SUMMARY While diet-induced obesity has been exclusively attributed to increased caloric intake from fat, animals fed high fat diet (HFD) ad libitum (ad lib) eat frequently throughout day and night disrupting the normal feeding cycle. To test whether obesity and metabolic diseases result from HFD or disruption of metabolic cycles, we subjected mice to either ad lib or time restricted feeding (tRF) of a HFD for 8 h/day. Mice under tRF consume equivalent calories from HFD as those with ad lib access, yet are protected against obesity, hyperinsulinemia, hepatic steatosis, inflammation, and have improved motor coordination. The tRF regimen improved CREB, mTOR and AMPK pathway function and oscillations of the circadian clock and their target genes' expression. These changes in catabolic and anabolic pathways altered liver metabolome, improved nutrient utilization and energy expenditure. We demonstrate in mice that tRF regimen is a non-pharmacological strategy against obesity and associated diseases.

Protoplasmic astrocytes are increasingly thought to interact extensively with neuronal elements in the brain and to influence their activity. Recent reports have also begun to suggest that physiologically, and perhaps functionally, diverse forms of these cells may be present in the CNS. Our current understanding of astrocyte form and distribution is based predominantly on studies that used the astrocytic marker glial fibrillary acidic protein (GFAP) and on studies using metal-impregnation techniques. The prevalent opinion, based on studies using these methods, is that astrocytic processes overlap extensively and primarily share the underlying neuropil. However, both of these techniques have serious shortcomings for visualizing the interactions among these structurally complex cells. In the present study, intracellular injection combined with immunohistochemistry for GFAP show that GFAP delineates only approximately 15% of the total volume of the astrocyte. As a result, GFAP-based images have led to incorrect conclusions regarding the interaction of processes of neighboring astrocytes. To investigate these interactions in detail, groups of adjacent protoplasmic astrocytes in the CA1 stratum radiatum were injected with fluorescent intracellular tracers of distinctive emissive wavelengths and analyzed using three-dimensional (3D) confocal analysis and electron microscopy. Our findings show that protoplasmic astrocytes establish primarily exclusive territories. The knowledge of how the complex morphology of protoplasmic astrocytes affects their 3D relationships with other astrocytes, oligodendroglia, neurons, and vasculature of the brain should have important implications for our understanding of nervous system function.

Although new and functional neurons are produced in the adult brain, little is known about how they integrate into mature networks. Here we explored the mechanisms of synaptogenesis on neurons born in the adult mouse hippocampus using confocal microscopy, electron microscopy and live imaging. We report that new neurons, similar to mature granule neurons, were contacted by axosomatic, axodendritic and axospinous synapses. Consistent with their putative role in synaptogenesis, dendritic filopodia were more abundant during the early stages of maturation and, when analyzed in three dimensions, the tips of all filopodia were found within 200 nm of preexisting boutons that already synapsed on other neurons. Furthermore, dendritic spines primarily synapsed on multiple-synapse boutons, suggesting that initial contacts were preferentially made with preexisting boutons already involved in a synapse. The connectivity of new neurons continued to change until at least 2 months, long after the formation of the first dendritic protrusions.

It is assumed that synaptic strengthening and weakening balance throughout learning to avoid runaway potentiation and memory interference. However, energetic and informational considerations suggest that potentiation should occur primarily during wake, when animals learn, and depression should occur during sleep. We measured 6,920 synapses in mouse motor and sensory cortices using 3D-electron microscopy. The axon-spine interface (ASI) decreased ~18% after sleep compared with wake. This decrease was proportional to ASI size, which is indicative of scaling. Scaling was selective, sparing synapses that were large and lacked recycling endosomes. Similar scaling occurred for spine head volume, suggesting a distinction between weaker, more plastic synapses (~80%) and stronger, more stable synapses. These results support the hypothesis that a core function of sleep is to renormalize overall synaptic strength increased by wake.

Summary The inability to purify and culture astrocytes has long hindered studies of their function. Whereas astrocyte progenitor cells can be cultured from neonatal brain, culture of mature astrocytes from postnatal brain has not been possible. Here we report a new method to prospectively purify astrocytes by immunopanning. These astrocytes undergo apoptosis in culture, but vascular cells and HBEGF promote their survival in serum-free culture. We found that some developing astrocytes normally undergo apoptosis in vivo and that the vast majority of astrocytes contact blood vessels, suggesting that astrocytes are matched to blood vessels by competing for vascular-derived trophic factors such as HBEGF. Compared to traditional astrocyte cultures, the gene profiles of the cultured purified postnatal astrocytes much more closely resemble those of in vivo astrocytes. Although these astrocytes strongly promote synapse formation and function, they do not secrete glutamate in response to stimulation.

