Percentage Of Myelinated Axons Research Articles

Early microglial response, myelin deterioration and lethality in mice deficient for very long chain ceramide synthesis in oligodendrocytes.

The sphingolipids galactosylceramide (GalCer), sulfatide (ST) and sphingomyelin (SM) are essential for myelin stability and function. GalCer and ST are synthesized mostly from C22-C24 ceramides, generated by Ceramide Synthase 2 (CerS2). To clarify the requirement for C22-C24 sphingolipid synthesis in myelin biosynthesis and stability, we generated mice lacking CerS2 specifically in myelinating cells (CerS2ΔO/ΔO ). At 6 weeks of age, normal-appearing myelin had formed in CerS2ΔO/ΔO mice, however there was a reduction in myelin thickness and the percentage of myelinated axons. Pronounced loss of C22-C24 sphingolipids in myelin of CerS2ΔO/ΔO mice was compensated by greatly increased levels of C18 sphingolipids. A distinct microglial population expressing high levels of activation and phagocytic markers such as CD64, CD11c, MHC class II, and CD68 was apparent at 6 weeks of age in CerS2ΔO/ΔO mice, and had increased by 10 weeks. Increased staining for denatured myelin basic protein was also apparent in 6-week-old CerS2ΔO/ΔO mice. By 16 weeks, CerS2ΔO/ΔO mice showed pronounced myelin atrophy, motor deficits, and axon beading, a hallmark of axon stress. 90% of CerS2ΔO/ΔO mice died between 16 and 26 weeks of age. This study highlights the importance of sphingolipid acyl chain length for the structural integrity of myelin, demonstrating how a modest reduction in lipid chain length causes exposure of a denatured myelin protein epitope and expansion of phagocytic microglia, followed by axon pathology, myelin degeneration, and motor deficits. Understanding the molecular trigger for microglial activation should aid the development of therapeutics for demyelinating and neurodegenerative diseases.

Nuclear Inhibitor of Protein Phosphatase 1 (NIPP1) Regulates CNS Tau Phosphorylation and Myelination During Development.

Nuclear inhibitor of protein phosphatase 1 (NIPP1) is a known regulator of gene expression and plays roles in many physiological or pathological processes such as stem cell proliferation and skin inflammation. While NIPP1 has many regulatory roles in proliferating cells, its function in the central nervous system (CNS) has not been directly investigated. In the present study, we examined NIPP1 CNS function using a conditional knockout (cKO) mouse model in which the Nipp1 gene is excised from neural precursor cells. These mice exhibited severe developmental impairments that led to premature lethality. To delineate the neurological changes occurring in these animals, we first assessed microtubule-associated protein tau, a known target of NIPP1 activity. We found that phosphorylation of tau is significantly enhanced in NIPP1 cKO mice. Consistent with this, we found altered AKT and PP1 activity in NIPP1 cKO mice, suggesting that increased tau phosphorylation likely results from a shift in kinase/phosphatase activity. Secondly, we observed tremors in the NIPP1 cKO mice which prompted us to explore the integrity of the myelin sheath, an integral structure for CNS function. We demonstrated that in NIPP1 cKO mice, there is a significant decrease in MBP protein expression in the cortex, along with deficits in both the conduction of compound action potentials (CAP) and the percentage of myelinated axons in the optic nerve. Our study suggests that NIPP1 in neural precursor cells regulates phosphorylation of tau and CNS myelination and may represent a novel therapeutic target for neurodegenerative diseases.

H-ferritin expression in astrocytes is necessary for proper oligodendrocyte development and myelination.

How iron is delivered to the CNS for myelination is poorly understood. Astrocytes are the most abundant glial cells in the brain and are the only cells in close contact with blood vessels. Therefore, they are strategically located to obtain nutrients, such as iron, from circulating blood. To determine the importance of astrocyte iron uptake and storage in myelination and remyelination, we conditionally knocked-out the expression of the divalent metal transporter 1 (DMT1), the transferrin receptor 1 (Tfr1), and the ferritin heavy subunit (Fth) in Glast-1-positive astrocytes. DMT1 or Tfr1 ablation in astrocytes throughout early brain development did not significantly affects oligodendrocyte maturation or iron homeostasis. However, blocking Fth production in astrocytes during the first postnatal week drastically delayed oligodendrocyte development and myelin synthesis. Fth knockout animals presented an important decrease in the number of myelinating oligodendrocytes and a substantial reduction in the percentage of myelinated axons. This postnatal hypomyelination was accompanied by a decline in oligodendrocyte iron uptake and with an increase in brain oxidative stress. We also tested the relevance of astrocytic Fth expression in the cuprizone model of myelin damage and repair. Fth deletion in Glast1-positive astrocytes significantly reduced myelin production and the density of mature myelinating oligodendrocytes throughout the complete remyelination process. These results indicate that Fth iron storage in astrocytes is vital for early oligodendrocyte development as well as for the remyelination of the CNS.

