Abstract:
The rhythm of sleep and wakefulness is a dynamic behavioral phenomenon present in virtually every animal species investigated to date. It is regulated by a fine-tuned network of sleep- and wakepromoting neurons in the brain. Broad evidence suggests that a consequence of staying awake is a progressive increase in synaptic strength – synaptic upscaling –, as the waking brain adapts to the changing environment primarily through synaptic changes. Enhanced synaptic strength, however, might soon become unsustainable, as stronger synapses consume more energy, take up more space, and require more supplies. It is known that sleep induces synaptic downscaling to re-establish synaptic homeostasis in the cortex and hippocampus. It is not clear, however, whether sleep-dependent changes in synaptic plasticity happen in other brain regions and, moreover, how changes in energy supply interact with the impact of sleep and wakefulness on synaptic homeostasis. This dissertation aimed to investigate net synaptic changes in the hypothalamus, the main center integrating metabolic and sleep-wake regulatory signals, resulting from disruptions of the sleep-wake rhythm and high-calorie food intake. It was hypothesized that sleep induces synaptic downscaling in the hypothalamus, whereas wakefulness and sleep deprivation are associated with upscaling. It was furthermore assumed that high-fat food intake leads to hypothalamus-specific synaptic upscaling. In order to explore net changes in synaptic strength in the hypothalamus across the sleep-wake cycle and during sleep deprivation, α-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid receptors (AMPARs) that contain the subunits GluA1 and GluA2 were determined in hypothalamic and cortical synaptoneurosomes of rats (study I). Since wakefulness- and sleep-dependent up- and, respectively, downscaling has been previously described in the cortex, this region was chosen to verify our approach. It was found that GluA1-containing AMPARs both in hypothalamus and cortex are high during wakefulness and low during sleep, indicating net synaptic upscaling during wakefulness and downscaling during sleep. Moreover, sleep deprivation (extension of the wake period by six hours) prevented synaptic downscaling in both structures. Furthermore, the impact of high-fat feeding on hypothalamic and cortical GluA1- and GluA2-containing AMPARs (study II) was investigated. Three days, but not one day of high-fat diet (HFD) decreased the levels of AMPAR GluA1 and GluA2 subunits, as well as GluA1 phosphorylation at Ser845, in the hypothalamus but not cortex. In a control experiment, the reversibility of HFD-induced changes was investigated by comparing the 3-day HFD with a 3-day HFD followed by four recovery days of normal chow. This experiment corroborated the suppressive effect of high-fat feeding on hypothalamic but not cortical AMPAR GluA1, GluA2 and GluA1 phosphorylation at Ser845, and indicated that the effects can be reversed by normal chow feeding. In both experiments, high-fat feeding stimulated energy intake and raised body weight as well as serum concentrations of insulin, leptin, free fatty acids and corticosterone. Only the 3-day HFD increased wakefulness assessed via video analysis in the six hours before tissue collection. These results demonstrate (i) that in accordance with our hypothesis, wakefulness is associated with net synaptic upscaling in the hypothalamus and sleep induces respective synaptic downscaling; sleep deprivation interferes with the latter process and induces upscaling. (ii) Contrary to our expectations, short-term high-fat feeding downregulates rather than upregulates hypothalamic synaptic strength; this effect is rapidly reversible and does not extend to the cortex. These findings suggest that global downscaling processes accompany the initial phase of high-calorie intake to shift the overall hypothalamic activity balance and, possibly, to counteract anabolic drive and weight gain. Sleep in comparison to wakefulness or sleep deprivation likewise attenuates synaptic strength in the hypothalamus, corroborating the assumption that sleep exerts a normalizing effect on neuronal metabolic homeostasis that is thwarted by sleep deprivation. It will be important to investigate how long-term exposure to high-calorie food and the eventual development of obesity tip these synaptic scales, and how manipulations of sleep and wakefulness might be employed to recalibrate the hypothalamic regulation of energy homeostasis.
Sleep
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