Remote and reversible inhibition of neurons and circuits by small molecule induced potassium channel stabilization

Manipulating the function of neurons and circuits that translate electrical and chemical signals into behavior represents a major challenges in neuroscience. In addition to optogenetic methods using light-activatable channels, pharmacogenetic methods with ligand induced modulation of cell signaling and excitability have been developed. However, they are largely based on ectopic expression of exogenous or chimera proteins. Now, we describe the remote and reversible expression of a Kir2.1 type potassium channel using the chemogenetic technique of small molecule induced protein stabilization. Based on shield1-mediated shedding of a destabilizing domain fused to a protein of interest and inhibition of protein degradation, this principle has been adopted for biomedicine, but not in neuroscience so far. Here, we apply this chemogenetic approach in brain research for the first time in order to control a potassium channel in a remote and reversible manner. We could show that shield1-mediated ectopic Kir2.1 stabilization induces neuronal silencing in vitro and in vivo in the mouse brain. We also validated this novel pharmacogenetic method in different neurobehavioral paradigms. The DD-Kir2.1 may complement the existing portfolio of pharmaco- and optogenetic techniques for specific neuron manipulation, but it may also provide an example for future applications of this principle in neuroscience research

Time and dosage effects of meprobamate on visual detection

Sixteen male subjects, 21 years and older, were run in a doubleblind own-control design under three dosages of meprobamate (400, 800 and 1600 mg) and matching placebo, to determine response curves on a visual detection task over a four and one-half hour period, from 5:00 to 9:30 p.m. To control for possible daily cycle effects two additional runs under placebo and 1600 mg meprobamate were conducted from 8:00 a.m. to 12:30 p.m.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/46406/1/213_2004_Article_BF00710913.pd

Induction of erythroferrone in healthy humans by micro-dose recombinant erythropoietin or high-altitude exposure

The erythropoietin (Epo)-erythroferrone (ERFE)-hepcidin axis coordinates erythropoiesis and iron homeostasis. While mouse studies have established that Epo-induced ERFE production represses hepcidin synthesis by inhibiting hepatic BMP/SMAD signaling, evidence for the role of ERFE in humans is limited. To investigate the role of ERFE as a physiological erythroid regulator in humans, we conducted two studies: first, 24 males received six injections of saline (placebo), recombinant Epo (rhEpo) 20 UI kg-1 (micro-dose) or 50 UI kg-1 (low-dose). Second, we quantified ERFE in 22 subjects exposed to high altitude (3800 m) for 15 hours. In the first study, total hemoglobin mass (Hbmass) increased after low- but not after micro-dose injections, when compared to placebo. Serum ERFE levels were enhanced by rhEpo, remaining higher than after placebo for 48 (micro-dose) or 72 hours (low-dose) post-injections. Conversely, hepcidin levels decreased when Epo and ERFE arose, before any changes in serum iron parameters occurred. In the second study, serum Epo and ERFE increased at high altitude. The present results demonstrate that in healthy humans ERFE responds to slightly increased Epo levels not associated with Hbmass expansion and down-regulates hepcidin in an apparently iron-independent way. Notably, ERFE flags micro-dose Epo, thus holding promise as novel anti-doping biomarker

The brain as 'immunoprecipitator' of serum autoantibodies against N-Methyl-D-Aspartate receptor subunit NR1

Autoantibodies (AB) against N-methyl-D-aspartate receptor subunit NR1 (NMDAR1) are highly seroprevalent in health and disease. Symptomatic relevance may arise upon compromised blood-brain barrier (BBB). However, it remained unknown whether circulating NMDAR1 AB appear in the cerebrospinal fluid (CSF). Of n5271 subjects with CSF-serum pairs, 26 were NMDAR1 AB seropositive, but only 1 was CSF positive. Contrariwise, tetanus AB (non-brain-binding) were present in serum and CSF of all subjects, with CSF levels higher upon BBB dysfunction. Translational mouse experiments proved the hypothesis that the brain acts as an 'immunoprecipitator'; simultaneous injection of NMDAR1 AB and the non-brain-binding green fluorescent protein AB resulted in high detectability of the former in brain and the latter in CSF

Lipid raft integrity affects GABAA receptor, but not NMDA receptor modulation by psychopharmacological compounds

