We have previously reported that the reducing agent dithiothreitol (DTT) strongly increases thermally induced activity of the transient receptor potential vanilloid receptor-1 (TRPV1) channel. Here, we show that exposure to oxidizing agents also enhances the heat-induced activation of TRPV1. The actions of sulfhydryl modifiers on heat-evoked whole-cell membrane currents were examined in TRPV1-transfected human embryonic kidney 293T cells. The sensitizing effects of the membrane-permeable oxidizing agents diamide (1 mM), chloramine-T (1 mM), and the copper-o-complex (100:400 microM) were not reversed by washout, consistent with the stable nature of covalently modified sulfhydryl groups. In contrast, the membrane-impermeable cysteine-specific oxidant 5,5'-dithio-bis-(2-nitrobenzoic acid) (0.5 mM) was ineffective. The alkylating agent N-ethylmaleimide (1 mM) strongly and irreversibly affected heat-evoked responses in a manner that depended on DTT pretreatment. Extracellular application of the membrane-impermeable reducing agent glutathione (10 mM) mimicked the effects of 10 mM DTT in potentiating the heat-induced and voltage-induced membrane currents. Using site-directed mutagenesis, we identified Cys621 as the residue responsible for the extracellular modulation of TRPV1 by reducing agents. These data suggest that the vanilloid receptor is targeted by redox-active substances that directly modulate channel activity at sites located extracellularly as well as within the cytoplasmic domains. The results obtained demonstrate that an optimal redox state is crucial for the proper functioning of the TRPV1 channel and both its reduced and oxidized states can result in an increase in responsiveness to thermal stimuli.

• MeSH
• buněčné linie MeSH
• diamid farmakologie   MeSH
• dithiothreitol farmakologie   MeSH
• ethylmaleimid farmakologie   MeSH
• financování organizované MeSH
• kapsaicin farmakologie   MeSH
• kationtové kanály TRPV fyziologie   genetika   MeSH
• krysa rodu rattus MeSH
• kyselina dithionitrobenzoová farmakologie   MeSH
• lidé MeSH
• membránové potenciály účinky léků   MeSH
• metoda terčíkového zámku MeSH
• missense mutace genetika   MeSH
• mutace genetika   MeSH
• mutantní proteiny fyziologie   genetika   MeSH
• oxidancia farmakologie   MeSH
• peroxid vodíku farmakologie   MeSH
• redukční činidla farmakologie   MeSH
• sulfhydrylová reagencia farmakologie   MeSH
• transfekce MeSH
• vysoká teplota MeSH
• vztah mezi dávkou a účinkem léčiva MeSH
• zvířata MeSH
• Check Tag
• krysa rodu rattus MeSH
• lidé MeSH
• zvířata MeSH

At a concentration of 0.5 to 3 mmol/l, ATP stimulates the activity of mitochondrial DNA polymerase of Neurospora crassa under the optimum reaction conditions; at higher concentrations, an inhibitory effect is observed. 4-Chloromercuribenzoate (1 mmol/L), a thiol inhibitor, decreases the enzyme activity two-fold, while N-ethylmalcimide (2 mmol/L) has no effect. Ethidium bromide (up to 10 mumol/L) and heparin (up to 0.4 micrograms/mL) reduce the activity by 60%. ddTTP does not affect the DNA polymerase reaction. The best in vitro template is the activated calf-thymus DNA.

• MeSH
• adenosintrifosfát farmakologie   MeSH
• aktivace enzymů účinky léků   MeSH
• dideoxynukleotidy MeSH
• DNA-dependentní DNA-polymerasy metabolismus   MeSH
• ethidium farmakologie   MeSH
• heparin farmakologie   MeSH
• inhibitory syntézy nukleových kyselin * MeSH
• mitochondrie enzymologie   MeSH
• Neurospora crassa účinky léků   enzymologie   MeSH
• sulfhydrylová reagencia farmakologie   MeSH
• thiminnukleotidy farmakologie   MeSH
• Publikační typ
• časopisecké články MeSH