Ceruloplasmin deletion in myelinating glial cells induces myelin disruption and oxidative stress in the central and peripheral nervous systems

Ceruloplasmin (Cp) is a ferroxidase enzyme that is essential for cell iron efflux and has been postulated to have a neuroprotective role. During the myelination process, oligodendrocytes (OLs) and Schwann cells (SCs) express high levels of Cp, but the role of this enzyme in glial cell development and function is completely unknown. To define the function of Cp in the myelination of the central and peripheral nervous systems, we have conditionally knocked-out Cp specifically in OLs and SCs during early postnatal development as well as in aged mice. Cp ablation in early OLs (postnatal day 2, P2) significantly affects the differentiation of these cells and the synthesis of myelin through the first four postnatal weeks. The total number of mature myelinating OLs was reduced, and the density of apoptotic OLs was increased. These changes were accompanied with reductions in the percentage of myelinated axons and increases in the g-ratio of myelinated fibers. Cp ablation in young myelinating OLs (P30 or P60) did not affect myelin synthesis and/or OL numbers, however, Cp loss in aged OLs (8 months) induced cell iron overload, apoptotic cell death, brain oxidative stress, neurodegeneration and myelin disruption. Furthermore, Cp deletion in SCs affected postnatal SC development and myelination and produced motor coordination deficits as well as oxidative stress in young and aged peripheral nerves. Together, our data indicate that Cp ferroxidase activity is essential for OLs and SCs maturation during early postnatal development and iron homeostasis in matured myelinating cells. Additionally, our results suggest that Cp expression in myelinating glial cells is crucial to prevent oxidative stress and neurodegeneration in the central and peripheral nervous systems.

BALB/c mice infected with Angiostrongylus cantonensis: A new model for demyelination in the brain.

In this study, we present a new model for demyelination of the central nervous system (CNS). BALB/c mice were infected with Angiostrongylus cantonensis and analyzed 7, 14, and 21 days postinfection. Neurological scale evaluation, magnetic resonance imaging (MRI), histology, real-time quantitative polymerase chain reaction, and western blotting were all performed on days 7, 14, and 21. The results showed that the neurological functions and weight of A. cantonensis-infected mice decreased markedly after 21 days of infection. MRI showed subdural effusion and white high signals in the corpus callosum in both T1WI and T2WI, while hematoxylin and eosin and luxol fast blue staining showed hemorrhage and demyelination in the corpus callosum. Transmission electron microscopy revealed that the ultrastructure of the myelin sheath in the corpus callosum was dispersed or disintegrated. The percentage of myelinated axons was significantly decreased, and the g-ratio was lower than that in the normal group. Both protein and mRNA levels of myelin basic protein decreased markedly at 21 days postinfection. Immunofluorescence revealed that the number of CC1 positive cells in the corpus callosum also decreased, which confirmed the damage of A. cantonensis to oligodendrocytes. Our experiments confirmed that A. cantonensis infection caused demyelination in the CNS of BALB/c mice after 21 days, and its clinical manifestations and pathological changes were similar to those of multiple sclerosis and other CNS demyelination models. Thus, mice infected with A. cantonensis could be used as a new model to study acute demyelination of the CNS.

Impaired Postnatal Myelination in a Conditional Knockout Mouse for the Ferritin Heavy Chain in Oligodendroglial Cells.

