Circadian Modulation of Gene Expression, but not Glutamate Uptake, in Mouse and Rat Cortical Astrocytes

Circadian clocks control daily rhythms including sleep-wake, hormone secretion, and metabolism. These clocks are based on intracellular transcription-translation feedback loops that sustain daily oscillations of gene expression in many cell types. Mammalian astrocytes display circadian rhythms in the expression of the clock genes Period1 (Per1) and Period2 (Per2). However, a functional role for circadian oscillations in astrocytes is unknown. Because uptake of extrasynaptic glutamate depends on the presence of Per2 in astrocytes, we asked whether glutamate uptake by glia is circadian.We measured glutamate uptake, transcript and protein levels of the astrocyte-specific glutamate transporter, Glast, and the expression of Per1 and Per2 from cultured cortical astrocytes and from explants of somatosensory cortex. We found that glutamate uptake and Glast mRNA and protein expression were significantly reduced in Clock/Clock, Per2- or NPAS2-deficient glia. Uptake was augmented when the medium was supplemented with dibutyryl-cAMP or B27. Critically, glutamate uptake was not circadian in cortical astrocytes cultured from rats or mice or in cortical slices from mice.We conclude that glutamate uptake levels are modulated by CLOCK, PER2, NPAS2, and the composition of the culture medium, and that uptake does not show circadian variations

Persistent neuronal Ube3a expression in the suprachiasmatic nucleus of Angelman syndrome model mice

Mutations or deletions of the maternal allele of the UBE3A gene cause Angelman syndrome (AS), a severe neurodevelopmental disorder. The paternal UBE3A/Ube3a allele becomes epigenetically silenced in most neurons during postnatal development in humans and mice; hence, loss of the maternal allele largely eliminates neuronal expression of UBE3A protein. However, recent studies suggest that paternal Ube3a may escape silencing in certain neuron populations, allowing for persistent expression of paternal UBE3A protein. Here we extend evidence in AS model mice (Ube3a(m–/p+)) of paternal UBE3A expression within the suprachiasmatic nucleus (SCN), the master circadian pacemaker. Paternal UBE3A-positive cells in the SCN show partial colocalization with the neuropeptide arginine vasopressin (AVP) and clock proteins (PER2 and BMAL1), supporting that paternal UBE3A expression in the SCN is often of neuronal origin. Paternal UBE3A also partially colocalizes with a marker of neural progenitors, SOX2, implying that relaxed or incomplete imprinting of paternal Ube3a reflects an overall immature molecular phenotype. Our findings highlight the complexity of Ube3a imprinting in the brain and illuminate a subpopulation of SCN neurons as a focal point for future studies aimed at understanding the mechanisms of Ube3a imprinting

Interobservational variation in determining fusion rates in anterior cervical discectomy and fusion procedures

The fusion rate represents one of the most commonly used criteria for evaluating the efficacy of spinal surgical techniques and the effectiveness of newly developed instrumentation and spinal implants. Reported fusion rates are not frequently supported by adequate information regarding by whom and how fusion was defined. In our prospective study we examined the fusion rate in patients undergoing first time anterior cervical discectomy and fusion for degenerative disease. Separate, well-defined radiographic fusion criteria were used and the 12-month post-operative X-rays were reviewed independently by a neurosurgeon, a neuroradiologist and an orthopedic surgeon, who were not involved in the patients’ management. The observed fusion rates were 77.3, 87.8 and 84.7% respectively. Statistical analysis demonstrated concordance rates of 87.8, 91 and 91.4% and Kappa coefficients of 0.585, 0.620 and 0.723 for each pair of evaluators. Another set of ratings of the same radiographs, by the same interviewers, was obtained 6 weeks after the initial one. The reported fusion rates were 78.2% for the neurosurgeon, 87.4% for the orthopedic surgeon, and 86.1% for the neuroradiologist. Statistical analysis demonstrated intra-observer concordance rates of 98.7, 92.2 and 97.9% respectively, while the Kappa coefficients were 0.963, 0.677 and 0.907 for each reviewer. Our findings confirm the necessity of defining and describing criteria for fusion whenever this rate is reported in clinical series. The lack of widely accepted, well-defined criteria makes comparison of these results difficult. The development of a well organized, prospective clinical study in which fusion and outcome will be assessed by both clinical and radiographic parameters could significantly contribute to a more accurate evaluation of overall outcome of cervical spinal procedures