TRPA1 is a calcium-permeable nonselective cation transient receptor potential (TRP) channel that functions as an excitatory ionotropic receptor in nociceptive neurons. TRPA1 is robustly activated by ...pungent substances in mustard oil, cinnamon, and garlic and mediates the inflammatory actions of environmental irritants and proalgesic agents. Here, we demonstrate a bimodal sensitivity of TRPA1 to menthol, a widely used cooling agent and known activator of the related cold receptor TRPM8. In whole-cell and single-channel recordings of heterologously expressed TRPA1, submicromolar to low-micromolar concentrations of menthol cause channel activation, whereas higher concentrations lead to a reversible channel block. In addition, we provide evidence for TRPA1-mediated menthol responses in mustard oil-sensitive trigeminal ganglion neurons. Our data indicate that TRPA1 is a highly sensitive menthol receptor that very likely contributes to the diverse psychophysical sensations after topical application of menthol to the skin or mucous membranes of the oral and nasal cavities.
Steviol glycosides (SGs), such as stevioside and rebaudioside A, are natural, non-caloric sweet-tasting organic molecules, present in extracts of the scrub plant Stevia rebaudiana, which are widely ...used as sweeteners in consumer foods and beverages. TRPM5 is a Ca
-activated cation channel expressed in type II taste receptor cells and pancreatic β-cells. Here we show that stevioside, rebaudioside A and their aglycon steviol potentiate the activity of TRPM5. We find that SGs potentiate perception of bitter, sweet and umami taste, and enhance glucose-induced insulin secretion in a Trpm5-dependent manner. Daily consumption of stevioside prevents development of high-fat-diet-induced diabetic hyperglycaemia in wild-type mice, but not in Trpm5
mice. These results elucidate a molecular mechanism of action of SGs and identify TRPM5 as a potential target to prevent and treat type 2 diabetes.
The Transient Receptor Potential Ankyrin 1 cation channel (TRPA1) is a broadly-tuned chemosensor expressed in nociceptive neurons. Multiple TRPA1 agonists are chemically unrelated non-electrophilic ...compounds, for which the mechanisms of channel activation remain unknown. Here, we assess the hypothesis that such chemicals activate TRPA1 by inducing mechanical perturbations in the plasma membrane. We characterized the activation of mouse TRPA1 by non-electrophilic alkylphenols (APs) of different carbon chain lengths in the para position of the aromatic ring. Having discarded oxidative stress and the action of electrophilic mediators as activation mechanisms, we determined whether APs induce mechanical perturbations in the plasma membrane using dyes whose fluorescence properties change upon alteration of the lipid environment. APs activated TRPA1, with potency increasing with their lipophilicity. APs increased the generalized polarization of Laurdan fluorescence and the anisotropy of the fluorescence of 1,6-diphenyl-1,3,5-hexatriene (DPH), also according to their lipophilicity. Thus, the potency of APs for TRPA1 activation is an increasing function of their ability to induce lipid order and membrane rigidity. These results support the hypothesis that TRPA1 senses non-electrophilic compounds by detecting the mechanical alterations they produce in the plasma membrane. This may explain how structurally unrelated non-reactive compounds induce TRPA1 activation and support the role of TRPA1 as an unspecific sensor of potentially noxious compounds.
Here we provide evidence for a critical role of the transient receptor potential cation channel, subfamily V, member 4 (TRPV4) in normal bladder function. Immunofluorescence demonstrated TRPV4 ...expression in mouse and rat urothelium and vascular endothelium, but not in other cell types of the bladder. Intracellular Ca2+ measurements on urothelial cells isolated from mice revealed a TRPV4-dependent response to the selective TRPV4 agonist 4alpha-phorbol 12,13-didecanoate and to hypotonic cell swelling. Behavioral studies demonstrated that TRPV4-/- mice manifest an incontinent phenotype but show normal exploratory activity and anxiety-related behavior. Cystometric experiments revealed that TRPV4-/- mice exhibit a lower frequency of voiding contractions as well as a higher frequency of nonvoiding contractions. Additionally, the amplitude of the spontaneous contractions in explanted bladder strips from TRPV4-/- mice was significantly reduced. Finally, a decreased intravesical stretch-evoked ATP release was found in isolated whole bladders from TRPV4-/- mice. These data demonstrate a previously unrecognized role for TRPV4 in voiding behavior, raising the possibility that TRPV4 plays a critical role in urothelium-mediated transduction of intravesical mechanical pressure.
Allyl isothiocyanate (AITC; aka, mustard oil) is a powerful irritant produced by Brassica plants as a defensive trait against herbivores and confers pungency to mustard and wasabi. AITC is widely ...used experimentally as an inducer of acute pain and neurogenic inflammation, which are largely mediated by the activation of nociceptive cation channels transient receptor potential ankyrin 1 and transient receptor potential vanilloid 1 (TRPV1). Although it is generally accepted that electrophilic agents activate these channels through covalent modification of cytosolic cysteine residues, the mechanism underlying TRPV1 activation by AITC remains unknown. Here we show that, surprisingly, AITC-induced activation of TRPV1 does not require interaction with cysteine residues, but is largely dependent on S513, a residue that is involved in capsaicin binding. Furthermore, AITC acts in a membrane-delimited manner and induces a shift of the voltage dependence of activation toward negative voltages, which is reminiscent of capsaicin effects. These data indicate that AITC acts through reversible interactions with the capsaicin binding site. In addition, we show that TRPV1 is a locus for cross-sensitization between AITC and acidosis in nociceptive neurons. Furthermore, we show that residue F660, which is known to determine the stimulation by low pH in human TRPV1, is also essential for the cross-sensitization of the effects of AITC and low pH. Taken together, these findings demonstrate that not all reactive electrophiles stimulate TRPV1 via cysteine modification and help understanding the molecular bases underlying the surprisingly large role of this channel as mediator of the algesic properties of AITC.