To define the importance of iron storage in oligodendrocyte development and function, the ferritin heavy subunit (Fth) was specifically deleted in oligodendroglial cells. Blocking Fth synthesis in Sox10 or NG2-positive oligodendrocytes during the first or the third postnatal week significantly reduces oligodendrocyte iron storage and maturation. The brain of Fth KO animals presented an important decrease in the expression of myelin proteins and a substantial reduction in the percentage of myelinated axons. This hypomyelination was accompanied by a decline in the number of myelinating oligodendrocytes and with a reduction in proliferating oligodendrocyte progenitor cells (OPCs). Importantly, deleting Fth in Sox10-positive oligodendroglial cells after postnatal day 60 has no effect on myelin production and/or oligodendrocyte quantities. We also tested the capacity of Fth-deficient OPCs to remyelinate the adult brain in the cuprizone model of myelin injury and repair. Fth deletion in NG2-positive OPCs significantly reduces the number of mature oligodendrocytes and myelin production throughout the remyelination process. Furthermore, the corpus callosum of Fth KO animals presented a significant decrease in the percentage of remyelinated axons and a substantial reduction in the average myelin thickness. These results indicate that Fth synthesis during the first three postnatal weeks is important for an appropriate oligodendrocyte development, and suggest that Fth iron storage in adult OPCs is also essential for an effective remyelination of the mouse brain.SIGNIFICANCE STATEMENT To define the importance of iron storage in oligodendrocyte function, we have deleted the ferritin heavy chain (Fth) specifically in the oligodendrocyte lineage. Fth ablation in oligodendroglial cells throughout early postnatal development significantly reduces oligodendrocyte maturation and myelination. In contrast, deletion of Fth in oligodendroglial cells after postnatal day 60 has no effect on myelin production and/or oligodendrocyte numbers. We have also tested the consequences of disrupting Fth iron storage in oligodendrocyte progenitor cells (OPCs) after demyelination. We have found that Fth deletion in NG2-positive OPCs significantly delays the remyelination process in the adult brain. Therefore, Fth iron storage is essential for early oligodendrocyte development as well as for OPC maturation in the demyelinated adult brain.

Transient Hypothyroidism During Lactation Alters the Development of the Corpus Callosum in Rats. An in vivo Magnetic Resonance Image and Electron Microscopy Study.

Magnetic resonance imaging (MRI) data of children with late diagnosed congenital hypothyroidism and cognitive alterations such as abnormal verbal memory processing suggest altered telencephalic commissural connections. The corpus callosum (CC) is the major inter-hemispheric commissure that contra-laterally connects neocortical areas. However, in late diagnosed neonates with congenital hypothyroidism, the possible effect of early transient and chronic postnatal hypothyroidism still remains unknown. We have studied the development of the anterior, middle and posterior CC, using in vivo MRI and electron microscopy in hypothyroid and control male rats. Four groups of methimazole (MMI) treated rats were studied. One group, as a model for early transient hypothyroidism, was MMI-treated from postnatal day (P) 0 to P21; some of these rats were also treated with L-thyroxine (T4) from P15 to 21. Another group modeling chronic hypothyroid, were treated with MMI from P0 to 150 and from embryonic day 10 to P170. The results obtained from these groups were compared with same age control rats. The normalized T2 signal obtained using MRI was higher in MMI-treated rats and correlated with a low number and percentage of myelinated axons. The number and density of myelinated axons decreased in transient and chronic hypothyroid rats at P150. The g-ratio (inner to outer diameter ratio) and the estimated conduction velocity of myelinated axons were similar between MMI-treated and controls, but the conduction delay decreased in the posterior CC of MMI-treated rats compared to controls. These data show that early postnatal transient and chronic hypothyroidism alters CC maturation in a way that may affect the callosal transfer of information. These alterations cannot be reversed after delayed T4-treatment. Our data support the findings of neurocognitive delay in late T4-treated children with congenital hypothyroidism.

Iron Metabolism in the Peripheral Nervous System: The Role of DMT1, Ferritin, and Transferrin Receptor in Schwann Cell Maturation and Myelination.

Iron is an essential cofactor for many cellular enzymes involved in myelin synthesis, and iron homeostasis unbalance is a central component of peripheral neuropathies. However, iron absorption and management in the PNS are poorly understood. To study iron metabolism in Schwann cells (SCs), we have created 3 inducible conditional KO mice in which three essential proteins implicated in iron uptake and storage, the divalent metal transporter 1 (DMT1), the ferritin heavy chain (Fth), and the transferrin receptor 1 (Tfr1), were postnatally ablated specifically in SCs. Deleting DMT1, Fth, or Tfr1 in vitro significantly reduce SC proliferation, maturation, and the myelination of DRG axons. This was accompanied by an important reduction in iron incorporation and storage. When these proteins were KO in vivo during the first postnatal week, the sciatic nerve of all 3 conditional KO animals displayed a significant reduction in the synthesis of myelin proteins and in the percentage of myelinated axons. Knocking out Fth produced the most severe phenotype, followed by DMT1 and, last, Tfr1. Importantly, DMT1 as well as Fth KO mice showed substantial motor coordination deficits. In contrast, deleting these proteins in mature myelinating SCs results in milder phenotypes characterized by small reductions in the percentage of myelinated axons and minor changes in the g-ratio of myelinated axons. These results indicate that DMT1, Fth, and Tfr1 are critical proteins for early postnatal iron uptake and storage in SCs and, as a consequence, for the normal myelination of the PNS.SIGNIFICANCE STATEMENT To determine the function of the divalent metal transporter 1, the transferrin receptor 1, and the ferritin heavy chain in Schwann cell (SC) maturation and myelination, we created 3 conditional KO mice in which these proteins were postnatally deleted in Sox10-positive SCs. We have established that these proteins are necessary for normal SC iron incorporation and storage, and, as a consequence, for an effective myelination of the PNS. Since iron is indispensable for SC maturation, understanding iron metabolism in SCs is an essential prerequisite for developing therapies for demyelinating diseases in the PNS.

