Cellular Mechanisms of Sleep Homeostasis : Studies on Brain Energy Metabolism and Aging

Sleep is governed by a homeostatic process in which the duration and quality of previous wake regulate the subsequent sleep. Active wakefulness is characterized with high frequency cortical oscillations and depends on stimulating influence of the arousal systems, such as the cholinergic basal forebrain (BF), while cessation of the activity in the arousal systems is required for slow wave sleep (SWS) to occur. The site-specific accumulation of adenosine (a by-product of ATP breakdown) in the BF during prolonged waking /sleep deprivation (SD) is known to induce sleep, thus coupling energy demand to sleep promotion. The adenosine release in the BF is accompanied with increases in extracellular lactate and nitric oxide (NO) levels. This thesis was aimed at further understanding the cellular processes by which the BF is involved in sleep-wake regulation and how these processes are affected by aging. The BF function was studied simultaneously at three levels of organization: 1) locally at a cellular level by measuring energy metabolites 2) globally at a cortical level (the out-put area of the BF) by measuring EEG oscillations and 3) at a behavioral level by studying changes in vigilance states. Study I showed that wake-promoting BF activation, particularly with glutamate receptor agonist N-methyl-D-aspatate (NMDA), increased extracellular adenosine and lactate levels and led to a homeostatic increase in the subsequent sleep. Blocking NMDA activation during SD reduced the high frequency (HF) EEG theta (7-9 Hz) power and attenuated the subsequent sleep. In aging, activation of the BF during SD or experimentally with NMDA (studies III, IV), did not induce lactate or adenosine release and the increases in the HF EEG theta power during SD and SWS during the subsequent sleep were attenuated as compared to the young. These findings implicate that increased or continuous BF activity is important for active wake maintenance during SD as well as for the generation of homeostatic sleep pressure, and that in aging these mechanisms are impaired. Study II found that induction of the inducible NO synthase (iNOS) during SD is accompanied with activation of the AMP-activated protein kinase (AMPK) in the BF. Because decreased cellular energy charge is the most common cause for AMPK activation, this finding implicates that the BF is selectively sensitive to the metabolic demands of SD as increases were not found in the cortex. In aging (study III), iNOS expression and extracellular levels of NO and adenosine were not significantly increased during SD in the BF. Furthermore, infusion of NO donor into the BF did not lead to sleep promotion as it did in the young. These findings indicated that the NO (and adenosine) mediated sleep induction is impaired in aging and that it could at least partly be due to the reduced sensitivity of the BF to sleep-inducing factors. Taken together, these findings show that reduced sleep promotion by the BF contributes to the attenuated homeostatic sleep response in aging.

Sleep deprivation and brain energy metabolism : in vivo studies in rats and humans

Sleep deprivation leads to increased subsequent sleep length and depth and to deficits in cognitive performance in humans. In animals extreme sleep deprivation is eventually fatal. The cellular and molecular mechanisms causing the symptoms of sleep deprivation are unclear. This thesis was inspired by the hypothesis that during wakefulness brain energy stores would be depleted, and they would be replenished during sleep. The aim of this thesis was to elucidate the energy metabolic processes taking place in the brain during sleep deprivation. Endogenous brain energy metabolite levels were assessed in vivo in rats and in humans in four separate studies (Studies I-IV). In the first part (Study I) the effects of local energy depletion on brain energy metabolism and sleep were studied in rats with the use of in vivo microdialysis combined with high performance liquid chromatography. Energy depletion induced by 2,4-dinitrophenol infusion into the basal forebrain was comparable to the effects of sleep deprivation: both increased extracellular concentrations of adenosine, lactate, and pyruvate, and elevated subsequent sleep. This result supports the hypothesis of a connection between brain energy metabolism and sleep. The second part involved healthy human subjects (Studies II-IV). Study II aimed to assess the feasibility of applying proton magnetic resonance spectroscopy (1H MRS) to study brain lactate levels during cognitive stimulation. Cognitive stimulation induced an increase in lactate levels in the left inferior frontal gyrus, showing that metabolic imaging of neuronal activity related to cognition is possible with 1H MRS. Study III examined the effects of sleep deprivation and aging on the brain lactate response to cognitive stimulation. No physiologic, cognitive stimulation-induced lactate response appeared in the sleep-deprived and in the aging subjects, which can be interpreted as a sign of malfunctioning of brain energy metabolism. This malfunctioning may contribute to the functional impairment of the frontal cortex both during aging and sleep deprivation. Finally (Study IV), 1H MRS major metabolite levels in the occipital cortex were assessed during sleep deprivation and during photic stimulation. N-acetyl-aspartate (NAA/H2O) decreased during sleep deprivation, supporting the hypothesis of sleep deprivation-induced disturbance in brain energy metabolism. Choline containing compounds (Cho/H2O) decreased during sleep deprivation and recovered to alert levels during photic stimulation, pointing towards changes in membrane metabolism, and giving support to earlier observations of altered brain response to stimulation during sleep deprivation. Based on these findings, it can be concluded that sleep deprivation alters brain energy metabolism. However, the effects of sleep deprivation on brain energy metabolism may vary from one brain area to another. Although an effect of sleep deprivation might not in all cases be detectable in the non-stimulated baseline state, a challenge imposed by cognitive or photic stimulation can reveal significant changes. It can be hypothesized that brain energy metabolism during sleep deprivation is more vulnerable than in the alert state. Changes in brain energy metabolism may participate in the homeostatic regulation of sleep and contribute to the deficits in cognitive performance during sleep deprivation.

