TRPA1 functions as an excitatory ionotropic receptor in sensory neurons. It was originally described as a noxious cold-activated channel, but its cold sensitivity has been disputed in later studies, and the contribution of TRPA1 to thermosensing is currently a matter of strong debate. Here, we provide several lines of evidence to establish that TRPA1 acts as a cold sensor in vitro and in vivo. First, we demonstrate that heterologously expressed TRPA1 is activated by cold in a Ca²⁺-independent and Ca²⁺store-independent manner; temperature-dependent gating of TRPA1 is mechanistically analogous to that of other temperature-sensitive TRP channels, and it is preserved after treatment with the TRPA1 agonist mustard oil. Second, we identify and characterize a specific subset of cold-sensitive trigeminal ganglion neurons that is absent in TRPA1-deficient mice. Finally, cold plate and tail-flick experiments reveal TRPA1-dependent, cold-induced nociceptive behavior in mice. We conclude that TRPA1 acts as a major sensor for noxious cold.

Reduced functional bladder capacity and concomitant increased micturition frequency (pollakisuria) are common lower urinary tract symptoms associated with conditions such as cystitis, prostatic hyperplasia, neurological disease, and overactive bladder syndrome. These symptoms can profoundly affect the quality of life of afflicted individuals, but available pharmacological treatments are often unsatisfactory. Recent work has demonstrated that the cation channel TRPV4 is highly expressed in urothelial cells and plays a role in sensing the normal filling state of the bladder. In this article, we show that the development of cystitis-induced bladder dysfunction is strongly impaired in Trpv4 ^−/− mice. Moreover, we describe HC-067047, a previously uncharacterized, potent, and selective TRPV4 antagonist that increases functional bladder capacity and reduces micturition frequency in WT mice and rats with cystitis. HC-067047 did not affect bladder function in Trpv4 ^−/− mice, demonstrating that its in vivo effects are on target. These results indicate that TRPV4 antagonists may provide a promising means of treating bladder dysfunction.

TRPM4 is a Ca²⁺-activated but Ca²⁺-impermeable cation channel. An increase of [Ca²⁺]_i induces activation and subsequent reduction of currents through TRPM4 channels. This inactivation is strikingly decreased in cell-free patches. In whole cell and cell-free configuration, currents through TRPM4 deactivate rapidly at negative potentials. At positive potentials, currents are much larger and activate slowly. This voltage-dependent behavior induces a striking outward rectification of the steady state currents. The instantaneous current-voltage relationship, derived from the amplitude of tail currents following a prepulse to positive potentials, is linear. Currents show a Boltzmann type of activation; the fraction of open channels increases at positive potentials and is low at negative potentials. Voltage dependence is not due to block by divalent cations or to voltage-dependent binding of intracellular Ca²⁺ to an activator site, indicating that TRPM4 is a transient receptor potential channel with an intrinsic voltage-sensing mechanism. Voltage dependence of TRPM4 may be functionally important, especially in excitable tissues generating plateau-like or bursting action potentials.

Transient receptor potential (TRP) channel, melastatin subfamily (TRPM)4 is a Ca2+‐activated monovalent cation channel that depolarizes the plasma membrane and thereby modulates Ca2+ influx through Ca2+‐permeable pathways. A typical feature of TRPM4 is its rapid desensitization to intracellular Ca2+ ([Ca2+]i). Here we show that phosphatidylinositol 4,5‐biphosphate (PIP2) counteracts desensitization to [Ca2+]i in inside‐out patches and rundown of TRPM4 currents in whole‐cell patch‐clamp experiments. PIP2 shifted the voltage dependence of TRPM4 activation towards negative potentials and increased the channel's Ca2+ sensitivity 100‐fold. Conversely, activation of the phospholipase C (PLC)‐coupled M1 muscarinic receptor or pharmacological depletion of cellular PIP2 potently inhibited currents through TRPM4. Neutralization of basic residues in a C‐terminal pleckstrin homology (PH) domain accelerated TRPM4 current desensitization and strongly attenuated the effect of PIP2, whereas mutations to the C‐terminal TRP box and TRP domain had no effect on the PIP2 sensitivity. Our data demonstrate that PIP2 is a strong positive modulator of TRPM4, and implicate the C‐terminal PH domain in PIP2 action. PLC‐mediated PIP2 breakdown may constitute a physiologically important brake on TRPM4 activity.

The recently cloned epithelial Ca²⁺ channel (ECaC) constitutes the Ca²⁺influx pathway in 1,25-dihydroxyvitamin D₃-responsive epithelia. We have combined patch-clamp analysis and fura-2 fluorescence microscopy to functionally characterize ECaC heterologously expressed in HEK293 cells. The intracellular Ca²⁺ concentration in ECaC-expressing cells was closely correlated with the applied electrochemical Ca²⁺ gradient, demonstrating the distinctive Ca²⁺ permeability and constitutive activation of ECaC. Cells dialyzed with 10 mm1,2-bis(2-aminophenoxy)ethane-N,N,N′,N′-tetraacetic acid displayed large inward currents through ECaC in response to voltage ramps. The corresponding current-voltage relationship showed pronounced inward rectification. Currents evoked by voltage steps to potentials below −40 mV partially inactivated with a biexponential time course. This inactivation was less pronounced if Ba²⁺or Sr²⁺ replaced Ca²⁺ and was absent in Ca²⁺-free solutions. ECaC showed an anomalous mole fraction behavior. The permeability ratioP _Ca:P _Na calculated from the reversal potential at 30 mm[Ca²⁺]_o was larger than 100. The divalent cation selectivity profile is Ca²⁺ > Mn²⁺ > Ba²⁺ ∼ Sr²⁺. Repetitive stimulation of ECaC-expressing cells induced a decay of the current response, which was greatly reduced if Ca²⁺ was replaced by Ba²⁺ and was virtually abolished if [Ca²⁺]_o was lowered to 1 nm. In conclusion, ECaC is a Ca²⁺ selective channel, exhibiting Ca²⁺-dependent autoregulatory mechanisms, including fast inactivation and slow down-regulation.