Demyelination and shrinkage of axons in the retinal nerve fiber layer in chickens developing deprivation myopia

Placing diffusers in front of the eyes induces deprivation myopia in a variety of animal models. As a result of the low pass filtering of the retinal images, less spatial information is available to the retina which should reduce neural activity. Since it has been found that myelination of axons in the central nervous system is modulated by neuronal activity, we have studied whether ganglion cell axons may shrink in response to the restricted visual input. Young chickens were treated for 5 h or 7 days with frosted diffusers to induce deprivation myopia. Nerve fiber layer thickness was measured in vivo, using B-scan OCT. Refractive states were tracked by IR photoretinoscopy, and UV fundus reflectivity by a custom-built device which flashed an LED centered in the camera aperture and recorded pupil brightness after refractive errors were corrected by trial lenses. Moreover, structure and histology of the retinal nerve fibers layer (RNFL) were analyzed ex vivo using transmission electron microscopy and immunohistochemistry. Since chicks have both non-myelinated and myelinated fibers in their RNFL, the thickness of myelin sheaths (G ratio) was measured, as well as the percentage of myelinated axons and the diameters of unmyelinated axons. Short-term deprivation caused an increase in UV fundus reflectivity already after 5 h (measured as pixel grey levels in the pupil: 28 ± 5 vs. 36 ± 10, p < 0.05) and thinning of the myelin sheaths (higher G ratio), compared to untreated control eyes (0.74 ± 0.01 vs. 0.79 ± 0.03, p < 0.05). Neither axon diameters (0.81 ± 0.05 μm vs. 0.82 ± 0.15 μm) nor thickness of the RNFL had changed after only 5 h (42.9 ± 1.3 μm vs. 42.3 ± 2.5 μm). However, after 7 days of diffuser wear, axons had become thinner (0.56 ± 0.14 μm vs. 0.78 ± 0.09 μm vs, p < 0.05), which could explain the thinning of the RNFL (36.3 ± 2.7 μm vs. 42.1 ± 2.4 μm, p < 0.01). Furthermore, myopic eyes had 38% less myelinated axons than untreated eyes as determined by immunohistochemical labelling against myelin basic protein (immunopositive areas in the central retina 1406 ± 341 μm2 vs. 2185 ± 290 μm2 in controls, p < 0.001). Myelin sheaths in the remaining axons remained unchanged (G ratio 0.76 ± 0.02 vs. 0.76 ± 0.03). Our study shows that deprivation myopia is associated with a significant loss of myelinated axons and shrinkage of the axon diameters of certain fibers in the RNFL. Early changes were already detected after 5 h and were accompanied by an increased fundus reflectivity in UV light. These parameters could therefore serve as the biomarkers for myopia development, at least in the chicken.

Ontogenetic Shifts in the Number of Axons in the Olfactory Tract and Optic Nerve in Two Species of Deep-Sea Grenadier Fish (Gadiformes: Macrouridae: Coryphaenoides)