Lipid rafts have been shown to play an important role for G-protein mediated signal transduction and the function of ligand-gated ion channels including their modulation by psychopharmacological compounds. In this study, we investigated the functional significance of the membrane distribution of NMDA and GABAA receptor subunits in relation to the accumulation of the tricyclic antidepressant desipramine (DMI) and the benzodiazepine diazepam (Diaz). In the presence of Triton X-100, which allowed proper separation of the lipid raft marker proteins caveolin-1 and flotillin-1 from the transferrin receptor, all receptor subunits were shifted to the non-raft fractions. In contrast, under detergent-free conditions, NMDA and GABAA receptor subunits were detected both in raft and non-raft fractions. Diaz was enriched in non-raft fractions without Triton X-100 in contrast to DMI, which preferentially accumulated in lipid rafts. Impairment of lipid raft integrity by methyl-β-cyclodextrine (MβCD)-induced cholesterol depletion did not change the inhibitory effect of DMI at the NMDA receptor, whereas it enhanced the potentiating effect of Diaz at the GABAA receptor at non-saturating concentrations of GABA. These results support the hypothesis that the interaction of benzodiazepines with the GABAA receptor likely occurs outside of lipid rafts while the antidepressant DMI acts on ionotropic receptors both within and outside these membrane microdomains

Towards the clinical implementation of pharmacogenetics in bipolar disorder.

BackgroundBipolar disorder (BD) is a psychiatric illness defined by pathological alterations between the mood states of mania and depression, causing disability, imposing healthcare costs and elevating the risk of suicide. Although effective treatments for BD exist, variability in outcomes leads to a large number of treatment failures, typically followed by a trial and error process of medication switches that can take years. Pharmacogenetic testing (PGT), by tailoring drug choice to an individual, may personalize and expedite treatment so as to identify more rapidly medications well suited to individual BD patients.DiscussionA number of associations have been made in BD between medication response phenotypes and specific genetic markers. However, to date clinical adoption of PGT has been limited, often citing questions that must be answered before it can be widely utilized. These include: What are the requirements of supporting evidence? How large is a clinically relevant effect? What degree of specificity and sensitivity are required? Does a given marker influence decision making and have clinical utility? In many cases, the answers to these questions remain unknown, and ultimately, the question of whether PGT is valid and useful must be determined empirically. Towards this aim, we have reviewed the literature and selected drug-genotype associations with the strongest evidence for utility in BD.SummaryBased upon these findings, we propose a preliminary panel for use in PGT, and a method by which the results of a PGT panel can be integrated for clinical interpretation. Finally, we argue that based on the sufficiency of accumulated evidence, PGT implementation studies are now warranted. We propose and discuss the design for a randomized clinical trial to test the use of PGT in the treatment of BD

Behavioral effects of chronic administration of psychoactive drugs to anxious patients

The effects of chronic administration of (a) 1600 mg. meprobamate, (b) five Tranquil tablets (the maximum recommended daily dose), and (c) placebos were examined, in a counterbalanced partially blinded design. Thirty-two subjects, including 23 anxious patients, employed as their own controls, were given behavioral tests, psychiatric interviews, and ratings at the end of each 21 day treatment period.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/46328/1/213_2004_Article_BF00409114.pd

Two Host Factors Regulate Persistence of H7a-Specific T Cells Injected in Tumor-Bearing Mice

BACKGROUND: Injection of CD8 T cells primed against immunodominant minor histocompatibility antigens (MiHA) such as H7(a) can eradicate leukemia and solid tumors. To understand why MiHA-targeted T cells have such a potent antitumor effect it is essential to evaluate their in vivo behavior. In the present work, we therefore addressed two specific questions: what is the proliferative dynamics of H7(a)-specifc T cells in tumors, and do H7(a)-specific T cells persist long-term after adoptive transfer? METHODOLOGY/PRINCIPAL FINDINGS: By day 3 after adoptive transfer, we observed a selective infiltration of melanomas by anti-H7(a) T cells. Over the next five days, anti-H7(a) T cells expanded massively in the tumor but not in the spleen. Thus, by day 8 after injection, anti-H7(a) T cells in the tumor had undergone more cell divisions than those in the spleen. These data strongly suggest that anti-H7(a) T cells proliferate preferentially and extensively in the tumors. We also found that two host factors regulated long-term persistence of anti-H7(a) memory T cells: thymic function and expression of H7(a) by host cells. On day 100, anti-H7(a) memory T cells were abundant in euthymic H7(a)-negative (B10.H7(b)) mice, present in low numbers in thymectomized H7(a)-positive (B10) hosts, and undetectable in euthymic H7(a)-positive recipients. CONCLUSIONS/SIGNIFICANCE: Although in general the tumor environment is not propitious to T-cell invasion and expansion, the present work shows that this limitation may be overcome by adoptive transfer of primed CD8 T cells targeted to an immunodominant MiHA (here H7(a)). At least in some cases, prolonged persistence of adoptively transferred T cells may be valuable for prevention of late cancer relapse in adoptive hosts. Our findings therefore suggest that it may be advantageous to target MiHAs with a restricted tissue distribution in order to promote persistence of memory T cells and thereby minimize the risk of cancer recurrence