Legal Requirements for Admission to Public Schools

Advanced driver assistance systems for heavy duty vehicles, such as lookahead cruise and gearshift controllers, rely on high quality map data. Current digital maps do not offer the required level of road grade information. This contribution presents an algorithm for on-board road grade estimation based on fusion of GPS and vehicle sensor data with measurements from previous runs over the same road segment. An incremental update scheme is utilized to ensure that data storage requirements are independent of the number of measurement runs. Results of the implemented system based on six traversals of a known road with three different vehicles are presented.QC 20120216</p

Presumptions—The Uniform Rules in the Federal Courts

Emerging heavy duty vehicle control systems increasingly rely on advance knowledge of the road topography, described by the longitudinal road grade. Highway road grade profiles are restricted by road design specifications to be piecewise affine. This characteristic is used herein to derive a method for road grade estimation based on standard on-vehicle sensors and optimal piecewise linear estimation through dynamic programming. The proposed method is demonstrated with on-road experiments. It is able to represent the road grade profile for two studied 15 km road sections, by 20 linear segments for each, with a root mean square error between 0.42 % and 0.55 % grade.QC 20120215</p

Voltage sensitivity of dopamine D2-like receptors

G protein coupled receptors (GPCRs) mediate a multitude of responses serving hormonal, neurotransmitter, and sensory functions. These receptors are important drug targets; in fact, about 27 % of prescribed drugs are GPCR ligands. The dopamine D2 receptor is prominently expressed within the CNS as two distinct isoforms; D2L (long isoform) and D2S (short isoform). The former is mainly expressed postsynaptically, whereas the latter functions primarily as an inhibitory auto- and heteroreceptor. The D2 receptor is of considerable pharmacological interest, as it constitutes the main target for most antiparkinsonian and antipsychotic drugs in clinical use. While many ion channels have long been known to be voltage sensitive, this property has not been attributed to GPCRs until quite recently. As a notable example, the muscarinic M2 receptor was shown to display depolarization-induced decreases in agonist binding and functional potency. M2 receptor voltage sensitivity has been implicated in the autoreceptor function of this GPCR, by permitting rapid control of neurotransmitter release kinetics by membrane voltage. The present work investigated the voltage sensitivities of the three D2-like dopamine receptors; D2, D3, and D4. The bulk of the experiments were carried out in Xenopus oocytes heterologously expressing D2-like receptors with G protein-coupled inwardly rectifying potassium channel (GIRK) subunits. GIRK channels are activated by Gβγ subunits from inhibitory G proteins and were used as readout of receptor activation. It was found that dopamine potency was reduced by depolarization to a similar extent at both isoforms of the D2 receptor. However, at the dopamine D3 receptor dopamine potency was not significantly affected, while a weak, albeit significant potency decrease was observed at the dopamine D4 receptor. Moreover, in mammalian cells expressing fluorescent G protein subunits, changes in inter-subunit Förster Resonance Energy Transfer (FRET) were used as readout of D2S receptor activation. Determination of dopamine concentration-response relationships in single cells under simultaneous patch clamp revealed similar depolarization-induced potency shifts as when studying GIRK channel activation in oocytes. Furthermore, radioligand binding experiments carried out on oocytes in hyperpolarizing vs. depolarizing buffer established that dopamine binding is reduced by depolarization. Interestingly, the effect of voltage was different for different agonists at the D2S receptor, including efficacious, high-affinity antiparkinsonian agonists. This agonist-specificity did not reflect selective signalling via distinct G protein subtypes. However, contacts between agonist hydroxyl groups and receptor serine residues, as well as between the agonist amine group and a conserved aspartate residue, were found to be important for the voltage induced potency shift of phenethylamine agonists, such as dopamine. In conclusion, the findings presented in this thesis suggest that the dopamine D2-like receptors are differentially affected by voltage. At the D2S receptor, specific agonistreceptor interactions determine the effect of the receptor’s voltage sensitivity on agonist potency and efficacy. This information demonstrates the relevance of GPCR voltage sensitivity to dopaminergic signalling, reveals new details about the mechanism of voltage sensitive agonism, and points to the possibility of using differentially voltagemodulated agonists to investigate the relevance of this phenomenon in native tissue

Dopamine-induced arrestin recruitment and desensitization of the dopamine D4 receptor is regulated by G protein-coupled receptor kinase-2

