Deuteration as a tool in investigating the role of protons in cell signaling

• Published in: Biochimica et biophysica acta 2011-02, Vol.1810 (2), p.218-225
• Main Authors: Hirakura, Yutaka, Sugiyama, Takashi, Takeda, Masahiro, Ikeda, Masayuki, Yoshioka, Tohru
• Format: Article
• Language: English
• Subjects: Animals; Calcium - metabolism; Calcium Signaling - drug effects; Calorimetry; CHO Cells; Cricetinae; Cricetulus; Deuterium Exchange Measurement; Deuterium Oxide - chemistry; Deuterium Oxide - pharmacology; Estrenes - pharmacology; Inositol 1,4,5-Trisphosphate - metabolism; Methoxyhydroxyphenylglycol - analogs & derivatives; Methoxyhydroxyphenylglycol - pharmacology; Microscopy, Fluorescence; Phosphodiesterase Inhibitors - pharmacology; Protons; Pyrrolidinones - pharmacology; Receptors, Metabotropic Glutamate - genetics; Receptors, Metabotropic Glutamate - metabolism; Spectrometry, Fluorescence; Thermodynamics
• ISSN: 0006-3002; 0304-4165
• DOI: 10.1016/j.bbagen.2010.10.005

The mechanisms underlying the inhibitory effects of deuterium oxide (D₂O; heavy water) are likely to provide insight into the fundamental significance of hydrogen bonds in biological functions. Previously, to begin elucidating the effect of D₂O on physiological functions in living cells, we studied the effects of D₂O on voltage-sensitive Ca²(+) channels in AtT 20 cells and showed that actin distribution, Ca²(+) currents, and β-endorphin release were affected. However, the molecular mechanisms underlying the inhibitory effects of D₂O in whole animals and living cells remain obscure, especially in the effects of D₂O on the cell signaling. We investigated the molecular mechanisms underlying the inhibitory effects of D₂O on the IP₃-mediated Ca²(+) signaling pathway using Ca²(+) imaging and micro-calorimetric measurements in mGluR1-expressing CHO cells. DHPG-induced Ca²(+) elevations were markedly reduced in D₂O. Moreover, the Ca²(+) elevations were completely suppressed in H₂O after receptor activation with DHPG in D₂O, recovering gradually in H₂O medium. Without prior stimulation in D₂O, however, DHPG-induced Ca²(+) elevations in H₂O were not affected. Micro-calorimetric measurements showed reduced total DHPG-evoked heat generation in D₂O, while initial heat production and absorption associated with receptor activation were found to be larger. The reduction of DHPG-induced Ca²(+) elevation and heat generation in D₂O medium may be due to decreased amount of IP₃ by the reduced hydrolysis of PIP₂. Protein structure changes due to the replacement of hydrogen with deuterium will induce the inhibitory effects of D₂O by reduction of the frequency of -OH bonds.