Penitrem A as a Tool for Understanding the Role of Large Conductance Ca2+/Voltage-Sensitive K+ Channels in Vascular Function
Creator
Asano S; Bratz I N; Berwick Z C; Fancher I S; Tune J D; Dick G M
Publisher
Journal of Pharmacology and Experimental Therapeutics
Date
2012
2012-08
Description
Large conductance, Ca2+/voltage-sensitive K+ channels (BK channels) are well characterized, but their physiological roles, often determined through pharmacological manipulation, are less clear. Iberiotoxin is considered the "gold standard" antagonist, but cost and membrane-impermeability limit its usefulness. Economical and membrane-permeable alternatives could facilitate the study of BK channels. Thus, we characterized the effect of penitrem A, a tremorigenic mycotoxin, on BK channels and demonstrate its utility for studying vascular function in vitro and in vivo. Whole-cell currents from human embryonic kidney 293 cells transfected with hSlo alpha or alpha + beta 1 were blocked >95% by penitrem A (IC50 6.4 versus 64.4 nM; p < 0.05). Furthermore, penitrem A inhibited BK channels in inside-out and cell-attached patches, whereas iberiotoxin could not. Inhibitory effects of penitrem A on whole-cell K+ currents were equivalent to iberiotoxin in canine coronary smooth muscle cells. As for specificity, penitrem A had no effect on native delayed rectifier K+ currents, cloned voltage-dependent Kv1.5 channels, or native ATP-dependent K-ATP current. Penitrem A enhanced the sensitivity to K+-induced contraction in canine coronary arteries by 23 +/- 5% (p < 0.05) and increased the blood pressure response to phenylephrine in anesthetized mice by 36 +/- 11% ( p < 0.05). Our data indicate that penitrem A is a useful tool for studying the role of BK channels in vascular function and is practical for cell and tissue (in vitro)
TRPA1 is functionally co-expressed with TRPV1 in cardiac muscle: Co-localization at z-discs, costameres and intercalated discs.
Creator
Andrei Spencer R; Sinharoy Pritam; Bratz Ian N; Damron Derek S
Publisher
Channels (Austin, Tex.)
Date
2016
2016-09
Description
Transient receptor potential channels of the ankyrin subtype-1 (TRPA1) and vanilloid subtype-1 (TRPV1) are structurally related, non-selective cation channels that show a high permeability to calcium. Previous studies indicate that TRP channels play a prominent role in the regulation of cardiovascular dynamics and homeostasis, but also contribute to the pathophysiology of many diseases and disorders within the cardiovascular system. However, no studies to date have identified the functional expression and/or intracellular localization of TRPA1 in primary adult mouse ventricular cardiomyocytes (CMs). Although TRPV1 has been implicated in the regulation of cardiac function, there is a paucity of information regarding functional expression and localization of TRPV1 in adult CMs. Our current studies demonstrate that TRPA1 and TRPV1 ion channels are co-expressed at the protein level in CMs and both channels are expressed throughout the endocardium, myocardium and epicardium. Moreover, immunocytochemical localization demonstrates that both channels predominantly colocalize at the Z-discs, costameres and intercalated discs. Furthermore, specific TRPA1 and TRPV1 agonists elicit dose-dependent, transient rises in intracellular free calcium concentration ([Ca(2+)]i) that are abolished in CMs obtained from TRPA1(-/-) and TRPV1(-/-) mice. Similarly, we observed a dose-dependent attenuation of the TRPA1 and TRPV1 agonist-induced increase in [Ca(2+)]i when WT CMs were pretreated with increasing concentrations of selective TRPA1 or TRPV1 channel antagonists. In summary, these findings demonstrate functional expression and the precise ultrastructural localization of TRPA1 and TRPV1 ion channels in freshly isolated mouse CMs. Crosstalk between TRPA1 and TRPV1 may be important in mediating cellular signaling events in cardiac muscle.