The dopamine D4 receptor (D4R) is expressed in the retina, prefrontal cortex, and autonomic nervous system and has been implicated in attention deficit hyperactivity disorder (ADHD), substance use disorders, and erectile dysfunction. D4R has also been investigated as a target for antipsychotics due to its high affinity for clozapine. As opposed to the closely related dopamine D2 receptor (D2R), dopamine-induced arrestin recruitment and desensitization at the D4R have not been studied in detail. Indeed, some earlier investigations could not detect arrestin recruitment and desensitization of this receptor upon its activation by agonist. Here, we used a novel nanoluciferase complementation assay to study dopamine-induced recruitment of β-arrestin2 (βarr2; also known as arrestin3) and G protein-coupled receptor kinase-2 (GRK2) to the D4R in HEK293T cells. We also studied desensitization of D4R-evoked G protein-coupled inward rectifier potassium (GIRK; also known as Kir3) current responses in Xenopus oocytes. Furthermore, the effect of coexpression of GRK2 on βarr2 recruitment and GIRK response desensitization was examined. The results suggest that coexpression of GRK2 enhanced the potency of dopamine to induce βarr2 recruitment to the D4R and accelerated the rate of desensitization of D4R-evoked GIRK responses. The present study reveals new details about the regulation of arrestin recruitment to the D4R and thus increases our understanding of the signaling and desensitization of this receptor

Pridopidine Reverses Phencyclidine-Induced Memory Impairment

Pridopidine is in clinical trials for Huntington's diseasetreatment. Originally developedas a dopamine D2receptor (D2R) ligand, pridopidine displays about 100-fold higheraffinity for the sigma-1 receptor (sigma-1R). Interestingly, pridopidine slows diseaseprogression and improves motor function in Huntington's disease model mice and,in preliminarily reports, Huntington's disease patients.The present study examinedthe anti-amnesic potential of pridopidine. Thus, memory impairment was produced inmice by administration of phencyclidine (PCP, 10 mg/kg/day) for 10 days, followedby 14 days' treatment with pridopidine (6 mg/kg/day), or saline. Finally, novel objectrecognition performance was assessed in the animals. Mice receiving PCP andsaline exhibited deficits in novel object recognition, as expected, while pridopidinetreatment counteracted PCP-induced memory impairment. The effect of pridopidine wasattenuated by co-administration of the sigma receptor antagonist, NE-100 (10 mg/kg).Our results suggest that pridopidine exerts anti-amnesic and potentially neuroprotectiveactions. These data provide new insights into the therapeutic potential of pridopidine asa pro-cognitive drug

Inhibition of tryptophan hydroxylases and monoamino oxidase-A by the proton pump inhibitor, omeprazole - in vitro and in vivo investigations

Serotonin (5-HT) is a hormone and neurotransmitter that modulates neural activity as well as a wide range of other physiological processes including cardiovascular function, bowel motility, and platelet aggregation. 5-HT synthesis is catalyzed by tryptophan hydroxylase (TPH) which exists as two distinct isoforms; TPH1 and TPH2, which are responsible for peripheral and central 5-HT, respectively. Due to the implication of 5-HT in a number of pathologies, including depression, anxiety, autism, sexual dysfunction, irritable bowel syndrome, inflammatory bowel disease, and carcinoid syndrome, there has been a growing interest in finding modulators of these enzymes in recent years. We thus performed high-throughput screening (HTS) using a fluorescence-based thermal shift assay (DSF) to search the Prestwick Chemical Library containing 1,280 compounds, mostly FDA-approved drugs, for TPH1 binders. We here report the identification of omeprazole, a proton pump inhibitor, as an inhibitor of TPH1 and TPH2 with low micromolar potency and high selectivity over the other aromatic amino acid hydroxylases. The S-enantiomer of omeprazole, esomeprazole, has recently also been described as an inhibitor of monoamine oxidase-A (MAO-A), the main enzyme responsible for 5-HT degradation, albeit with lower potency compared to the effect on TPH1 and TPH2. In order to investigate the net effect of simultaneous inhibition of TPH and MAO-A in vivo, we administered high-dose (100 mg/kg) omeprazole to CD-1 mice for 4 days, after which the animals were subjected to the tail suspension test. Finally, central (whole brain) and peripheral (serum) 5-HT content was measured using liquid chromatography-mass spectrometry (LC-MS). Omeprazole treatment significantly increased 5-HT concentrations, both in brain and in serum, and reduced the time spent immobile in the tail suspension test relative to vehicle control. Thus, the MAO-A inhibition afforded by high-dose omeprazole appears to overcome the opposing effect on 5-HT produced by inhibition of TPH1 and TPH2. Further modification of proton pump inhibitor scaffolds may yield more selective modulators of 5-HT metabolism