Concentrate feeding strategies for growing and finishing dairy bulls offered grass silage-based diets

The first aim of this thesis was to produce data for evaluating, developing and recommending biologically and economically efficient energy and protein feeding strategies for growing and finishing dairy bulls offered grass silage-based diets. The second aim was to calculate the energy and protein supplies of the dairy bulls fed different grass silage-cereal-based diets and, based on this, to estimate the possible need to revise the current Finnish energy and protein recommendations for growing dairy bulls. The third aim was to demonstrate the phosphorus supply of dairy bulls fed grass silage-cereal-based diets with or without protein supplementation in relation to current feeding recommendations for phosphorus. The results indicate that protein supplement is not needed for finishing dairy bulls (live weight more than 250 kg) when they are fed good-quality grass silage (digestible organic matter more than 650 g/kg dry matter, restricted fermentation with low concentrations of fermentation acids and ammonia N) and grain-based concentrate with a moderate (300-700 g/kg dry matter) concentrate level. The results also suggest that with total mixed ration feeding it is possible to use rather high concentrate proportions (700 g/kg dry matter) in feeding dairy bulls. According to this study, barley fibre is a suitable energy supplement with good-quality silage for growing dairy bulls. The results suggest that 50% of barley grain can be replaced with barley fibre without affecting growth. Also oats is a suitable energy supplement for dairy bulls. However, as a consequence of decreased energy intake, the gain and feed conversion of the bulls were slightly reduced in this study when barley grain was replaced by oats in the diet. Ultimately, the rationality of the use of barley fibre and oats in the future will depend on the price in relation to other concentrates. During the feeding experiments the calculated supply of energy was 10% higher than in the Finnish feeding recommendations for the present growth rate. This indicates that there is a need to update the Finnish feeding recommendations for dairy-breed growing bulls, and further calculations are needed for the energy supply of growing dairy bulls. The calculated supply of AAT (amino acids absorbed from the small intestine) was 38% higher than in the Finnish feeding recommendations for the present growth. Possibly, the present AAT-PBV system is not an optimal protein evaluation system for growing dairy bulls more than 250 kg live weight. The calculations based on the feeding experiments and the Finnish feeding recommendations indicate that in most cases the dairy bulls (live weight more than 250 kg) received enough P from the basic grass silage cereal-based diets without additional mineral feeds. Therefore there is no need to add P in the form of mineral mixtures.

