Research field
The Department of Neurochemistry deals with physiology and pharmacology of cholinergic neurons. Cholinergic neurotransmission is involved in many central as well as peripheral functions. Among them, brain muscarinic cholinergic transmission takes part in cognitive functions. A serious disturbances of muscarinic transmission underlie characteristic features of neurodegeneration accompanying e.g. Parkinson´s and Alzheimer´s disease. The department have long-term experience with investigations on mechanisms of cholinergic transmission (ref. 1-11), including presynaptic functions like synthesis, storage, and release of ACh, regulation of ACh release, and with mechanisms of signal transduction and molecular biology of muscarinic receptors.
Current projects
Physiological operation of cholinergic signaling necessitates availability of signaling molecule acetylcholine properly distributed in time and space, and intact receptive machinery comprising receptor itself and its coupling pathway connecting receptor stimulation to functional outcome. We investigate mechanisms which govern metabolism of acetylcholine in cholinergic nerve endings and transduction of signal through muscarinic receptors. There are five subtypes of muscarinic receptors (M1-M5) encoded by five different genes. They share a high degree of homology in transmembrane domains while extracellular and intracellular loops are less conserved. Acetylcholine binding site that activates all receptor subtypes is located in a pocket located among conserved transmembrane domains. It makes difficult to find a subtype selective agonists that could be used for targetting muscarinic receptor subtype-specific signaling pathways. Allosteric modulators of muscarinic receptors offer a remarkable selectivity that depend both on the receptor subtype and the pair orthosteric ligand/allosteric ligand. We are investigating interactions of allosteric modulators and orthosteric ligands as well as of atopic agonists (activating receptor from a site different then is acetylcholine binding site) with the aim to determine molecular mechanisms involved in subtype specificity of allosteric and atopic ligands in influencing signal transduction across plasma membrane and their effects on receptor cycling (ref. 12-16, 18, and 20). In the context of cholinergic deregulation in Alzheimer´s disease we are investigating mechanisms of deleterious effects of amyloid-β on brain muscarinic transmission (ref. 16, 17, and 19).
Methods
Running techniques comprise kinetic and equilibrium radioligand binding studies, real-time PCR, scintillation proximity assay, radioenzymatic, fluorometric, and spectrophotometric measurement of enzyme activities and metabolites, microfluorimetric single cell measurement of intracellular calcium, western blotting, and basic molecular biology techniques. The Lab is equipped for work with radioisotopes, cell cultures, and genetically modified organisms.
Selected references
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