Neuroanatomical studies of the peripheral sense organs and brains of deep-sea fishes are particularly useful for predicting their sensory capabilities and ultimately their behaviour. Over the abyssal plane (between 2,000 and 6,000 m), communities of grenadiers (Gadiformes: Macrouridae) play an important ecological role as predator-scavengers. Previous studies suggest that these fishes rely heavily on chemosensation, especially olfaction. Furthermore, at least one species, Coryphaenoides armatus, undergoes an ontogenetic shift in the relative size of the optic tectum and the olfactory bulbs, suggesting. a shift from a reliance on vision to olfaction during ontogeny, apparently in association with a shift to a more scavenging lifestyle. Here, we compared the olfactory and visual sensory inputs to the brain in C. armatus, and in a second, closely-related species, Coryphaenoides profundicolus, by assessing the total number of axons (myelinated and unmyelinated) in the olfactory tract and optic nerve in a range of individuals from both species. In C. armatus, the numbers of axons in both tract and nerve increased with body size, with the total number of axons in the olfactory tract being far greater than the number of axons in the optic nerve. These differences became more pronounced in larger animals. In the two smaller C. profundicolus individuals (≤315 mm SL), there were more axons in the optic nerve than in the olfactory tract, but the opposite situation was found in larger individuals. As in C. armatus, the number of olfactory tract axons also increased with body size in C. profundicolus, but in contrast, the number of optic nerve axons decreased in this species. These results suggest that both C. armatus and C. profundicolus undergo an ontogenetic shift in sensory orientation, with olfaction becoming relatively more important than vision in larger animals. The differences in the ratio of olfactory tract to optic nerve axons in C. armatus indicate that olfaction is of particular importance to larger individuals of this species. In both species, the percentage of myelinated axons in the olfactory tract was relatively low, but we found evidence for interspecific and ontogenetic variation in the percentages of myelinated axons in the optic nerve.

Alterations in synaptic density and myelination in response to exposure to high-energy charged particles.

High-energy charged particles are considered particularly hazardous components of the space radiation environment. Such particles include fully ionized energetic nuclei of helium, silicon, and oxygen, among others. Exposure to charged particles causes reactive oxygen species production, which has been shown to result in neuronal dysfunction and myelin degeneration. Here we demonstrate that mice exposed to high-energy charged particles exhibited alterations in dendritic spine density in the hippocampus, with a significant decrease of thin spines in mice exposed to helium, oxygen, and silicon, compared to sham-irradiated controls. Electron microscopy confirmed these findings and revealed a significant decrease in overall synapse density and in nonperforated synapse density, with helium and silicon exhibiting more detrimental effects than oxygen. Degeneration of myelin was also evident in exposed mice with significant changes in the percentage of myelinated axons and g-ratios. Our data demonstrate that exposure to all types of high-energy charged particles have a detrimental effect, with helium and silicon having more synaptotoxic effects than oxygen. These results have important implications for the integrity of the central nervous system and the cognitive health of astronauts after prolonged periods of space exploration.

The Divalent Metal Transporter 1 (DMT1) Is Required for Iron Uptake and Normal Development of Oligodendrocyte Progenitor Cells.

The divalent metal transporter 1 (DMT1) is a multimetal transporter with a primary role in iron transport. Although DMT1 has been described previously in the CNS, nothing was known about the role of this metal transporter in oligodendrocyte maturation and myelination. To determine whether DMT1 is required for oligodendrocyte progenitor cell (OPC) maturation, we used siRNAs and the Cre-lox system to knock down/knock out DMT1 expression in vitro as well as in vivo Blocking DMT1 synthesis in primary cultures of OPCs reduced oligodendrocyte iron uptake and significantly delayed OPC development. In vivo, a significant hypomyelination was found in DMT1 conditional knock-out mice in which DMT1 was postnatally deleted in NG2- or Sox10-positive OPCs. The brain of DMT1 knock-out animals presented a decrease in the expression levels of myelin proteins and a substantial reduction in the percentage of myelinated axons. This reduced postnatal myelination was accompanied by a decrease in the number of myelinating oligodendrocytes and a rise in proliferating OPCs. Furthermore, using the cuprizone model of demyelination, we established that DMT1 deletion in NG2-positive OPCs lead to less efficient remyelination of the adult brain. These results indicate that DMT1 is vital for OPC maturation and for the normal myelination of the mouse brain.SIGNIFICANCE STATEMENT To determine whether divalent metal transporter 1 (DMT1), a multimetal transporter with a primary role in iron transport, is essential for oligodendrocyte development, we created two conditional knock-out mice in which DMT1 was postnatally deleted in NG2- or Sox10-positive oligodendrocyte progenitor cells (OPCs). We have established that DMT1 is necessary for normal OPC maturation and is required for an efficient remyelination of the adult brain. Since iron accumulation by OPCs is indispensable for myelination, understanding the iron incorporation mechanism as well as the molecules involved is critical to design new therapeutic approaches to intervene in diseases in which the myelin sheath is damaged or lost.

Enhanced oligodendrocyte maturation and myelination in a mouse model of Timothy syndrome.