Reactive astrocytes in neurotrauma, stroke, or neurodegeneration are thought to undergo cellular hypertrophy, based on their morphological appearance revealed by immunohistochemical detection of glial fibrillary acidic protein, vimentin, or nestin, all of them forming intermediate filaments, a part of the cytoskeleton. Here, we used a recently established dye-filling method to reveal the full three-dimensional shape of astrocytes assessing the morphology of reactive astrocytes in two neurotrauma models. Both in the denervated hippocampal region and the lesioned cerebral cortex, reactive astrocytes increased the thickness of their main cellular processes but did not extend to occupy a greater volume of tissue than nonreactive astrocytes. Despite this hypertrophy of glial fibrillary acidic protein-containing cellular processes, interdigitation between adjacent hippocampal astrocytes remained minimal. This work helps to redefine the century-old concept of hypertrophy of reactive astrocytes.astrocyte domains ͉ astrocyte hypertrophy O nce considered to be merely a cellular layer filling the interneuronal space and gluing neurons together (hence the term ''glia''), astrocytes are receiving ever-increasing attention. The rising recognition of their importance builds on knowledge of their role in maintaining CNS homeostasis, providing nutrition for neuronal cells, and neurotransmitter recycling. Recently, astrocytes were shown to control the number and function of neuronal synapses (1, 2) and blood flow in the brain (3, 4).Astrocytes exhibit an intricate bushy or spongiform morphology, and their very fine processes are in close contact with synapses and other components of brain parenchyma (5-8). These fine terminal processes appear postnatally in the final stage of astrocyte maturation. The subsequent elaboration of spongiform processes results in the development of boundaries between neighboring astrocyte domains, thus establishing exclusive territories for individual astrocytes, a phenomenon termed ''tiling'' (7, 9).With earlier methods, including immunohistochemical detection of astrocyte markers and impregnation techniques, the extent of overlap between astrocyte territories was not amenable to investigation. However, more recent staining methods, optical imaging techniques, and 3D reconstruction paradigms using dye-filled astrocytes in semifixed tissue allowed the assessment of the boundaries of protoplasmic astrocyte territories in the CA1 area of the uninjured rat hippocampus. Neighboring astrocytes invariably touched each other but showed little interdigitation, basically tiling to form unique domains (8).Astrocytes react to many CNS challenges, and reactive astrocytes are a hallmark of many neuropathologies. Reactive astrocytes are characterized by high-level expression of glial fibrillary acidic protein (GFAP), an intermediate filament protein, and by upregulation of intermediate filaments in the cytoplasm. Antibodies against GFAP, the most frequently used astrocyte marker (10), reveal the cytoskeletal struct...

Glioblastoma multiforme (GBM) is the most common and aggressive malignant primary brain tumor in humans. Here, we show that gliomas can originate from differentiated cells in the central nervous system (CNS), including cortical neurons. Transduction by oncogenic lentiviral vectors of neural stem cells (NSCs), astrocytes, or even mature neurons in the brain of mice can give rise to malignant gliomas. All the tumors, irrespective of the site of injection (initiating population), share common features of high expression of stem or progenitor markers and low expression of differentiation markers. Microarray analysis revealed that tumors of astrocytic and neuronal origin match the mesenchymal GBM subtype. We propose that most differentiated cells in the CNS upon defined genetic alterations undergo dedifferentiation to generate a NSC or progenitor state to initiate and maintain the tumor progression, as well as to give rise to the heterogeneous populations observed in malignant gliomas.

The formation and recall of sensory, motor, and cognitive representations require coordinated fast communication among multiple cortical areas. Interareal projections are mainly mediated by glutamatergic pyramidal cell projections; only few long-range GABAergic connections have been reported. Using in vivo recording and labeling of single cells and retrograde axonal tracing, we demonstrate novel long-range GABAergic projection neurons in the rat hippocampus: (1) somatostatin-and predominantly mGluR1␣-positive neurons in stratum oriens project to the subiculum, other cortical areas, and the medial septum; (2) neurons in stratum oriens, including somatostatin-negative ones; and (3) trilaminar cells project to the subiculum and/or other cortical areas but not the septum. These three populations strongly increase their firing during sharp wave-associated ripple oscillations, communicating this network state to the septotemporal system. Finally, a large population of somatostatin-negative GABAergic cells in stratum radiatum project to the molecular layers of the subiculum, presubiculum, retrosplenial cortex, and indusium griseum and fire rhythmically at high rates during theta oscillations but do not increase their firing during ripples. The GABAergic projection axons have a larger diameter and thicker myelin sheet than those of CA1 pyramidal cells. Therefore, rhythmic IPSCs are likely to precede the arrival of excitation in cortical areas (e.g., subiculum) that receive both glutamatergic and GABAergic projections from the CA1 area. Other areas, including the retrosplenial cortex, receive only rhythmic GABAergic CA1 input. We conclude that direct GABAergic projections from the hippocampus to other cortical areas and the septum contribute to coordinating oscillatory timing across structures.