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Streptozotocin Stimulates the Ion Channel TRPA1 Directly: INVOLVEMENT OF PEROXYNITRITE.

ABSTRACT: Streptozotocin (STZ)-induced diabetes is the most commonly used animal model of diabetes. Here, we have demonstrated that intraplantar injections of low dose STZ evoked acute polymodal hypersensitivities in mice. These hypersensitivities were inhibited by a TRPA1 antagonist and were absent in TRPA1-null mice. In wild type mice, systemic STZ treatment (180 mg/kg) evoked a loss of cold and mechanical sensitivity within an hour of injection, which lasted for at least 10 days. In contrast, Trpa1(-/-) mice developed mechanical, cold, and heat hypersensitivity 24 h after STZ. The TRPA1-dependent sensory loss produced by STZ occurs before the onset of diabetes and may thus not be readily distinguished from the similar sensory abnormalities produced by the ensuing diabetic neuropathy. In vitro, STZ activated TRPA1 in isolated sensory neurons, TRPA1 cell lines, and membrane patches. Mass spectrometry studies revealed that STZ oxidizes TRPA1 cysteines to disulfides and sulfenic acids. Furthermore, incubation of tyrosine with STZ resulted in formation of dityrosine, suggesting formation of peroxynitrite. Functional analysis of TRPA1 mutants showed that cysteine residues that were oxidized by STZ were important for TRPA1 responsiveness to STZ. Our results have identified oxidation of TRPA1 cysteine residues, most likely by peroxynitrite, as a novel mechanism of action of STZ. Direct stimulation of TRPA1 complicates the interpretation of results from STZ models of diabetic sensory neuropathy and strongly argues that more refined models of diabetic neuropathy should replace the use of STZ.

SUBMITTER: Andersson DA

PROVIDER: S-EPMC4463460 | biostudies-literature | 2015 Jun

REPOSITORIES: biostudies-literature

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Streptozotocin Stimulates the Ion Channel TRPA1 Directly: INVOLVEMENT OF PEROXYNITRITE.

Andersson David A DA Filipović Milos R MR Gentry Clive C Eberhardt Mirjam M Vastani Nisha N Leffler Andreas A Reeh Peter P Bevan Stuart S

The Journal of biological chemistry 20150422 24

Streptozotocin (STZ)-induced diabetes is the most commonly used animal model of diabetes. Here, we have demonstrated that intraplantar injections of low dose STZ evoked acute polymodal hypersensitivities in mice. These hypersensitivities were inhibited by a TRPA1 antagonist and were absent in TRPA1-null mice. In wild type mice, systemic STZ treatment (180 mg/kg) evoked a loss of cold and mechanical sensitivity within an hour of injection, which lasted for at least 10 days. In contrast, Trpa1(-/- ...[more]

PMID: 25903127

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Project description:RationaleTransient receptor potential channels of the ankyrin subtype-1 (TRPA1) are non-selective cation channels that show high permeability to calcium. Previous studies from our laboratory have demonstrated that TRPA1 ion channels are expressed in adult mouse ventricular cardiomyocytes (CMs) and are localized at the z-disk, costamere and intercalated disk. The functional significance of TRPA1 ion channels in the modulation of CM contractile function have not been explored.ObjectiveTo identify the extent to which TRPA1 ion channels are involved in modulating CM contractile function and elucidate the cellular mechanism of action.Methods and resultsFreshly isolated CMs were obtained from murine heart and loaded with Fura-2 AM. Simultaneous measurement of intracellular free Ca2+ concentration ([Ca2+]i) and contractility was performed in individual CMs paced at 0.3 Hz. Our findings demonstrate that TRPA1 stimulation with AITC results in a dose-dependent increase in peak [Ca2+]i and a concomitant increase in CM fractional shortening. Further analysis revealed a dose-dependent acceleration in time to peak [Ca2+]i and velocity of shortening as well as an acceleration in [Ca2+]i decay and velocity of relengthening. These effects of TRPA1 stimulation were not observed in CMs pre-treated with the TRPA1 antagonist, HC-030031 (10 µmol/L) nor in CMs obtained from TRPA1-/- mice. Moreover, we observed no significant increase in cAMP levels or PKA activity in response to TRPA1 stimulation and the PKA inhibitor peptide (PKI 14-22; 100 nmol/L) failed to have any effect on the TRPA1-mediated increase in CM contractile function. However, TRPA1 stimulation resulted in a rapid phosphorylation of Ca2+/calmodulin-dependent kinase II (CaMKII) (1-5 min) that correlated with increases in CM [Ca2+]i and contractile function. Finally, all aspects of TRPA1-dependent increases in CM [Ca2+]i, contractile function and CaMKII phosphorylation were virtually abolished by the CaMKII inhibitors, KN-93 (10 µmol/L) and autocamtide-2-related peptide (AIP; 20 µmol/L).ConclusionsThese novel findings demonstrate that stimulation of TRPA1 ion channels in CMs results in activation of a CaMKII-dependent signaling pathway resulting in modulation of intracellular Ca2+ availability and handling leading to increases in CM contractile function. Cardiac TRPA1 ion channels may represent a novel therapeutic target for increasing the inotropic and lusitropic state of the heart.

Dataset Information

Streptozotocin Stimulates the Ion Channel TRPA1 Directly: INVOLVEMENT OF PEROXYNITRITE.

Publications

Streptozotocin Stimulates the Ion Channel TRPA1 Directly: INVOLVEMENT OF PEROXYNITRITE.

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