Glucose homeostasis is critically dependent on insulin release from pancreatic β-cells, which is strictly regulated by glucose-induced oscillations in membrane potential (Vm) and the cytosolic ...calcium level (Ca²⁺cyt). We propose that TRPM5, a Ca²⁺-activated monovalent cation channel, is a positive regulator of glucose-induced insulin release. Immunofluorescence revealed expression of TRPM5 in pancreatic islets. A Ca²⁺-activated nonselective cation current with TRPM5-like properties is significantly reduced in Trpm5⁻/⁻ cells. Ca²⁺-imaging and electrophysiological analysis show that glucose-induced oscillations of Vm and Ca²⁺cyt have on average a reduced frequency in Trpm5⁻/⁻ islets, specifically due to a lack of fast oscillations. As a consequence, glucose-induced insulin release from Trpm5⁻/⁻ pancreatic islets is significantly reduced, resulting in an impaired glucose tolerance in Trpm5⁻/⁻ mice.
The cation channel TRPM8 plays a central role in the somatosensory system, as a key sensor of innocuously cold temperatures and cooling agents. Although increased functional expression of TRPM8 has ...been implicated in various forms of pathological cold hypersensitivity, little is known about the cellular and molecular mechanisms that determine TRPM8 abundance at the plasma membrane. Here we demonstrate constitutive transport of TRPM8 towards the plasma membrane in atypical, non-acidic transport vesicles that contain lysosomal-associated membrane protein 1 (LAMP1), and provide evidence that vesicle-associated membrane protein 7 (VAMP7) mediates fusion of these vesicles with the plasma membrane. In line herewith, VAMP7-deficient mice exhibit reduced functional expression of TRPM8 in sensory neurons and concomitant deficits in cold avoidance and icilin-induced cold hypersensitivity. Our results uncover a cellular pathway that controls functional plasma membrane incorporation of a temperature-sensitive TRP channel, and thus regulates thermosensitivity in vivo.
Transient Receptor Potential (TRP) channels form a broadly expressed and functionally diverse family of cation channels involved in various (patho)physiological processes. Whereas the mechanisms that ...control opening of TRP channels have been extensively studied, little is known about the transport processes of TRP channels to and within the plasma membrane. Here we used Total Internal Reflection--Fluorescence Recovery after Photobleaching (TIR-FRAP) to selectively visualize and bleach the fluorescently labeled TRP channels TRPV2 and TRPM4 in close proximity of the glass-plasma membrane interface, allowing detailed analysis of their perimembrane dynamics. We show that recovery of TRPM4 occurs via 200-nm diameter transport vesicles, and demonstrate the full fusion of such vesicles with the plasma membrane. In contrast, TRPV2 recovery proceeded mainly via lateral diffusion from non-bleached areas of the plasma membrane. Analysis of the two-dimensional channel diffusion kinetics yielded 2D diffusion coefficients ranging between 0.1 and 0.3 μm(2)/s, suggesting that these TRP channels move relatively unrestricted within the plasma membrane. These data demonstrate distinct modes of TRP channel turnover at the plasma membrane and illustrate the usefulness of TIR-FRAP to monitor these processes with high resolution.
We recently proposed that the transient receptor potential melastatin 5 (TRPM5) cation channel contributes to glucose-induced electrical activity of the β cell and positively influences ...glucose-induced insulin release and glucose homeostasis. In this study, we investigated
Trpm5
expression and function in pancreatic islets from mouse models of type II diabetes. Gene expression analysis revealed a strong reduction of
Trpm5
mRNA levels in pancreatic islets of
db
/
db
and
ob
/
ob
mice. The glucose-induced Ca
2+
oscillation pattern in
db
/
db
and
ob
/
ob
islets mimicked those of
Trpm5
−/−
islets. Leptin treatment of
ob
/
ob
mice not only reversed the diabetic phenotype seen in these mice but also upregulated
Trpm5
expression. Leptin treatment had no additional effect on
Trpm5
expression levels when plasma insulin levels were comparable to those of the vehicle-injected control group. In murine β cell line, MIN6, insulin downregulated TRPM5 expression in a dose-dependent manner, unlike glucose or leptin. In conclusion, our data show that increased plasma insulin levels downregulate TRPM5 expression in pancreatic islets from leptin-deficient mouse models of type 2 diabetes.
Acute pain represents a crucial alarm signal to protect us from injury. Whereas the nociceptive neurons that convey pain signals were described more than a century ago, the molecular sensors that ...detect noxious thermal or mechanical insults have yet to be fully identified. Here we show that acute noxious heat sensing in mice depends on a triad of transient receptor potential (TRP) ion channels: TRPM3, TRPV1, and TRPA1. We found that robust somatosensory heat responsiveness at the cellular and behavioural levels is observed only if at least one of these TRP channels is functional. However, combined genetic or pharmacological elimination of all three channels largely and selectively prevents heat responses in both isolated sensory neurons and rapidly firing C and Aδ sensory nerve fibres that innervate the skin. Strikingly, Trpv1
Trpm3
Trpa1
triple knockout (TKO) mice lack the acute withdrawal response to noxious heat that is necessary to avoid burn injury, while showing normal nociceptive responses to cold or mechanical stimuli and a preserved preference for moderate temperatures. These findings indicate that the initiation of the acute heat-evoked pain response in sensory nerve endings relies on three functionally redundant TRP channels, representing a fault-tolerant mechanism to avoid burn injury.
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