To study the role of L-type voltage-gated Ca++ channels in oligodendrocyte development, we used a mouse model of Timothy syndrome (TS) in which a gain-of-function mutation in the α1 subunit of the L-type Ca++ channel Cav1.2 gives rise to an autism spectrum disorder (ASD). Oligodendrocyte progenitor cells (OPCs) isolated from the cortex of TS mice showed greater L-type Ca++ influx and displayed characteristics suggestive of advanced maturation compared to control OPCs, including a more complex morphology and higher levels of myelin protein expression. Consistent with this, expression of Cav1.2 channels bearing the TS mutation in wild-type OPCs triggered process formation and promoted oligodendrocyte-neuron interaction via the activation of Ca++ /calmodulin-dependent protein kinase II. To ascertain whether accelerated OPC maturation correlated with functional enhancements, we examined myelination in the TS brain at different postnatal time points. The expression of myelin proteins was significantly higher in the corpus callosum, cortex and striatum of TS animals, and immunohistochemical analysis for oligodendrocyte stage-specific markers revealed an increase in the density of myelinating oligodendrocytes in several areas of the TS brain. Along the same line, electron microscopy studies in the corpus callosum of TS animals showed significant increases both in the percentage of myelinated axons and in the thickness of myelin sheaths. In summary, these data indicate that OPC development and oligodendrocyte myelination is enhanced in the brain of TS mice, and suggest that this mouse model of a syndromic ASD is a useful tool to explore the role of L-type Ca++ channels in myelination.

Transient Hypothyroidism During Lactation Arrests Myelination in the Anterior Commissure of Rats. A Magnetic Resonance Image and Electron Microscope Study.

Thyroid hormone deficiency at early postnatal ages affects the cytoarchitecture and function of neocortical and telencephalic limbic areas, leading to impaired associative memory and in a wide spectrum of neurological and mental diseases. Neocortical areas project interhemispheric axons mostly through the corpus callosum and to a lesser extent through the anterior commissure (AC), while limbic areas mostly project through the AC and hippocampal commissures. Functional magnetic resonance data from children with late diagnosed congenital hypothyroidism and abnormal verbal memory processing, suggest altered ipsilateral and contralateral telencephalic connections. Gestational hypothyroidism affects AC development but the possible effect of transient and chronic postnatal hypothyroidism, as occurs in late diagnosed neonates with congenital hypothyroidism and in children growing up in iodine deficient areas, still remains unknown. We studied AC development using in vivo magnetic resonance imaging and electron microscopy in hypothyroid and control male rats. Four groups of methimazole (MMI) treated rats were studied. One group was MMI-treated from postnatal day (P) 0 to P21; some of these rats were also treated with L-thyroxine (T4) from P15 to P21, as a model for early transient hypothyroidism. Other rats were MMI-treated from P0 to P150 and from embryonic day (E) 10 to P170, as a chronic hypothyroidism group. The results were compared with age paired control rats. The normalized T2 signal using magnetic resonance image was higher in MMI-treated rats and correlated with the number and percentage of myelinated axons. Using electron microscopy, we observed decreased myelinated axon number and density in transient and chronic hypothyroid rats at P150, unmyelinated axon number increased slightly in chronic hypothyroid rats. In MMI-treated rats, the myelinated axon g-ratio and conduction velocity was similar to control rats, but with a decrease in conduction delays. These data show that early postnatal transient and chronic hypothyroidism alters AC maturation that may affect the transfer of information through the AC. The alterations cannot be recovered after delayed T4-treatment. Our data support the neurocognitive delay found in late T4-treated children with congenital hypothyroidism.

Conditional Deletion of the L-Type Calcium Channel Cav1.2 in NG2-Positive Cells Impairs Remyelination in Mice.