Influence of harvesting strategy on nutrient supply and production of dairy cows consuming diets based on grass and red clover silage

The main objective of this thesis was to elucidate the effects of regrowth grass silage and red clover silage on nutrient supply and milk production of dairy cows as compared with primary growth grass silages. In the first experiment (publication I), two primary growth and four regrowth grass silages were harvested at two stages of growth. These six silages were fed to 24 lactating dairy cows with two levels of concentrate allowance. Silage intake and energy corrected milk yield (ECM) responses, and the range in these response variables between the diets, were smaller when regrowth silages rather than primary growth silages were fed. Milk production of dairy cows reflected the intake of metabolizable energy (ME), and no differences in the ME utilization were found between the diets based on silages harvested from primary growth and regrowth. The ECM response to increased concentrate allowance was, on average, greater when regrowth rather than primary growth silages were fed. In the second experiment (publication II), two silages from primary growth and two from regrowth used in I were fed to rumen cannulated lactating dairy cows. Cows consumed less feed dry matter (DM), energy and protein, and produced less milk, when fed diets based on regrowth silages rather than primary growth silages. Lower milk production responses of regrowth grass silage diets were mainly due to the lower silage DM intake, and could not be accounted for by differences in energy or protein utilization. Regrowth grass silage intake was not limited due to neutral detergent fibre (NDF) digestion or rumen fill or passage kinetics. However, lower intake may be at least partly attributable to plant diseases such as leaf spot infections, dead deteriorating material or abundance of weeds, which are all higher in regrowth compared with primary growth, and increase with advancing regrowth. In the third experiment (publications III and IV), red clover silages and grass silages harvested at two stages of growth, and a mixed diet of red clover and grass silages, were fed to five rumen cannulated lactating dairy cows. In spite of the lower average ME intake for red clover diets, the ECM production remained unchanged suggesting more efficient utilisation of ME for red clover diets compared with grass diets. Intake of N, and omasal canal flows of total non-ammonia N (NAN), microbial and non-microbial NAN were higher for red clover than for grass silage diets, but were not affected by forage maturity. Delaying the harvest tended to decrease DM intake of grass silage and increase that of red clover silage. The digestion rate of potentially digestible NDF was faster for red clover diets than for grass silage diets. Delaying the harvest decreased the digestion rate for grass but increased it for red clover silage diets. The low intake of early-cut red clover silage could not be explained by silage digestibility, fermentation quality, or rumen fill but was most likely related to the nutritionally suboptimal diet composition because inclusion of moderate quality grass silage in mixed diet increased silage DM intake. Despite the higher total amino acid supply of cows fed red clover versus grass silage diets, further milk production responses on red clover diets were possibly compromised by an inadequate supply of methionine as evidenced by lower methionine concentration in the amino acid profile of omasal digesta and plasma. Increasing the maturity of ensiled red clover does not seem to affect silage DM intake as consistently as that of grasses. The efficiency of N utilization for milk protein synthesis was lower for red clover diets than for grass diets. It was negatively related to diet crude protein concentration similarly to grass silage diets.

Urban energy fluxes in Helsinki and their high frequency spectral corrections

Inadvertent climate modification has led to an increase in urban temperatures compared to the surrounding rural area. The main reason for the temperature rise is the altered energy portioning of input net radiation to heat storage and sensible and latent heat fluxes in addition to the anthropogenic heat flux. The heat storage flux and anthropogenic heat flux have not yet been determined for Helsinki and they are not directly measurable. To the contrary, turbulent fluxes of sensible and latent heat in addition to net radiation can be measured, and the anthropogenic heat flux together with the heat storage flux can be solved as a residual. As a result, all inaccuracies in the determination of the energy balance components propagate to the residual term and special attention must be paid to the accurate determination of the components. One cause of error in the turbulent fluxes is the fluctuation attenuation at high frequencies which can be accounted for by high frequency spectral corrections. The aim of this study is twofold: to assess the relevance of high frequency corrections to water vapor fluxes and to assess the temporal variation of the energy fluxes. Turbulent fluxes of sensible and latent heat have been measured at SMEAR III station, Helsinki, since December 2005 using the eddy covariance technique. In addition, net radiation measurements have been ongoing since July 2007. The used calculation methods in this study consist of widely accepted eddy covariance data post processing methods in addition to Fourier and wavelet analysis. The high frequency spectral correction using the traditional transfer function method is highly dependent on relative humidity and has an 11% effect on the latent heat flux. This method is based on an assumption of spectral similarity which is shown not to be valid. A new correction method using wavelet analysis is thus initialized and it seems to account for the high frequency variation deficit. Anyhow, the resulting wavelet correction remains minimal in contrast to the traditional transfer function correction. The energy fluxes exhibit a behavior characteristic for urban environments: the energy input is channeled to sensible heat as latent heat flux is restricted by water availability. The monthly mean residual of the energy balance ranges from 30 Wm-2 in summer to -35 Wm-2 in winter meaning a heat storage to the ground during summer. Furthermore, the anthropogenic heat flux is approximated to be 50 Wm-2 during winter when residential heating is important.

