Silke Kreitz

Blockade of TNF-α rapidly inhibits pain responses in the central nervous system

There has been a consistent gap in understanding how TNF-α neutralization affects the disease state of arthritis patients so rapidly, considering that joint inflammation in rheumatoid arthritis is a chronic condition with structural changes. We thus hypothesized that neutralization of TNF-α acts through the CNS before directly affecting joint inflammation. Through use of functional MRI (fMRI), we demonstrate that within 24 h after neutralization of TNF-α, nociceptive CNS activity in the thalamus and somatosensoric cortex, but also the activation of the limbic system, is blocked. Brain areas showing blood-oxygen level-dependent signals, a validated method to assess neuronal activity elicited by pain, were significantly reduced as early as 24 h after an infusion of a monoclonal antibody to TNF-α. In contrast, clinical and laboratory markers of inflammation, such as joint swelling and acute phase reactants, were not affected by anti-TNF-α at these early time points. Moreover, arthritic mice overexpressing human TNF-α showed an altered pain behavior and a more intensive, widespread, and prolonged brain activity upon nociceptive stimuli compared with wild-type mice. Similar to humans, these changes, as well as the rewiring of CNS activity resulting in tight clustering in the thalamus, were rapidly reversed after neutralization of TNF-α. These results suggest that neutralization of TNF-α affects nociceptive brain activity in the context of arthritis, long before it achieves anti-inflammatory effects in the joints.

A Genome-wide Drosophila Screen for Heat Nociception Identifies α2δ3 as an Evolutionarily Conserved Pain Gene

Worldwide, acute, and chronic pain affects 20% of the adult population and represents an enormous financial and emotional burden. Using genome-wide neuronal-specific RNAi knockdown in Drosophila, we report a global screen for an innate behavior and identify hundreds of genes implicated in heat nociception, including the α2δ family calcium channel subunit straightjacket (stj). Mice mutant for the stj ortholog CACNA2D3 (α2δ3) also exhibit impaired behavioral heat pain sensitivity. In addition, in humans, α2δ3 SNP variants associate with reduced sensitivity to acute noxious heat and chronic back pain. Functional imaging in α2δ3 mutant mice revealed impaired transmission of thermal pain-evoked signals from the thalamus to higher-order pain centers. Intriguingly, in α2δ3 mutant mice, thermal pain and tactile stimulation triggered strong cross-activation, or synesthesia, of brain regions involved in vision, olfaction, and hearing.

Association of brain functional magnetic resonance activity with response to tumor necrosis factor inhibition in rheumatoid arthritis

OBJECTIVE To test whether brain activity predicts the response to tumor necrosis factor inhibitors (TNFi) in patients with rheumatoid arthritis (RA). Since clinical and laboratory parameters have proven unsuccessful in predicting response, we followed a radically different concept, hypothesizing that response to TNFi depends on central nervous system activity rather than the clinical signs of disease. METHODS Sequential testing by functional magnetic resonance imaging (MRI) of the brain, anatomic MRI of the hand, and clinical assessment of arthritis were carried out in 10 patients with active RA before and 3, 7, and 28 days after the start of TNFi treatment. RESULTS Baseline demographic and disease-specific parameters were identical in TNFi responders and nonresponders. The mean ± SEM decrease in the Disease Activity Score in 28 joints after 28 days was -1.8 ± 0.3 in TNFi responders (n = 5) and -0.2 ± 0.1 in nonresponders (n = 5). Responders showed significantly higher baseline activation in thalamic, limbic, and associative areas of the brain than nonresponders. Moreover, brain activity decreased within 3 days after TNFi exposure in the responders, preceding clinical responses (day 7) and responses observed on the anatomic hand MRI (day 28). CONCLUSION These data suggest that response to TNFi depends on brain activity in RA patients, reflecting the subjective perception of disease.

Substrate Utilization Patterns of Extractable and Non-Extractable Bacterial Fractions in Neutral and Acidic Beech Forest Soils

Substrate utilization patterns of four neutral and four acidic beech forest soils were examined using BIOLOG GN microplates. Soil bacteria were extracted using a three batch fractionated centrifugation procedure that produces two fractions. One contains the extracted and one the non-extractable bacteria and the fungi. Substrate utilization patterns of both fractions were determined with inhibition of fungal growth. All samples cluster distinctly into four different groups which correspond to the two fractions of the acidic and the neutral soils. Bacteria in acidic soils have generally a lower overall colour development indicating lower utilization activity. The extracted bacteria from acidic soils show a lower functional diversity than those extracted from neutral soils, but no difference in diversity was observed when the results of the extracted and the non-extractable bacteria of each soil were combined. In acidic soils substrate utilization abilities are heterogenously distributed between the two fractions. This effect mainly occurs in carbohydrate utilization. Bacterial communities in acidic soils are especially unable to degrade carboxylic acids. As decarboxylation is a main process of the initial decomposition of organic residues, this inability may be one reason for the retarded degradation of freshly fallen litter in acidic soils.