Exploring the molecular mechanisms that drive the maturation of oligodendrocyte progenitor cells (OPCs) during the remyelination process is essential to developing new therapeutic tools to intervene in demyelinating diseases such as multiple sclerosis. To determine whether L-type voltage-gated calcium channels (L-VGCCs) are required for OPC development during remyelination, we generated an inducible conditional knock-out mouse in which the L-VGCC isoform Cav1.2 was deleted in NG2-positive OPCs (Cav1.2KO). Using the cuprizone (CPZ) model of demyelination and mice of either sex, we establish that Cav1.2 deletion in OPCs leads to less efficient remyelination of the adult brain. Specifically, Cav1.2KO OPCs mature slower and produce less myelin than control oligodendrocytes during the recovery period after CPZ intoxication. This reduced remyelination was accompanied by an important decline in the number of myelinating oligodendrocytes and in the rate of OPC proliferation. Furthermore, during the remyelination phase of the CPZ model, the corpus callosum of Cav1.2KO animals presented a significant decrease in the percentage of myelinated axons and a substantial increase in the mean g-ratio of myelinated axons compared with controls. In addition, in a mouse line in which the Cav1.2KO OPCs were identified by a Cre reporter, we establish that Cav1.2KO OPCs display a reduced maturational rate through the entire remyelination process. These results suggest that Ca2+ influx mediated by L-VGCCs in oligodendroglial cells is necessary for normal remyelination and is an essential Ca2+ channel for OPC maturation during the remyelination of the adult brain.SIGNIFICANCE STATEMENT Ion channels implicated in oligodendrocyte differentiation and maturation may induce positive signals for myelin recovery. Voltage-gated Ca2+ channels (VGCCs) are important for normal myelination by acting at several critical steps during oligodendrocyte progenitor cell (OPC) development. To determine whether voltage Ca2+ entry is involved in oligodendrocyte differentiation and remyelination, we used a conditional knockout mouse for VGCCs in OPCs. Our results indicate that VGCCs can modulate oligodendrocyte maturation in the demyelinated brain and suggest that voltage-gated Ca2+ influx in OPCs is critical for remyelination. These findings could lead to novel approaches for obtaining a better understanding of the factors that control OPC maturation in order to stimulate this pool of progenitors to replace myelin in demyelinating diseases.

Conditional Deletion of the L-Type Calcium Channel Cav1.2 in Oligodendrocyte Progenitor Cells Affects Postnatal Myelination in Mice.

Overall, it is clear that cells in the oligodendrocyte lineage exhibit remarkable plasticity with regard to the expression of Ca2+ channels and that perturbation of Ca2+ homeostasis likely plays an important role in the pathogenesis underlying demyelinating diseases. To determine whether voltage-gated Ca2+ entry is involved in oligodendrocyte maturation and myelination, we used a conditional knock-out mouse for voltage-operated Ca2+ channels in oligodendrocyte progenitor cells. Our results indicate that voltage-operated Ca2+ channels can modulate oligodendrocyte development in the postnatal brain and suggest that voltage-gated Ca2+ influx in oligodendroglial cells is critical for normal myelination. These findings could lead to novel approaches to intervene in neurodegenerative diseases in which myelin is lost or damaged.

Enhanced Femoral Nerve Regeneration After Tubulization with a Tyrosine-Derived Polycarbonate Terpolymer: Effects of Protein Adsorption and Independence of Conduit Porosity

Following complete nerve transection, entubulation of the nerve stumps helps guide axons to reconnect distally. In this study, a biodegradable and noncytotoxic tyrosine-derived polycarbonate terpolymer composed of 89.5 mol% desaminotyrosyl tyrosine ethyl ester (DTE), 10 mol% desaminotyrosyl tyrosine (DT), and 0.5 mol% poly(ethylene glycol) (PEG, molecular weight [Mw]=1 kDa) [designated as E10-0.5(1K)] was used to fabricate conduits for peripheral nerve regeneration. These conduits were evaluated against commercially available nonporous polyethylene (PE) tubes. The two materials are characterized in vitro for differences in surface properties, and the conduits are then evaluated in vivo in a critical-sized nerve defect in the mouse femoral nerve model. Conduits were fabricated from E10-0.5(1K) in both porous [P-E10-0.5(1K)] and nonporous [NP-E10-0.5(1K)] configurations. The results illustrate that adsorption of laminin, fibronectin, and collagen type I was enhanced on E10-0.5(1K) compared to PE. In addition, in vivo the E10-0.5(1K) conduits improved functional recovery over PE conduits, producing regenerated nerves with a fivefold increase in the number of axons, and an eightfold increase in the percentage of myelinated axons. These increases were observed for both P-E10-0.5(1K) and NP-E10-0.5(1K) after 15 weeks. When conduits were removed at 7 or 14 days following implantation, an increase in Schwann cell proteins and fibrin matrix formation was observed in E10-0.5(1K) conduits over PE conduits. These results indicate that E10-0.5(1K) is a pro-regenerative material for peripheral nerves and that the porosity of P-E10-0.5(1K) conduits was inconsequential in this model of nerve injury.

