On the Feasibility of Using an Acedan-Based Fluorescent Probe to Monitor Hydrogen Sulfide in Primary Neuronal Cultures

• Published in: Biochemistry (Moscow). Supplement series A, Membrane and cell biology Vol. 18; no. 2; pp. 90 - 99
• Main Authors: Sharipov, R. R., Tarzhanov, I. A., Zgodova, A. A., Bakaeva, Z. V., Surin, A. M.
• Format: Journal Article
• Language: English
• Published: Moscow Pleiades Publishing 01-06-2024; Springer Nature B.V
• Subjects: Biomedical and Life Sciences; Calcium homeostasis; Calcium signalling; Carbon monoxide; Cell Biology; Cell culture; Cerebral cortex; Composition; Cytosol; Deregulation; Dimethyl sulfoxide; Fluorescence; Fluorescent indicators; Glycine; Homeostasis; Hydrogen sulfide; Hydrophobicity; Intracellular; Life Sciences; Magnesium; Nitric oxide; Sodium bicarbonate; Sodium sulfide; fluorescent probe; hydrogen sulfide; excitotoxicity; primary neuronal cultures; glutamate
• ISSN: 1990-7478; 1990-7494
• DOI: 10.1134/S1990747824700119

Hydrogen sulfide (H
2
S), which under physiological conditions exists in cells mainly in the form of the HS
–
anion, is considered as a gaseous transmitter of inter- and intracellular signals along with nitrogen monoxide and carbon monoxide. Analysis of the dynamics of H
2
S content in living cells is impossible without the creation of sensitive and specific probes. The group of K.H. Ahn synthesized several acedan-based compounds, which in the presence of H
2
S attached a sulfhydryl group, forming fluorescent carbocyclic compounds. According to the spectral characteristics and reaction rate with H
2
S, the optimal substance was P3, which forms the carbocyclic compound csP3 with the same large Stokes shift as P3 (approx. 130 nm) and has a brighter fluorescence. In this work, we tested the suitability of csP3 for recording changes in H
2
S in solutions simulating the minimum salt composition of the intracellular medium, as well as in cells of primary neuronal culture from the rat cerebral cortex. It was found that the fluorescence intensity of csP3, which was formed when Na
2
S (H
2
S donor, 100 and 300 µM) was added to the P3 solution, differed for solutions corresponding in salt composition to the extracellular medium and cytosol. In both cases, fluorescence increased in the presence of bicarbonate (NaHCO
3
, 10 mM). A decrease in the polarity of solutions due to the addition of dimethyl sulfoxide (30% by volume) shifted the emission by approx. 10 nm to the shorter wavelength region and doubled the intensity. Glutamate (10 µM, in the presence of 10 µM of glycine, 0 Mg
2+
) increased the fluorescence of the probe, but only in those neurons in which delayed deregulation of calcium homeostasis did not occur. The addition of P3 or csP3 to the cell culture caused a rapid increase in the fluorescent signal, which was replaced by a slow signal growth after 3–5 min. It was concluded that the product of the reaction of P3 with H
2
S was sensitive to changes in the salt composition of the intracellular medium and could be redistributed in cells between an aqueous and more hydrophobic environment. These circumstances made it difficult to interpret the growth of P3 fluorescence in cells as a quantitative indicator of the presence of H
2
S and required additional studies of the properties of this and structurally related H
2
S probes.