Aspects of Inflationary Models at Low Energy Scales

Cosmological inflation is the dominant paradigm in explaining the origin of structure in the universe. According to the inflationary scenario, there has been a period of nearly exponential expansion in the very early universe, long before the nucleosynthesis. Inflation is commonly considered as a consequence of some scalar field or fields whose energy density starts to dominate the universe. The inflationary expansion converts the quantum fluctuations of the fields into classical perturbations on superhorizon scales and these primordial perturbations are the seeds of the structure in the universe. Moreover, inflation also naturally explains the high degree of homogeneity and spatial flatness of the early universe. The real challenge of the inflationary cosmology lies in trying to establish a connection between the fields driving inflation and theories of particle physics. In this thesis we concentrate on inflationary models at scales well below the Planck scale. The low scale allows us to seek for candidates for the inflationary matter within extensions of the Standard Model but typically also implies fine-tuning problems. We discuss a low scale model where inflation is driven by a flat direction of the Minimally Supersymmetric Standard Model. The relation between the potential along the flat direction and the underlying supergravity model is studied. The low inflationary scale requires an extremely flat potential but we find that in this particular model the associated fine-tuning problems can be solved in a rather natural fashion in a class of supergravity models. For this class of models, the flatness is a consequence of the structure of the supergravity model and is insensitive to the vacuum expectation values of the fields that break supersymmetry. Another low scale model considered in the thesis is the curvaton scenario where the primordial perturbations originate from quantum fluctuations of a curvaton field, which is different from the fields driving inflation. The curvaton gives a negligible contribution to the total energy density during inflation but its perturbations become significant in the post-inflationary epoch. The separation between the fields driving inflation and the fields giving rise to primordial perturbations opens up new possibilities to lower the inflationary scale without introducing fine-tuning problems. The curvaton model typically gives rise to relatively large level of non-gaussian features in the statistics of primordial perturbations. We find that the level of non-gaussian effects is heavily dependent on the form of the curvaton potential. Future observations that provide more accurate information of the non-gaussian statistics can therefore place constraining bounds on the curvaton interactions.

Formation of structure in dark energy cosmologies

Acceleration of the universe has been established but not explained. During the past few years precise cosmological experiments have confirmed the standard big bang scenario of a flat universe undergoing an inflationary expansion in its earliest stages, where the perturbations are generated that eventually form into galaxies and other structure in matter, most of which is non-baryonic dark matter. Curiously, the universe has presently entered into another period of acceleration. Such a result is inferred from observations of extra-galactic supernovae and is independently supported by the cosmic microwave background radiation and large scale structure data. It seems there is a positive cosmological constant speeding up the universal expansion of space. Then the vacuum energy density the constant describes should be about a dozen times the present energy density in visible matter, but particle physics scales are enormously larger than that. This is the cosmological constant problem, perhaps the greatest mystery of contemporary cosmology. In this thesis we will explore alternative agents of the acceleration. Generically, such are called dark energy. If some symmetry turns off vacuum energy, its value is not a problem but one needs some dark energy. Such could be a scalar field dynamically evolving in its potential, or some other exotic constituent exhibiting negative pressure. Another option is to assume that gravity at cosmological scales is not well described by general relativity. In a modified theory of gravity one might find the expansion rate increasing in a universe filled by just dark matter and baryons. Such possibilities are taken here under investigation. The main goal is to uncover observational consequences of different models of dark energy, the emphasis being on their implications for the formation of large-scale structure of the universe. Possible properties of dark energy are investigated using phenomenological paramaterizations, but several specific models are also considered in detail. Difficulties in unifying dark matter and dark energy into a single concept are pointed out. Considerable attention is on modifications of gravity resulting in second order field equations. It is shown that in a general class of such models the viable ones represent effectively the cosmological constant, while from another class one might find interesting modifications of the standard cosmological scenario yet allowed by observations. The thesis consists of seven research papers preceded by an introductory discussion.

