Accepted

Real-time quantitative reverse transcription-PCR (qRT-PCR ) is a highly sensitive molecular biology method based on the amplification of the cDNA of mRNA to detect and quantify the levels of mRNA of interest. In this chapter, we describe real-time qRT-PCR to detect and quantify mRNA of opioid receptors in immune cells. Specifically, we analyze mouse immune cells isolated from the blood and sciatic nerves exposed to a chronic constriction injury, which represents a model of neuropathic pain. We describe in detail the requirements and techniques to induce the chronic constriction injury, to isolate immune cells from the blood and injured nerves, to isolate the total RNA from immune cells, to perform a cDNA reverse transcription from the total RNA, and to perform real-time qRT-PCR for μ-, δ-, and κ-opioid receptor mRNAs.

This study aims to systematically evaluate the effect of non-invasive brain stimulation (NIBS) on neuropathic pain (NP) after spinal cord injury and compare the effects of two different NIBS.

K (KCNK) potassium channels form "background" or "leak" currents that have critical roles in cell excitability control in the brain, cardiovascular system, and somatosensory neurons. Similar to many ion channel families, studies of Ks have been limited by poor pharmacology. Of six K subfamilies, the thermo- and mechanosensitive TREK subfamily comprising K2.1 (TREK-1), K4.1 (TRAAK), and K10.1 (TREK-2) are the first to have structures determined for each subfamily member. These structural studies have revealed key architectural features that underlie K function and have uncovered sites residing at every level of the channel structure with respect to the membrane where small molecules or lipids can control channel function. This polysite pharmacology within a relatively small (~70 kDa) ion channel comprises four structurally defined modulator binding sites that occur above (Keystone inhibitor site), at the level of (K modulator pocket), and below (Fenestration and Modulatory lipid sites) the C-type selectivity filter gate that is at the heart of K function. Uncovering this rich structural landscape provides the framework for understanding and developing subtype-selective modulators to probe K function that may provide leads for drugs for anesthesia, pain, arrhythmia, ischemia, and migraine.

In attempts to understand the migraine patients' overall brain functional architecture, blood oxygenation level-dependent (BOLD) signals in the white matter (WM) and gray matter (GM) were considered in the current study. Migraine, a severe and multiphasic brain condition, is characterized by recurrent attacks of headaches. BOLD fluctuations in a resting state exhibit similar temporal and spectral profiles in both WM and GM. It is feasible to explore the functional interactions between WM tracts and GM regions in migraine. Forty-eight migraineurs without aura (MWoA) and 48 healthy controls underwent resting-state functional magnetic resonance imaging. Pearson's correlations between the mean time courses of 48 white matter (WM) bundles and 82 gray matter (GM) regions were computed for each subject. Two-sample -tests were performed on the Pearson's correlation coefficients (CC) to compare the differences between the MWoA and healthy controls in the GM-averaged CC of each bundle and the WM-averaged CC of each GM region. The MWoAs exhibited an overall decreased average temporal CC between BOLD signals in 82 GM regions and 48 WM bundles compared with healthy controls, while little was increased. In particular, WM bundles such as left anterior corona radiata, left external capsule and bilateral superior longitudinal fasciculus had significantly decreased mean CCs with GM in MWoA. On the other hand, 16 GM regions had significantly decreased mean CCs with WM in MWoA, including some areas that are parts of the somatosensory regions, auditory cortex, temporal areas, frontal areas, cingulate cortex, and parietal cortex. Decreased functional connections between WM bundles and GM regions might contribute to disrupted functional connectivity between the parts of the pain processing pathway in MWoAs, which indicated that functional and connectivity abnormalities in cortical regions may not be limited to GM regions but are instead associated with functional abnormalities in WM tracts.

The value of understanding patients' illness experience and social contexts for advancing medicine and clinical care is widely acknowledged. However, methodologies for rigorous and inclusive data gathering and integrative analysis of biomedical, cultural, and social factors are limited. In this paper, we propose a digital strategy for large-scale qualitative health research, using (as a state of being, a communication mode or context, and a set of imaginative, expressive, and game-like activities) as a research method for recursive learning and action planning. Our proposal builds on Gregory Bateson's cybernetic approach to knowledge production. Using chronic pain as an example, we show how pragmatic, structural and cultural constraints that define the relationship of patients to the healthcare system can give rise to conflicted messaging that impedes inclusive health research. We then review existing literature to illustrate how different types of play including games, chatbots, virtual worlds, and creative art making can contribute to research in chronic pain. Inspired by Frederick Steier's application of Bateson's theory to designing a science museum, we propose DiSPORA (Digital Strategy for Play-Oriented Research and Action), a virtual citizen science laboratory which provides a framework for delivering health information, tools for play-based experimentation, and data collection capacity, but is flexible in allowing participants to choose the mode and the extent of their interaction. Combined with other data management platforms used in epidemiological studies of neuropsychiatric illness, DiSPORA offers a tool for large-scale qualitative research, digital phenotyping, and advancing personalized medicine.