Manganese-Enhanced Magnetic Resonance Imaging for Mapping of Whole Brain Activity Patterns Associated with the Intake of Snack Food in Ad Libitum Fed Rats

Non-homeostatic hyperphagia, which is a major contributor to obesity-related hyperalimentation, is associated with the diet’s molecular composition influencing, for example, the energy content. Thus, specific food items such as snack food may induce food intake independent from the state of satiety. To elucidate mechanisms how snack food may induce non-homeostatic food intake, it was tested if manganese-enhanced magnetic resonance imaging (MEMRI) was suitable for mapping the whole brain activity related to standard and snack food intake under normal behavioral situation. Application of the MnCl2 solution by osmotic pumps ensured that food intake was not significantly affected by the treatment. After z-score normalization and a non-affine three-dimensional registration to a rat brain atlas, significantly different grey values of 80 predefined brain structures were recorded in ad libitum fed rats after the intake of potato chips compared to standard chow at the group level. Ten of these areas had previously been connected to food intake, in particular to hyperphagia (e.g. dorsomedial hypothalamus or the anterior paraventricular thalamic nucleus) or to the satiety system (e.g. arcuate hypothalamic nucleus or solitary tract); 27 areas were related to reward/addiction including the core and shell of the nucleus accumbens, the ventral pallidum and the ventral striatum (caudate and putamen). Eleven areas associated to sleep displayed significantly reduced Mn2+-accumulation and six areas related to locomotor activity showed significantly increased Mn2+-accumulation after the intake of potato chips. The latter changes were associated with an observed significantly higher locomotor activity. Osmotic pump-assisted MEMRI proved to be a promising technique for functional mapping of whole brain activity patterns associated to nutritional intake under normal behavior.

Combining functional magnetic resonance imaging with mouse genomics: new options in pain research.

This functional magnetic imaging study investigated the functional implications of genetic modification and pharmacological intervention on cerebral processing of heat-induced nociception in mice. Comparing dynorphin-overexpressing dream−/− with wild-type mice, smaller activated cortical and limbic brain structure sizes could be observed. Moreover, significantly reduced blood oxygenation level-dependent signal amplitudes were found in pain-related brain structures: sensory input, thalamic regions, sensory cortex, limbic system, basal ganglia, hypothalamus and periaqueductal grey. Administration of the specific κ-opioid-receptor antagonist nor-binaltorphimine to dream−/− mice reversed this reduction to wild-type level in the same brain structures.These results show that blood oxygenation level-dependent functional magnetic imaging in the pain system of (transgenic) mice is feasible. Genetic modifications and pharmacological interventions modify brain responses in a structure-specific manner.

Fat/carbohydrate ratio but not energy density determines snack food intake and activates brain reward areas

The snack food potato chips induces food intake in ad libitum fed rats, which is associated with modulation of the brain reward system and other circuits. Here, we show that food intake in satiated rats is triggered by an optimal fat/carbohydrate ratio. Like potato chips, an isocaloric fat/carbohydrate mixture influenced whole brain activity pattern of rats, affecting circuits related e.g. to reward/addiction, but the number of modulated areas and the extent of modulation was lower compared to the snack food itself.

A New Analysis of Resting State Connectivity and Graph Theory Reveals Distinctive Short-Term Modulations due to Whisker Stimulation in Rats

Resting state (RS) connectivity has been increasingly studied in healthy and diseased brains in humans and animals. This paper presents a new method to analyze RS data from fMRI that combines multiple seed correlation analysis with graph-theory (MSRA). We characterize and evaluate this new method in relation to two other graph-theoretical methods and ICA. The graph-theoretical methods calculate cross-correlations of regional average time-courses, one using seed regions of the same size (SRCC) and the other using whole brain structure regions (RCCA). We evaluated the reproducibility, power, and capacity of these methods to characterize short-term RS modulation to unilateral physiological whisker stimulation in rats. Graph-theoretical networks found with the MSRA approach were highly reproducible, and their communities showed large overlaps with ICA components. Additionally, MSRA was the only one of all tested methods that had the power to detect significant RS modulations induced by whisker stimulation that are controlled by family-wise error rate (FWE). Compared to the reduced resting state network connectivity during task performance, these modulations implied decreased connectivity strength in the bilateral sensorimotor and entorhinal cortex. Additionally, the contralateral ventromedial thalamus (part of the barrel field related lemniscal pathway) and the hypothalamus showed reduced connectivity. Enhanced connectivity was observed in the amygdala, especially the contralateral basolateral amygdala (involved in emotional learning processes). In conclusion, MSRA is a powerful analytical approach that can reliably detect tiny modulations of RS connectivity. It shows a great promise as a method for studying RS dynamics in healthy and pathological conditions.

Functional Atlas of the Rat Brain

For functional mapping of human brain activity the Talairach atlas provides the spatial reference system for structure identification of activated spots. No such common reference system exists for fMRI studies on animals. Therefore, the aim of this study was to establish a set of different image processing algorithms in order to generate for a given stimulation paradigm a functional standard atlas of the rat brain. The goal, which should be achieved, was an automatic identification and structure assignment of activated voxel groups based on such a labelled standard atlas. Due to the smooth brain structure of rodents even basic affine registration techniques greatly reduce the interindividual variations in brains and activated structures and allowed precise identification and labelling of the activated structures.

Direct enumeration and size classification of soil bacteria using confocal laser scanning microscopy and image analysis

Direct measurement of fluorescently stained soil bacteria is a common method in microbiology. The tedium of making such measurements visually has prompted the use of automatic image analysis. Accurate measurements by image analysis require an reliable method of image processing that eliminates background particles and selects an appropriate threshold for automated counts and measurements. The authors tested two processing methods, a sharpening filter and a background correction, with subsequent threshold setting and direct threshold setting without previous image processing. The threshold was chosen according to the first maximum of the first derivative of the characteristic line of one sample. The background correction method was found to be superior for analyzing images of soil bacteria, although it leads to slight underestimation of the count as well as the area measurement.