Targeting repulsive guidance molecule A to promote regeneration and neuroprotection in multiple sclerosis.

Repulsive guidance molecule A (RGMa) is a potent inhibitor of neuronal regeneration and a regulator of cell death, and it plays a role in multiple sclerosis (MS). In autopsy material from progressive MS patients, RGMa was found in active and chronic lesions, as well as in normal-appearing gray and white matter, and was expressed by cellular meningeal infiltrates. Levels of soluble RGMa in the cerebrospinal fluid were decreased in progressive MS patients successfully treated with intrathecal corticosteroid triamcinolone acetonide (TCA), showing functional improvements.

Establishment of a secondary screening assay for P/Q-type calcium channel blockers.

Development of calcium channel blockers is attractive, but has in the past been hampered by lack of high throughput electrophysiological technology. This limitation has been overcome by the implementation of automated patch clamp systems that allow identification of state-dependent compounds, which preferentially target pathologically overactive channels. We recently presented a fluorescence-based high-throughput screen for P/Q-type calcium channels followed by automated electrophysiology.

Development of disease-modifying treatment of schizophrenia.

Development of disease-modifying therapies requires an innovative approach to drug development where novel drugs are designed to target mechanisms of interest rather than to produce preclinical effects similar to those of currently used antipsychotics. Application of such novel strategy will undoubtedly require a very deep understanding of the disease biology that is just starting to emerge. Alternatively, one may let environmental experiences of the diseased individual guide the repair process and use drugs only to facilitate the effects of experience.

Development and validation of a fluorescence-based HTS assay for the identification of P/Q-type calcium channel blockers.

Dysfunction of P/Q-type calcium channels is thought to underlie a variety of neurological diseases. There is evidence that migraine, Alzheimer's disease, and epilepsy involve a gain-of-function of the channel, leading to abnormal presynaptic vesicle release. P/Q-channel blockers may normalize current flow and consequently lead to an alleviation of disease symptoms.

Effects of a positive allosteric modulator of group II metabotropic glutamate receptors, LY487379, on cognitive flexibility and impulsive-like responding in rats.

Orthosteric group II metabotropic glutamate receptor (mGluR) agonists are regarded as novel, effective medications for all major symptom domains of schizophrenia, including cognitive disturbances. mGluR2s also can be affected in a more subtle way by positive allosteric modulators (PAMs) characterized by a unique degree of subtype selectivity and neuronal frequency-dependent activity. Because currently available treatments for schizophrenia do not improve cognitive dysfunction, the main aim of the present study was to examine the effects of a mGluR2 PAM, N-(4-(2-methoxyphenoxy)-phenyl-N-(2,2,2-trifluoroethylsulfonyl)-pyrid-3-ylmethylamine (LY487379), on rat cognitive flexibility and impulsive-like responding, assessed in an attentional set-shifting task (ASST) and a differential reinforcement of low-rate 72 s (DRL72) schedule of food reinforcement.

Generation and therapeutic efficacy of highly oligomer-specific beta-amyloid antibodies.

Oligomers of the beta-amyloid (Abeta) peptide have been indicated in early neuropathologic changes in Alzheimer's disease. Here, we present a synthetic Abeta(20-42) oligomer (named globulomer) with a different conformation to monomeric and fibrillar Abeta peptide, enabling the generation of highly Abeta oligomer-specific monoclonal antibodies. The globulomer-derived antibodies specifically detect oligomeric but not monomeric or fibrillar Abeta in various Abeta preparations.

Calpain inhibition prevents amyloid-beta-induced neurodegeneration and associated behavioral dysfunction in rats.

Amyloid-beta (Abeta) is toxic to neurons and such toxicity is - at least in part - mediated via the NMDA receptor. Calpain, a calcium dependent cystein protease, is part of the NMDA receptor-induced neurodegeneration pathway, and we previously reported that inhibition of calpain prevents excitotoxic lesions of the cholinergic nucleus basalis magnocellularis of Meynert. The present study reveals that inhibition of calpain is also neuroprotective in an in vivo model of Abeta oligomer-induced neurodegeneration in rats.

Glutamatergic (N-methyl-D-aspartate receptor) hypofrontality in schizophrenia: too little juice or a miswired brain?

