Abstract
Implementation of fluorescence methods in studies of ligand binding to their receptors opens new possibilities to characterise these processes. One of the potential approaches is the detection of changes of the fluorescence anisotropy (FA) and/or total fluorescence intensity (TFI) signals upon binding reaction. However, to achieve significant changes in the FA/TFI signal, some requirements need to be met – the concentration of receptor binding sites as well as the dissociation constant of the interaction should be in the same order as the fluorescent ligand’s concentration. We have used FA assay to investigate ligand binding properties to Melanocortin 4 (MC4) receptor [1]. Implementation of budded baculoviruses (BBV) that display G protein-coupled receptors on their surfaces significantly increased sensitivity and temporal stability of this assay [2]. For the first time we demonstrate the applicability of BBV experimental setup to study Y1 receptor system. Here we used TAMRA-PYY, an Y1 receptor specific fluorescent peptide ligand, as a reporter ligand. Besides real-time monitoring of FA signal changes, up to 5 fold decrease in TFI signal was observed within TAMRA-PYY binding to the Y1 receptor. Pharmacological characterization of Y1 receptors with receptor-specific unlabelled ligands gave the rank order of potencies consistent with previously reported values. Additionally, allosteric heterogeneous interactions were revealed as koff values of TAMRA-PYY differed more than 7 times depending on the nature of dissociation initiated ligand. These observations provide evidence for similar allosteric receptor-ligand binding mechanism as previously shown for MC4 receptors [3].
Highlights
Alzheimer’s disease (AD) is an incurable neurodegenerative disease characterized by progressive dementia
The results of the present study indicate that development of the neuronal hypoxic tolerance induced by the three-trial, in contrast to one-trial, mild hypoxic preconditioning is apparently largely associated with the activation of CREB, as well as brain-derived neurotrophic factor (BDNF) and Bcl-2 overexpression
No significant differences in serum level of Solubile form of RAGE (sRAGE) where found between rapidly progressing and slow progressing subgroup of multiple sclerosis (MS) patients.Our results suggest for the role of sRAGE in MS ethiopathogenesis, but we did not find any association of sRAGE in serum with the rate of MS disability progression
Summary
Alzheimer’s disease (AD) is an incurable neurodegenerative disease characterized by progressive dementia. The aim of the study was to characterize the effects of streptozocin (STZ)-indced diabetes on learning and memory of 5XFAD and wild-type (WT) mice in Morris water maze (MWM) at ages 2 and 6 months and on brain amyloid load. Existing evidence suggests GABAergic system is involved in pathophysiology of Alzheimer’s disease (AD) via inhibitory interneuron deficits (Verret et al, 2012) and decrease in functional GABAA receptors (Limon et al, 2012). Our concept: low doses of muscimol may prevent learning/memory deficits in intracerebroventricular (icv) streptozocin (STZ)-induced AD nontransgenic rat model. The Sigma-1 receptor is a chaperone protein that modulates intracellular calcium signalling of the endoplasmatic reticulum and is involved in learning and memory processes.The aim of the present study was to compare in vitro Ca2+ concentration modulating activity and in vivo behavioural effects of enantiomers of methylphenylpiracetam, a novel positive allosteric modulator of Sigma-1 receptors
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