Bottom-Up versus Top-Down Induction of Sleep by Zolpidem Acting on Histaminergic and Neocortex Neurons

Zolpidem, a GABAA receptor-positive modulator, is the gold-standard drug for treating insomnia. Zolpidem prolongs IPSCs to decrease sleep latency and increase sleep time, effects that depend on α2 and/or α3 subunit-containing receptors. Compared with natural NREM sleep, zolpidem also decreases the EEG power, an effect that depends on α1 subunit-containing receptors, and which may make zolpidem-induced sleep less optimal. In this paper, we investigate whether zolpidem needs to potentiate only particular GABAergic pathways to induce sleep without reducing EEG power. Mice with a knock-in F77I mutation in the GABAA receptor γ2 subunit gene are zolpidem-insensitive. Using these mice, GABAA receptors in the frontal motor neocortex and hypothalamic (tuberomammillary nucleus) histaminergic-neurons of γ2I77 mice were made selectively sensitive to zolpidem by genetically swapping the γ2I77 subunits with γ2F77 subunits. When histamine neurons were made selectively zolpidem-sensitive, systemic administration of zolpidem shortened sleep latency and increased sleep time. But in contrast to the effect of zolpidem on wild-type mice, the power in the EEG spectra of NREM sleep was not decreased, suggesting that these EEG power-reducing effects of zolpidem do not depend on reduced histamine release. Selective potentiation of GABAA receptors in the frontal cortex by systemic zolpidem administration also reduced sleep latency, but less so than for histamine neurons. These results could help with the design of new sedatives that induce a more natural sleep

GABA Receptors and the Pharmacology of Sleep

Current GABAergic sleep-promoting medications were developed pragmatically, without making use of the immense diversity of GABAA receptors. Pharmacogenetic experiments are leading to an understanding of the circuit mechanisms in the hypothalamus by which zolpidem and similar compounds induce sleep at α2βγ2-type GABAA receptors. Drugs acting at more selective receptor types, for example, at receptors containing the α2 and/or α3 subunits expressed in hypothalamic and brain stem areas, could in principle be useful as hypnotics/anxiolytics. A highly promising sleep-promoting drug, gaboxadol, which activates αβδ-type receptors failed in clinical trials. Thus, for the time being, drugs such as zolpidem, which work as positive allosteric modulators at GABAA receptors, continue to be some of the most effective compounds to treat primary insomnia

Cryogels with Affinity Ligands as Tools in Protein Purification

Affinity chromatography is one of the well-known separation techniques especially if high purity is desired. Introducing ligands on monolithic structure gives the possibility for purifying complex media such as plasma and crude extract. This chapter is focusing on the preparation of cryogels as monolithic column and immobilization of concanavalin A on its surface as ligand for capturing the glycoprotein horseradish peroxidase

Effects of increasing dietary protein and fibre intake with lupin on body weight and composition and blood lipids in overweight men and women

Background: Lupin kernel flour (LKF) is a novel food ingredient that is high in protein and fibre. We have previously shown that partial substitution of refined wheat-derived carbohydrate in bread with protein and fibre from LKF can reduce appetite and energy intake acutely. In addition, several studies have suggested that lupin may reduce cholesterol concentrations and benefit glucose and insulin metabolism. Aim: The aim of this study was to investigate the effects on body weight and composition and blood lipids, glucose and insulin of an ad libitum LKF-enriched diet higher in dietary protein and fibre. Subjects and methods: A total of 88 overweight and obese men and women were recruited for a 16-week parallel-design randomized controlled trial. Participants replaced 15–20% of their usual daily energy intake with white bread (control) or LKF-enriched bread (lupin) in an ad libitum diet. Measurements of body weight and composition, and fasting blood biochemical measurements were performed at baseline and 16 weeks. The primary analysis included 74 participants (37 per group) who completed the intervention. Results: At baseline, mean (±s.d.) body mass index and total cholesterol were 30.6±3.5 kg m−2 and 5.37±0.94 mmol l−1, respectively. Estimated (mean between-group difference (95% confidence interval)) protein (13.7 (2.28, 25.0) g per day) and fibre (12.5 (8.79, 16.2) g per day) intakes were higher during the intervention with lupin than with control. For lupin relative to control, the net effects on body weight (−0.4 (−1.3, 0.6) kg), fat mass (−0.5 (−1.1, 0.2) kg) and percentage (−0.5 (−1.1, 0.1)%), plasma leptin (−1.66 (−4.91, 1.59) ng ml−1) and adiponectin (0.20 (−0.73, 1.13) mg l−1, as well as serum total cholesterol (−0.08 (−0.38, 0.22) mmol l−1), triglycerides (0.09 (−0.10, 0.21) mmol l−1), glucose (0.10 (−0.11, 0.30) mmol l−1) and insulin (0.40 (−1.20, 2.00) mU l−1) were not significant. Conclusions: This study does not support the proposal that an ad libitum diet enriched in LKF resulting in moderate changes in both protein and fibre intakes can benefit body weight and composition or fasting blood lipids, glucose and insulin concentrations in overweight men and women with mildly elevated total cholesterol concentrations

Bone tissue engineering: Past-present-future

Bone is one of the few tissues to display a true potential for regeneration. Fracture healing is an obvious example where regeneration occurs through tightly regulated sequences of molecular and cellular events which recapitulate tissue formation seen during embryogenesis. Still in some instances, bone regeneration does not occur properly (i.e. critical size lesions) and an appropriate therapeutic intervention is necessary. Successful replacement of bone by tissue engineering will likely depend on the recapitulation of this flow of events. In fact, bone regeneration requires cross-talk between microenvironmental factors and cells; for example, resident mesenchymal progenitors are recruited and properly guided by soluble and insoluble signaling molecules. Tissue engineering attempts to reproduce and to mimic this natural milieu by delivering cells capable of differentiating into osteoblasts, inducing growth factors and biomaterials to support cellular attachment, proliferation, migration, and matrix deposition. In the last two decades, a significant effort has been made by the scientific community in the development of methods and protocols to repair and regenerate tissues such as bone, cartilage, tendons, and ligaments. In this same period, great advancements have been achieved in the biology of stem cells and on the mechanisms governing \u201cstemness\u201d. Unfortunately, after two decades, effective clinical translation does not exist, besides a few limited examples. Many years have passed since cell-based regenerative therapies were first described as \u201cpromising approaches\u201d, but this definition still engulfs the present literature. Failure to envisage translational cell therapy applications in routine medical practice evidences the existence of unresolved scientific and technical struggles, some of which still puzzle researchers in the field and are presented in this chapter