Dopamine D2 receptor blockade has been an obligate mechanism of action present in all medications that effectively treat positive symptoms of schizophrenia (e.g., delusions and hallucinations) and have been approved by regulatory agencies since the 1950s.

Amphetamine decreases behavioral inhibition by stimulation of dopamine D2, but not D3, receptors.

Behavioral disinhibition is a manifestation of impulsive behavior that is prominent in the psychopathology of various psychiatric disorders such as addiction, attention-deficit hyperactivity disorder, mania, and personality disorders. Impulsivity may be studied by measuring anticipatory responses made before the presentation of a food-predictive, brief light stimulus in a two-choice serial reaction time task. In such serial reaction time tasks, amphetamine has been shown to produce dose-dependent increases in premature responding in a manner dependent on dopamine D(2)-like receptor stimulation.

Improvement of an acid phosphatase/DHAP-dependent aldolase cascade reaction by using directed evolution.

To enhance the phosphorylating activity of the bacterial nonspecific acid phosphatase from Salmonella enterica ser. typhimurium LT2 towards dihydroxyacetone (DHA), a mutant library was generated from the native enzyme. Three different variants that showed enhanced activity were identified after one round of epPCR.

Complementary chemoenzymatic routes to both enantiomers of febrifugine.

Two complementary strategies for the synthesis of febrifugine are detailed based on previously developed chemoenzymatic approaches to the 3-hydroxypiperidine skeleton. The introduction of the quinazolone-containing side chain in both strategies was based on an N-acyliminium ion-mediated coupling reaction.

Stimulating neuroregeneration as a therapeutic drug approach for traumatic brain injury.

Traumatic brain injury, a silent epidemic of modern societies, is a largely neglected area in drug development and no drug is currently available for the treatment of patients suffering from brain trauma. Despite this grim situation, much progress has been made over the last two decades in closely related medical indications, such as spinal cord injury, giving rise to a more optimistic approach to drug development in brain trauma. Fundamental insights have been gained with animal models of central nervous system (CNS) trauma and spinal cord injury.

Running wheel activity is sensitive to acute treatment with selective inhibitors for either serotonin or norepinephrine reuptake.

Rationale: Most antidepressants (AD) directly or indirectly enhance the serotonergic tone in the CNS. Since the serotonin system is involved in both, the modulation of mood and motor behavior, it was reasoned that these drugs might also interfere with running wheel activity (RWA), a form of positively motivated motor behavior, which might be linked to pathological states like obsessive-compulsive disorder (OCD).

Objectives: We used RWA to characterize ADs from all major classes.

Inhibitors of GlyT1 affect glycine transport via discrete binding sites.

In the forebrain, synaptic glycine concentrations are regulated through the glycine transporter GlyT1. Because glycine is a coagonist of the N-methyl-D-aspartate (NMDA) receptor (NMDAR), which has been implicated in schizophrenia, inhibition of GlyT1 is thought to provide an option for the treatment of schizophrenia. In support of this hypothesis, GlyT1 inhibitors facilitate in vivo NMDAR function and demonstrate antipsychotic-like effects in animal models.

N,N-acetals as N-acyliminium ion precursors: synthesis and absolute stereochemistry of epiquinamide.

A stereoselective synthesis of (+)-epiquinamide is presented in combination with determination of the absolute configuration of the natural product. Key steps in the sequence involved chemoenzymatic formation of an enantiomerically pure cyanohydrin, reductive cyclization to the corresponding cyclic N,N-acetal, and subsequent conversion into a suitable N-acyliminium ion precursor to enable construction of the second ring.

Inhibition of Calpain Prevents N-Methyl-D-aspartate-Induced Degeneration of the Nucleus Basalis and Associated Behavioral Dysfunction.

N-Methyl-D-aspartate (NMDA) receptor-mediated excitotoxicity is thought to underlie a variety of neurological disorders, and inhibition of either the NMDA receptor itself, or molecules of the intracellular cascade, may attenuate neurodegeneration in these diseases. Calpain, a calcium-dependent cysteine protease, has been identified as part of such an NMDA receptor-induced excitotoxic signaling pathway. The present study addressed the question of whether inhibition of calpain can prevent neuronal cell death and associated behavioral deficits in a disease-relevant animal model, which is based on excitotoxic lesions of the cholinergic nucleus basalis magnocellularis of Meynert.