A truncated Kv1.1 protein in the brain of the megencephaly mouse: expression and interaction

BACKGROUND: The megencephaly mouse, mceph/mceph, is epileptic and displays a dramatically increased brain volume and neuronal count. The responsible mutation was recently revealed to be an eleven base pair deletion, leading to a frame shift, in the gene encoding the potassium channel Kv1.1. The predicted MCEPH protein is truncated at amino acid 230 out of 495. Truncated proteins are usually not expressed since nonsense mRNAs are most often degraded. However, high Kv1.1 mRNA levels in mceph/mceph brain indicated that it escaped this control mechanism. Therefore, we hypothesized that the truncated Kv1.1 would be expressed and dysregulate other Kv1 subunits in the mceph/mceph mice. RESULTS: We found that the MCEPH protein is expressed in the brain of mceph/mceph mice. MCEPH was found to lack mature (Golgi) glycosylation, but to be core glycosylated and trapped in the endoplasmic reticulum (ER). Interactions between MCEPH and other Kv1 subunits were studied in cell culture, Xenopus oocytes and the brain. MCEPH can form tetramers with Kv1.1 in cell culture and has a dominant negative effect on Kv1.2 and Kv1.3 currents in oocytes. However, it does not retain Kv1.2 in the ER of neurons. CONCLUSION: The megencephaly mice express a truncated Kv1.1 in the brain, and constitute a unique tool to study Kv1.1 trafficking relevant for understanding epilepsy, ataxia and pathologic brain overgrowth

A Neanderthal Sodium Channel Increases Pain Sensitivity in Present-Day Humans.

The sodium channel Nav1.7 is crucial for impulse generation and conduction in peripheral pain pathways [1]. In Neanderthals, the Nav1.7 protein carried three amino acid substitutions (M932L, V991L, and D1908G) relative to modern humans. We expressed Nav1.7 proteins carrying all combinations of these substitutions and studied their electrophysiological effects. Whereas the single amino acid substitutions do not affect the function of the ion channel, the full Neanderthal variant carrying all three substitutions, as well as the combination of V991L with D1908G, shows reduced inactivation, suggesting that peripheral nerves were more sensitive to painful stimuli in Neanderthals than in modern humans. We show that, due to gene flow from Neanderthals, the three Neanderthal substitutions are found in ∼0.4% of present-day Britons, where they are associated with heightened pain sensitivity

Pridopidine selectively occupies sigma-1 rather than dopamine D2 receptors at behaviorally active doses

Dopamine stabilizers have stimulatory actions under low dopamine tone and inhibitory actions under high dopamine tone without eliciting catalepsy. These compounds are dopamine D-2 receptor (D2R) antagonists or weak partial agonists and may have pro-mnemonic and neuroprotective effects. The mechanism underlying their stimulatory and neuroprotective actions is unknown but could involve sigma-1R binding. The present study examined sigma-1R and D2R occupancy by the dopamine stabilizer pridopidine (ACR16) at behaviorally relevant doses in living rats. Rats were administered 3 or 15 mg/kg pridopidine, or saline, before injection of the radiotracer C-11-SA4503 (sigma-1R) or C-11-raclopride (D2R). Some animals received 60 mg/kg pridopidine and were only scanned with C-11-raclopride. Cerebral C-11-SA4503 binding was quantified using metabolite-corrected plasma input data and distribution volume (V (T)) calculated by Logan graphical analysis. C-11-raclopride binding was quantified using striatum-to-cerebellum ratios and binding potentials calculated with a simplified reference tissue model. Cunningham-Lassen plots indicated sigma-1R occupancies of 57 +/- 2 and 85 +/- 2 % after pretreatment of animals with 3 and 15 mg/kg pridopidine. A significant (44-66 %) reduction of C-11-raclopride binding was only observed at 60 mg/kg pridopidine. At doses shown to elicit neurochemical and behavioral effects, pridopidine occupied a large fraction of sigma-1Rs and a negligible fraction of D(2)Rs. Significant D2R occupancy was only observed at a dose 20-fold higher than was required for sigma-1R occupancy. The characteristics of dopamine stabilizers may result from the combination of high sigma-1R and low D2R affinity