Quantitative ultrastructural evidence of myelin malformation in optic nerves of rats submitted to a multideficient diet

Pups were subjected to malnutrition by feeding the lactating mothers a multi-deficient (8% protein content) diet, known as regional basic diet (RBD), from birth up to weaning. The weanings were fed the same diet until 60 days of age. Ultrastructure of the optic nerve was analyzed at postnatal (P) day P8, P13, P21, P30 and P60. Electron microscopy revealed that at P8 the process of myelination has not started yet in neither groups. At P13 different stages of myelination were observed and, in the experimental group, the optic nerve showed non-organized axon bundles and empty spaces. Control optic nerve at P21 exhibited axons with fully developed myelin sheath; whereas malnourished group had many axons being enveloped by myelin with anomalous alteration. These alterations were present in malnourished group at P30 and P60. Quantitative analysis showed statistically significant difference between control and malnourished groups when compared to the percentage of myelinated axons, axons with myelin anomalous alterations (MAA) and non-myelinated axons. Also, the myelin area was significantly smaller in malnourished groups when compared to control group. Finally, a high percentage of large non-myelinated fibers were found in the malnourished group. In conclusion, our results show that early malnutrition by a multideficient diet (RBD) affects permanently the optic nerve organization and myelination, indicating an impairment of nerve transmission and a probable dysfunction in the visual ability.

In vivo actions of insulin-like growth factor-I (IGF-I) on brain myelination: studies of IGF-I and IGF binding protein-1 (IGFBP-1) transgenic mice

To study the effects and mechanisms of insulin-like growth factor I (IGF-I) on brain myelination in vivo, the morphology of myelinated axons and the expression of myelin specific protein genes have been examined in transgenic (Tg) mice that overexpress IGF-I and that those ectopically express IGF binding protein-1 (IGFBP-1), a protein that inhibits IGF-I actions when present in molar excess. Our data show that the percentage of myelinated axons and the thickness of myelin sheaths are significantly increased in IGF-I Tg and decreased in the IGFBP-1 mice. Cerebral cortical proteolipid protein (PLP) and myelin basic protein (MBP) mRNAs consistently exhibit approximately 200% increases in IGF-I Tg mice and approximately 50% decreases in IGFBP-1 Tg mice. The percentage of oligodendrocytes labeled with a PLP cRNA probe in the corpus callosum and cerebral cortex also is increased in IGF-I Tg mice and reduced in IGFBP-1 Tg mice, suggesting that IGF-I promotes oligodendrocyte survival and/or proliferation. The alterations in the number of oligodendrocytes, however, can not completely account for the changes in myelin gene expression. These results strongly indicate that IGF-I increases myelination by increasing the number of myelinated axons and the thickness of myelin sheaths, the latter by mechanisms that involve stimulation of the expression of myelin protein genes and increase of oligodendrocyte number.

Facial nerve regeneration through semipermeable chambers in the rabbit.

Peripheral neural regeneration, over a 10-mm transectional gap, was determined in 70 rabbit buccal divisions of the facial nerve using two entubational systems (semipermeable and impermeable silicone chambers) prefilled with three natural occurring media (serum, blood, and saline) during a 5-week period. The number of myelinated axonal regenerates at the midchamber and at 2 mm in the distal transected neural stump were counted in each group and compared to pooled myelinated axonal counts in 9 normal rabbit buccal divisions of the facial nerve. Semipermeable porous chambers had an overall greater regeneration success rate (75% vs. 42.8%) and regained, on the average, a higher number of myelinated axons (51.4% vs. 26.1%) than silicone chamber regenerates. Semipermeable chambers prefilled with serum or blood had significantly higher regeneration success rates, myelinated axonal counts, and percentages of neural innervation of the distal transected neural stump. Both entubational systems produced similar axonal counts with intraluminal saline. The highest overall success rate (93.7%) and average number of myelinated axons per chamber (3072) were achieved in semipermeable chambers prefilled with serum. The greatest variability in myelinated axonal counts (0 to 3266 axons) and percentage of distal stump innervation (5.5% to 98.1%) was seen in silicone chambers filled with saline. The percentage of myelinated axons from the midchamber that innervated the distal stump was greater in semipermeable chambers with blood (73%) and serum (54%) than in silicone saline chambers (43%). On the average, the distal stumps from semipermeable chambers filled with serum (47%) and blood (33.5%) regained a higher percentage of normal myelinated axonal counts than silicone-saline chambers (12.5%). These results suggest that both the construction of entubational chamber and the intraluminal medium can have significant influence on neurite regeneration. Semipermeable chambers prefilled with serum have a strong neurite-promoting potential in peripheral neural regeneration of rabbit facial nerves.