Behavioral characterization of the mGlu group II/III receptor antagonist, LY-341495, in animal models of anxiety and depression.

There is a growing body of evidence indicating that stimulation of metabotropic glutamate type II receptors (mGlu2/3) reduces anxiety in laboratory animals and humans. Surprisingly, it was reported that mGlu2/3 receptor antagonists have antidepressant- and anxiolytic-like activities in laboratory animal studies as well. The present study aimed to resolve this controversy by characterizing behavioral effects of a selective mGlu2/3 receptor antagonist, LY-341495, in a variety of animal models sensitive to clinically used anxiolytic and antidepressant agents.

Amyloid beta oligomers (A beta(1-42) globulomer) suppress spontaneous synaptic activity by inhibition of P/Q-type calcium currents.

Abnormal accumulation of soluble oligomers of amyloid beta (Abeta) is believed to cause malfunctioning of neurons in Alzheimer's disease. It has been shown that Abeta oligomers impair synaptic plasticity, thereby altering the ability of the neuron to store information. We examined the underlying cellular mechanism of Abeta oligomer-induced synaptic modifications by using a recently described stable oligomeric Abeta preparation called "Abeta(1-42) globulomer.

Biocatalytic reductions: from lab curiosity to "first choice".

Enzyme-catalyzed reductions have been studied for decades and have been introduced in more than 10 industrial processes for production of various chiral alcohols, alpha-hydroxy acids and alpha-amino acids. The earlier hurdle of expensive cofactors was taken by the development of highly efficient cofactor regeneration methods. In addition, the accessible number of suitable dehydrogenases and therefore the versatility of this technology is constantly increasing and currently expanding beyond asymmetric production of alcohols and amino acids.

Laser capture microdissection and microarray analysis of dividing neural progenitor cells from the adult rat hippocampus.

Neural progenitor cells reside in the hippocampus of adult rodents and humans and generate granule neurons throughout life. Knowledge about the molecular processes regulating these neurogenic cells is fragmentary. In order to identify genes with a role in the proliferation of adult neural progenitor cells, a protocol was elaborated to enable the staining and isolation of such cells under RNA-preserving conditions with a combination of immunohistochemistry and laser capture microdissection.

Habituation deficits induced by metabotropic glutamate receptors 2/3 receptor blockade in mice: reversal by antipsychotic drugs.

Cortical metabotropic glutamate receptors (mGluRs) seem to be involved in habituation of simple stimulus-bound behaviors (e.g., habituation to acoustic startle or odor-elicited orienting response).

Nitrile hydrolysis activity of Rhodococcus erythropolis NCIMB 11540 whole cells.

The nitrile hydrolyzing properties of the bacterium strain Rhodococcus erythropolis NCIMB 11540 have been investigated. Using whole cells of the microorganism, a wide variety of aromatic and aliphatic cyanide-containing substrates was successfully hydrolyzed to the corresponding amide or acid. In the case of dicyanides, selective monohydrolysis took place, which was further explored in the desymmetrization of malononitriles resulting in the corresponding cyano amides in enantiomeric excesses of up to 98%.

Simple enzymatic in situ generation of dihydroxyacetone phosphate and its use in a cascade reaction for the production of carbohydrates: increased efficiency by phosphate cycling.

A new enzymatic method for the generation of dihydroxyacetone phosphate (DHAP) using the acid phosphatase from Shigella flexneri (PhoN-Sf) and the cheap phosphate donor pyrophosphate (PPi) is described. The utility of this method was demonstrated in an aldolase-catalyzed condensation carried out in one pot in which DHAP was generated and coupled to propionaldehyde to give a yield of 53% of the isolated dephosphorylated end product.

An ELISA-based method for the quantification of incorporated BrdU as a measure of cell proliferation in vivo.

In this study, we describe a new rapid and versatile method to determine the BrdU content of DNA in brain tissues dissected from BrdU-treated rats. Different to already existing BrdU ELISAs the method is suitable for the assessment of BrdU incorporation in ex vivo experiments as it is based on the analysis of tissue extracts instead of immobilized cells. The method comprises the preparation of DNA extracts from dissected tissues, the immobilization of BrdU-containing DNA with an anti-BrdU antibody and quantification of the incorporated BrdU by a peroxidase-conjugated anti-BrdU antibody.