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16
XXIII Congress of I.P. Pavlov Physiology Society
New Trends in the Analysis of Function and Modulation of the Cells of Nervous System
P. Bregestovski
Aix Marseille University, INSERM, Institute of System Neurosciences, Marseille, France; Department of Physiology, Kazan; Medical State University, Kazan, Russia.
Several new areas for analysis and functional regulation of nervous system function have been developed in recent years. Among them connectomics, clarity, optogenetics, optosensorics and optopharmacology provide an excellent tools for detailed investigation of the nervous system organization, on clarifying of neuronal networks functioning and control of some biological organisms behaviour.
A crucial factor for the establishment of optogenetics become the discovery of algal light-gated ion channels (chan-nelrhodopsins) and pumps (halorhodopsins), which can be easily incorporated in different cell types. This provided unprecedented tolls for the control of neural activity with light. Consequently, a number of other light-sensitive modules were discovered and engineered to generate new photoswitches to control protein activity, protein localization, and gene expression. With the help of light one can investigate the function of cells controlling their activity, to measure the concentration of ions, ATP and other cellular components, to control the behaviour of organisms, as well as to seek for novel ways to treat certain diseases. Optogenetic approaches have been used in many models with medical orientation, including the study of stress, schizophrenia, memory disorders, drug addiction, psychiatry and motor functions, vision, pain, functional recovery after stroke and epilepsy.
Photopharmacology is a direction based on the creation of chemical compounds capable of controlling the functions of biological molecules possessing photosensitive switches. Photochromic compounds that activate or inhibit the activity of key cellular proteins, especially ion channels, represent a powerful tools for non-invasive control of neuronal network activity and, consequently, functional control of organisms and behavior. Photochromic switches can be divided into two main classes: (i) soluble photochromic ligands and (ii) compounds covalently bounding to target proteins. On this basis, potassium channel blockers, modulators of glutamate and GABA receptors, as well as cationic TRP channels were created. Photopharmacology offers great opportunities in the regulation of pain, restoration of the functions of the retina and other physiological functions.
Optosensorics uses genetically encoded biosensors for non-invasive imaging of concentrations of ions, the activity of enzymes, distribution of small molecules, proteins and organelles, and the protein interactions in living cells. These fluorescent molecules are used either at the transient expression in cultured cells or organisms or at stable expression producing transgenic animals possessing heritable and functional biosensors. Using the mouse Thy1 mini-promoter, we generated two lines of transgenic mice for the monitoring of intracellular chloride (Cl-) and for the simultaneous measurements of intracellular and pH. To reveal the ClopHensor expression pattern across the brain of transgenic mice we obtained transparent brain samples using CLARITY method and imaged them with confocal and light-sheet microscopy. This analysis provides the map of the brain areas available for non-invasive monitoring of intracellular Cl'/pH in normal and pathological conditions.
These and other directions will be presented in the talk.
Spatiotemporal Characteristics of Astrocyte Calcium Dynamics in Chronic Epilepsy
P. Denisov*, A. Lebedeva, A. Pimashkin, A. Semyanov
Institute of Neuroscience, Lobachevsky State University of Nizhny Novgorod, Nizhny Novgorod, Russia. * Presenting e-mail: denisov@neuro.nnov.ru
Epilepsy is diagnosed annually in 2.4 million people (WHO 2017). Epileptic status (ES) is the main symptom and is the most severe manifestation of generalized epilepsy. It is a series of seizures with tonic and clonic phases, leading to neurodegeneration and astrogliosis. The influence of these processes on the neuron-astrocytic interaction, in particular on the calcium events in the astrocytic network, remains poorly studied. The study of astrocytic activity can give new knowledge about the mechanisms of epilepsy and determine the direction of creating new methods of treatment.
As an experimental model in this project, a lithium-pilocarpine model of epileptogenesis was chosen. It reflects the various phenomenological features characteristic of temporal epilepsy in humans.
Sprague-Dawley rats (age 18-25 days) were used in the experiments. 2-4 weeks after ES, the changes in calcium dynamics in the neuron-astrocytic network stratum radiatum of the CA1 field on the hippocampal slices were stud-
OM&P
4 Opera Med Physiol 2017 Vol. 3 (S1)
