Agonist-induced Ca2+ entry into cells by both store-operated channels and channels activated independently of Ca2+-store depletion has been described in various cell types. The molecular structures of these channels are unknown as is, in most cases, their impact on various cellular functions. Here we describe a store-operated Ca2+ current in vascular endothelium and show that endothelial cells of mice deficient in TRP4 (also known as CCE1) lack this current. As a consequence, agonist-induced Ca2+ entry and vasorelaxation is reduced markedly, showing that TRP4 is an indispensable component of store-operated channels in native endothelial cells and that these channels directly provide an Ca2+-entry pathway essentially contributing to the regulation of blood vessel tone.

SUMMARY In the mammalian central nervous system, slow synaptic excitation involves the activation of metabotropic glutamate receptors (mGluRs). It has been proposed that C1-type transient receptor potential (TRPC1) channels underlie this synaptic excitation, but our analysis of TRPC1-deficient mice does not support this hypothesis. Here, we show unambiguously that it is TRPC3 that is needed for mGluR-dependent synaptic signaling in mouse cerebellar Purkinje cells. TRPC3 is the most abundantly expressed TRPC subunit in Purkinje cells. In mutant mice lacking TRPC3, both slow synaptic potentials and mGluR-mediated inward currents are completely absent, while the synaptically mediated Ca2+ release signals from intracellular stores are unchanged. Importantly, TRPC3 knockout mice exhibit an impaired walking behavior. Taken together, our results establish TRPC3 as a new type of postsynaptic channel that mediates mGluR-dependent synaptic transmission in cerebellar Purkinje cells and is crucial for motor coordination.

TRPM4 is a Ca 2؉ -activated but Ca 2؉ -impermeable cation channel. An increase of [Ca 2؉ ] 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. Cation channels of the transient receptor potential (TRP) 1 superfamily have received much attention during the last years because of their pivotal role in various cell functions linked to the modulation of intracellular Ca 2ϩ signals, mostly in nonexcitable cells (1-3). More than 20 mammalian TRP members are known, which are classified into the TRPC (C for "canonical"), TRPV (V for "vanilloid"), and TRPM (M for "melastatin") subfamilies (1, 2). However, the functional properties of most members of this novel and fast growing channel family are not yet analyzed in detail. The predicted transmembrane topology of TRPs is similar to that of voltage-gated and cyclic nucleotide gated channels; they consist of six transmembrane-spanning helices (TM1 to -6), cytoplasmic N and C termini, and a pore region between TM5 and TM6 (2, 3). Since the fourth transmembrane helix is not positively charged, TRP channels were considered as voltage-independent. The voltage-sensing features of some members of the TRPV subfamily could be attributed to voltage-dependent block of the channel pore by intra-or extracellular divalent cations (4, 5).Members of the TRPM subfamily are much less studied at the functional level than those of the TRPV and TRPC family. They are characterized by relatively long N and C termini, and some of them have entire enzyme domains linked to their C termini: an ADP-ribose pyrophosphatase in TRPM2 (6) and an atypical ␣-kinase domain in TRPM6 and TRPM7 (7-11). Surprisingly, TRPM4b, which is a Ca 2ϩ -impermeable monovalent cation channel of 25-picosiemens unitary conductance belonging to the TRPM subfamily, is in contrast to other TRP channels not inactivated but activated by intracellular Ca 2ϩ , [Ca 2ϩ ] i (12). A short form of TRPM4, TRPM4a, is characterized in less detail and displays completely different properties with regard to Ca 2ϩ permeability and activation (13). In this study, we report cloning of the human and mouse TRPM4 cDNAs. Transcripts of these genes are expressed in heart, kidney, and endothelial cells, indicating that this channel plays a role in the cardiovascular system. We demonstrate that TRPM4 is a Ca 2ϩ -and voltage-dependent channel. It could therefore modulate the electrical activity of cells that generate action potentials. This is, to our knowledge, the first description of voltage-dependent properties of a TRP channel, suggesting a special role for this channel in excitable cells. MATERIALS AND METHODS Cloning of Human and Mouse TRPM4 cDNAs-The human expressed sequence tag 885075 (GenBank TM ), homologous to the human TRPM1 cDNA, was identified and sequenced on both strands; it contained a ϳ1500-bp DNA fragment, which represented part of the...

Abstract-We investigated the possibility that the TRPC gene family of putative store-operated Ca 2ϩ entry channels contributes to the increase in microvascular endothelial permeability by prolonging the rise in intracellular Ca 2ϩ signaling. Studies were made in wild-type (wt) and TRPC4 knockout (TRPC4 Ϫ/Ϫ ) mice and lung vascular endothelial cells (LECs) isolated from these animals. RT-PCR showed expression of TRPC1, TRPC3, TRPC4, and TRPC6 mRNA in wt LECs, but TRPC4 mRNA expression was not detected in TRPC4

Lower vertebrates have an intrinsically-photosensitive iris and thus a local pupillary light reflex (PLR). In contrast, it has been a dogma that the PLR in mammals generally requires neuronal circuitry connecting the eye and the brain. We report here that an intrinsic component of the PLR is actually widespread in nocturnal and crepuscular mammals. In mouse, this intrinsic PLR requires the visual pigment, melanopsin. It also requires PLCβ4, the vertebrate homolog of the Drosophila NorpA phospholipase C mediating rhabdomeric phototransduction. The Plcβ4−/− genotype, besides removing the intrinsic PLR, also essentially eliminates the intrinsic light response of the M1-subtype of melanopsin-expressing, intrinsically-photosensitive retinal ganglion cells (M1-ipRGCs), by far the most photosensitive ipRGCs and with the largest responses. Ablating in mouse the expression of both TRPC6 and TRPC7, members of the TRP channel superfamily, likewise essentially eliminated the M1-ipRGC light response, but spared the intrinsic PLR. Thus, melanopsin signaling exists in both iris and retina, involving a PLCβ4-mediated pathway that nonetheless diverges in the two locations.