Factors controlling the surface energy budget over snow and ice

Polar Regions are an energy sink of the Earth system, as the Sun rays do not reach the Poles for half of the year, and hit them only at very low angles for the other half of the year. In summer, solar radiation is the dominant energy source for the Polar areas, therefore even small changes in the surface albedo strongly affect the surface energy balance and, thus, the speed and amount of snow and ice melting. In winter, the main heat sources for the atmosphere are the cyclones approaching from lower latitudes, and the atmosphere-surface heat transfer takes place through turbulent mixing and longwave radiation, the latter dominated by clouds. The aim of this thesis is to improve the knowledge about the surface and atmospheric processes that control the surface energy budget over snow and ice, with particular focus on albedo during the spring and summer seasons, on horizontal advection of heat, cloud longwave forcing, and turbulent mixing during the winter season. The critical importance of a correct albedo representation in models is illustrated through the analysis of the causes for the errors in the surface and near-surface air temperature produced in a short-range numerical weather forecast by the HIRLAM model. Then, the daily and seasonal variability of snow and ice albedo have been examined by analysing field measurements of albedo, carried out in different environments. On the basis of the data analysis, simple albedo parameterizations have been derived, which can be implemented into thermodynamic sea ice models, as well as numerical weather prediction and climate models. Field measurements of radiation and turbulent fluxes over the Bay of Bothnia (Baltic Sea) also allowed examining the impact of a large albedo change during the melting season on surface energy and ice mass budgets. When high contrasts in surface albedo are present, as in the case of snow covered areas next to open water, the effect of the surface albedo heterogeneity on the downwelling solar irradiance under overcast condition is very significant, although it is usually not accounted for in single column radiative transfer calculations. To account for this effect, an effective albedo parameterization based on three-dimensional Monte Carlo radiative transfer calculations has been developed. To test a potentially relevant application of the effective albedo parameterization, its performance in the ground-based retrieval of cloud optical depth was illustrated. Finally, the factors causing the large variations of the surface and near-surface temperatures over the Central Arctic during winter were examined. The relative importance of cloud radiative forcing, turbulent mixing, and lateral heat advection on the Arctic surface temperature were quantified through the analysis of direct observations from Russian drifting ice stations, with the lateral heat advection calculated from reanalysis products.

Search for single top quark production in pp̅ collisions at √s=1.96 TeV in the missing transverse energy plus jets topology

We report a search for single top quark production with the CDF II detector using 2.1 fb-1 of integrated luminosity of pbar p collisions at sqrt{s}=1.96 TeV. The data selected consist of events characterized by large energy imbalance in the transverse plane and hadronic jets, and no identified electrons and muons, so the sample is enriched in W -> tau nu decays. In order to suppress backgrounds, additional kinematic and topological requirements are imposed through a neural network, and at least one of the jets must be identified as a b-quark jet. We measure an excess of signal-like events in agreement with the standard model prediction, but inconsistent with a model without single top quark production by 2.1 standard deviations (sigma), with a median expected sensitivity of 1.4 sigma. Assuming a top quark mass of 175 GeV/c2 and ascribing the excess to single top quark production, the cross section is measured to be 4.9+2.5-2.2(stat+syst)pb, consistent with measurements performed in independent datasets and with the standard model prediction.

Search for Supersymmetry with Gauge-Mediated Breaking in Diphoton Events with Missing Transverse Energy at CDF II

We present the results of a search for supersymmetry with gauge-mediated breaking and $\NONE\to\gamma\Gravitino$ in the $\gamma\gamma$+missing transverse energy final state. In 2.6$\pm$0.2 \invfb of $p{\bar p}$ collisions at $\sqrt{s}$$=$1.96 TeV recorded by the CDF II detector we observe no candidate events, consistent with a standard model background expectation of 1.4$\pm$0.4 events. We set limits on the cross section at the 95% C.L. and place the world's best limit of 149\gevc on the \none mass at $\tau_{\tilde{\chi}_1^0}$$