Numerical analysis of neuronal responses for photolysis of MNI and Rubi caged glutamate

Abstract

Photo-labile caged compound are biologically inert state, but absorption of flash light unleash the cleavage of chemical bond so that bioactive molecules come out and have influence on cellular dynamics in various ways. Such uncaging method with advanced optical technique is possible to manipulate the function of cell with high subcellular resolution. However, there have been no suitable quantification method of the amount of photolysis in situ. Fluorescence indicators have not been made for caged compounds with the exception of specific bioactive molecules such as peptide and Ca2+. In this paper, we investigated evoked neuronal responses for photolysis of MNI and Rubi-caged glutamate, and suggested a new formula quantifying the extent of the uncaging. For those, primary hippocampal neurons were cultured on Microelectrode-array (MEA) with microfluidic devices for recording extracellular signals. Evoked neuronal responses was monitored according to optical stimulation parameters including wavelength, intensity, illumination duration and concentration of each chemicals respectively. Our experimental results revealed that the number of spikes per second was dependent on illumination power, wavelength, exposure time and concentration. Also, the first neural response was involved in illuminated intensity of light regardless of chemical species of caged glutamate. Those result indicated that three optical factors and concentration, were important factors to determine the amount of released glutamates. Finally, we established a new formula quantifying the amount of released glutamate. Through an empirical assessment, neuronal responses could be elicited by numerically modeling the amount of the released caged glutamates. We hoped that this formula was applied in quantification of the amount of photolysis of various caged compounds.