SUMMARY mTor kinase is involved in cell growth, proliferation, and differentiation. The roles of mTor activators, Rheb1 and Rheb2, have not been established in vivo. Here, we report that Rheb1, but not Rheb2, is critical for embryonic survival and mTORC1 signaling. Embryonic deletion of Rheb1 in neural progenitor cells abolishes mTORC1 signaling in developing brain and increases mTORC2 signaling. Remarkably, embryonic and early postnatal brain development appears grossly normal in these Rheb1f/f, Nes-cre mice with the notable exception of deficits of myelination. Conditional expression of Rheb1 transgene in neural progenitors increases mTORC1 activity and promotes myelination in the brain. In addition, the Rheb1 transgene rescues mTORC1 signaling and hypomyelination in the Rheb1f/f, Nes-cre mice. Our study demonstrates that Rheb1 is essential for mTORC1 signaling and myelination in the brain, and suggests that mTORC1 signaling plays a role in selective cellular adaptations, rather than general cellular viability.

Rheb1 is an immediate early gene that functions to activate mammalian target of rapamycin (mTor) selectively in complex 1 (mTORC1).We have demonstrated previously that Rheb1 is essential for myelination in the CNS using a Nestin-Cre driver line that deletes Rheb1 in all neural cell lineages, and recent studies using oligodendrocyte-specific CNP-Cre have suggested a preferential role for mTORC1 is myelination in the spinal cord. Here, we examine the role of Rheb1/mTORC1 in mouse oligodendrocyte lineage using separate Cre drivers for oligodendrocyte progenitor cells (OPCs) including Olig1-Cre and Olig2-Cre as well as differentiated and mature oligodendrocytes including CNP-Cre and Tmem10-Cre. Deletion of Rheb1 in OPCs impairs their differentiation to mature oligodendrocytes. This is accompanied by reduced OPC cell-cycle exit suggesting a requirement for Rheb1 in OPC differentiation. The effect of Rheb1 on OPC differentiation is mediated by mTor since Olig1-Cre deletion of mTor phenocopies Olig1-Cre Rheb1 deletion. Deletion of Rheb1 in mature oligodendrocytes, in contrast, does not disrupt developmental myelination or myelin maintenance. Loss of Rheb1 in OPCs or neural progenitors does not affect astrocyte formation in gray and white matter, as indicated by the pan-astrocyte marker Aldh1L1. We conclude that OPC-intrinsic mTORC1 activity mediated by Rheb1 is critical for differentiation of OPCs to mature oligodendrocytes, but that mature oligodendrocytes do not require Rheb1 to make myelin or maintain it in the adult brain. These studies reveal mechanisms that may be relevant for both developmental myelination and impaired remyelination in myelin disease.

Epilepsy and other neurological deficits are common, disabling manifestations of the genetic disorder, Tuberous Sclerosis Complex (TSC). Brain inflammation has been implicated in contributing to epileptogenesis in acquired epilepsy due to brain injury, but the potential role of inflammatory mechanisms in genetic epilepsies is relatively unexplored. In this study, we investigated activation of inflammatory mediators and tested the effects of anti-inflammatory treatment on epilepsy in the Tsc1-GFAP conditional knock-out mouse model of TSC (Tsc1GFAPCKO mice). Real-time quantitative RT-PCR, immunohistochemistry, and western blotting demonstrated increased expression of specific cytokines and chemokines, particularly IL-1β and CXCL10, in the neocortex and hippocampus of Tsc1GFAPCKO mice, which was reversed by treatment with a mammalian target of rapamycin complex 1 (mTORC1) inhibitor. Double-labeling immunohistochemical studies indicated that the increased IL-1β was localized primarily to astrocytes. Importantly, the increase in inflammatory markers was also observed in astrocyte culture in vitro and at 2 weeks of age in Tsc1GFAPCKO mice before the onset of epilepsy in vivo, indicating that the inflammatory changes were not secondary to seizures. Epicathechin-3-gallate, an inhibitor of IL-1β and CXCL10, at least partially reversed the elevated cytokine and chemokine levels, reduced seizure frequency, and prolonged survival of Tsc1GFAPCKO mice. These findings suggest that mTOR-mediated inflammatory mechanisms may be involved in epileptogenesis in the genetic epilepsy, TSC.

We propose an efficient optical electromagnetically induced transparency (EIT) cooling scheme for a cantilever with a nitrogen-vacancy center attached in a non-uniform magnetic field using dynamical Zeeman effect. In our scheme, the Zeeman effect combined with the quantum interference effect enhances the desired cooling transition and suppresses the undesired heating transitions. As a result, the cantilever can be cooled down to nearly the vibrational ground state under realistic experimental conditions within a short time. This efficient optical EIT cooling scheme can be reduced to the typical EIT cooling scheme under special conditions.

Summary Objective Tuberous sclerosis complex (TSC) is a genetic disorder, characterized by tumor formation in multiple organs and severe neurological manifestations, including epilepsy, intellectual disability, and autism. Abnormalities of both neurons and astrocytes have been implicated in contributing to the neurological phenotype of TSC, but the role of microglia in TSC has not been investigated. The objectives of this study were to characterize microglial activation in a mouse model of TSC, involving conditional inactivation of the Tsc1 gene predominantly in glial cells (Tsc1GFAPCKO mice), and to test the hypothesis that microglial activation contributes to epileptogenesis in this mouse model. Methods Microglial and astrocyte activation was examined in Tsc1GFAPCKO mice by Iba1 and GFAP immunohistochemistry. Cytokine and chemokine expression was evaluated with QPCR. Seizures were monitored by video-EEG. The effect of minocycline in inhibiting microglial and astrocyte activation, cytokine expression, and seizures was tested. Results Microglial cell number and size were increased in cortex and hippocampus of 3–4 wk old Tsc1GFAPCKO mice, which correlated with the onset of seizures. Minocycline treatment prevented the increase in number and cell size of microglia in 4 week old Tsc1GFAPCKO mice. However, minocycline treatment had no effect on astrocyte proliferation and cytokine/chemokine expression and the progression of seizures in Tsc1GFAPCKO mice. Significance Microglia cell number and size are abnormal in Tsc1GFAPCKO mice, and minocycline treatment inhibits this microglia activation, but does not suppress seizures. Microglia may play a role in the neurological manifestations of TSC, but additional studies are needed in other models and human studies to determine whether microglia are critical for epileptogenesis in TSC.