chromosome (PAC). The PAC transgene was faithfully expressed by OLPs in the postnatal CNS. By combining PdgfracreER T2 mice with the Rosa26-YFP reporter line, we were able to induce expression of yellow fluorescent protein (YFP) de novo in adult OLPs and identify their differentiated progeny. We found that OLPs generated mature, myelinating oligodendrocytes in adult mice until at least 8 months of age, raising questions about the function of the newly myelinated axons. We were unable to find any evidence for astrocyte production from adult OLPs in either gray or white matter. Notably, we found small numbers of YFP-labeled neurons in the forebrain, particularly in th...

After central nervous system (CNS) demyelination-such as occurs during multiple sclerosis-there is often spontaneous regeneration of myelin sheaths, mainly by oligodendrocytes but also by Schwann cells. The origins of the remyelinating cells have not previously been established. We have used Cre-lox fate mapping in transgenic mice to show that PDGFRA/NG2-expressing glia, a distributed population of stem/progenitor cells in the adult CNS, produce the remyelinating oligodendrocytes and almost all of the Schwann cells in chemically induced demyelinated lesions. In contrast, the great majority of reactive astrocytes in the vicinity of the lesions are derived from preexisting FGFR3-expressing cells, likely to be astrocytes. These data resolve a long-running debate about the origins of the main players in CNS remyelination and reveal a surprising capacity of CNS precursors to generate Schwann cells, which normally develop from the embryonic neural crest and are restricted to the peripheral nervous system.

No abstract

Summary At various stages of the visual system, visual responses are modulated by arousal. Here, we find that in mice this modulation operates as early as in the first synapse from the retina and even in retinal axons. To measure retinal activity in the awake, intact brain, we imaged the synaptic boutons of retinal axons in the superior colliculus. Their activity depended not only on vision but also on running speed and pupil size, regardless of retinal illumination. Arousal typically reduced their visual responses and selectivity for direction and orientation. Recordings from retinal axons in the optic tract revealed that arousal modulates the firing of some retinal ganglion cells. Arousal had similar effects postsynaptically in colliculus neurons, independent of activity in the other main source of visual inputs to the colliculus, the primary visual cortex. These results indicate that arousal modulates activity at every stage of the mouse visual system.

In order to link our knowledge of single neurons with theories of network function, it has been a long-standing goal to manipulate the activity and gene expression of identified subsets of mammalian neurons within the intact brain in vivo. This protocol describes a method for delivering plasmid DNA into single identified mammalian neurons in vivo, by combining two-photon imaging with single-cell electroporation. Surgery, mounting of a chronic recording chamber and targeted electroporation of identified neurons can be performed within 1-2 h. Stable transgene expression can reliably be induced with high success rates both in single neurons as well as in small, spatially defined networks of neurons in the cerebral cortex of rodents.

Summary Age-related macular degeneration and other macular diseases result in the loss of light-sensing cone photoreceptors, causing irreversible sight impairment. Photoreceptor replacement may restore vision by transplanting healthy cells, which must form new synaptic connections with the recipient retina. Despite recent advances, convincing evidence of functional connectivity arising from transplanted human cone photoreceptors in advanced retinal degeneration is lacking. Here, we show restoration of visual function after transplantation of purified human pluripotent stem cell-derived cones into a mouse model of advanced degeneration. Transplanted human cones elaborate nascent outer segments and make putative synapses with recipient murine bipolar cells (BCs), which themselves undergo significant remodeling. Electrophysiological and behavioral assessments demonstrate restoration of surprisingly complex light-evoked retinal ganglion cell responses and improved light-evoked behaviors in treated animals. Stringent controls exclude alternative explanations, including material transfer and neuroprotection. These data provide crucial validation for photoreceptor replacement therapy and for the potential to rescue cone-mediated vision.

BackgroundThe use of human pluripotent stem cell-derived retinal cells for cell therapy strategies and disease modelling relies on the ability to obtain healthy and organised retinal tissue in sufficient quantities. Generating such tissue is a lengthy process, often taking over 6 months of cell culture, and current approaches do not always generate large quantities of the major retinal cell types required.MethodsWe adapted our previously described differentiation protocol to investigate the use of stirred-tank bioreactors. We used immunohistochemistry, flow cytometry and electron microscopy to characterise retinal organoids grown in standard and bioreactor culture conditions.ResultsOur analysis revealed that the use of bioreactors results in improved laminar stratification as well as an increase in the yield of photoreceptor cells bearing cilia and nascent outer-segment-like structures.ConclusionsBioreactors represent a promising platform for scaling up the manufacture of retinal cells for use in disease modelling, drug screening and cell transplantation studies.Electronic supplementary materialThe online version of this article (10.1186/s13287-018-0907-0) contains supplementary material, which is available to authorized users.

The rd1 mouse with a mutation in the Pde6b gene was the first strain of mice identified with a retinal degeneration. However, AAV-mediated gene supplementation of rd1 mice only results in structural preservation of photoreceptors, and restoration of the photoreceptor-mediated a-wave, but not in restoration of the bipolar cell-mediated b-wave. Here we show that a mutation in Gpr179 prevents the full restoration of vision in rd1 mice. Backcrossing rd1 with C57BL6 mice reveals the complete lack of b-wave in a subset of mice, consistent with an autosomal recessive Mendelian inheritance pattern. We identify a mutation in the Gpr179 gene, which encodes for a G-protein coupled receptor localized to the dendrites of ON-bipolar cells. Gene replacement in rd1 mice that are devoid of the mutation in Gpr179 successfully restores the function of both photoreceptors and bipolar cells, which is maintained for up to 13 months. Our discovery may explain the failure of previous gene therapy attempts in rd1 mice, and we propose that Grp179 mutation status should be taken into account in future studies involving rd1 mice.