Send Orders of Reprints at [email protected] Current Hypertension Reviews, 2012, 8, 181-189 181 TRP Channels: Emerging Links Between Ca2+, and Hypertension

Paolo Menè*, Anna Giuliani, Jacopo Scrivano, Francesca Apponi, Giorgio Punzo and Nicola Pirozzi

Department of Clinical and Molecular Medicine, “Sapienza” University of Rome, School of Medicine and Psychology, Rome, Italy

Abstract: Cationic channels with variable selectivity for Ca2+, K+, and Na+ are ubiquitous in non-excitable tissues as well as contractile cardiac cells and vascular smooth muscle (VSMC). They are involved in multiple cell functions, including contraction and proliferation of VSMC, key to the regulation of vascular tone and blood pressure. A large superfamily includes at least 28 highly conserved heterotetramer homologues of Drosophila TRP (transient receptor potential) channels. Direct evidence exists for TRPC, TRPV, TRPP gating ion entry in vascular cells, as well as in epithelial cells of kidney and intestine, thus controlling homeostasis and external balance of divalent cations. Mice deficient in TRPV5 display phenotypic defects resembling human idiopathic hypercalciuria and impaired bone mineral density. , encoded by the PKD2 , is an epithelial transmembrane TRPP2 , whose mutation is associated with autosomal dominant polycystic kidney disease (ADPKD). TRP ion channels may therefore play a role in the pathogenesis of systemic arterial hypertension, pulmonary hypertension, as well as hypertension complicating renal disease via effects on vascular tone and/or on the electrolyte composition of body fluids. Keywords: Transient Receptor Potential Channels, Store-Operated Ca2+ entry, Pulmonary Hypertension, Arterial Hypertension.

INTRODUCTION as photoreceptors in insect retinal cells, and later recognized in a variety of sensing apparatuses [4-7]. These include Regulated permeability of plasma membranes to ions and visual and hearing structures, such as cochlea sensory hair water is key to preserving the internal microenvironment and cells, but are also widely distributed in the cilia of epithelial cell volume in living organisms, resisting to shifts of external cells of various organs [4-7]. Cation influx through these ion concentrations and osmotic/oncotic forces. This is channels often follows activation of a physical or chemical critical for both resting and activated cells, enabling sensor. Selectivity of TRP channels for one divalent cation thorough control of the transmembrane potential, which in such as Ca2+ or Mg2+ vs monovalent Na+ is variable from excitable cell drives electric currents, action potentials, and >100:1 to 1:1, based on isoform structure and tissue type contraction/relaxation. Permeability of the plasma membrane [4-7]. involves a host of channels, water and ion transporters constitutively expressed or translocated to the surface of TRP channels also serve as absorption mechanisms in cells in a highly regulated fashion. Channels or carrier epithelial cells, regulated by transmembrane concentration or not only control a cell’s microenvironment and electrochemical gradients. One such process is “bulk entry”, transmembrane potential, but do also serve for signalling, as in the case of Ca2+ or Mg2+ influx in the duodenum or inasmuch as ion fluxes encode for functional responses in renal tubules. In the kidney, TRPC5 and 6 channels mediate target cells. Vasoconstrictors or vasorelaxant agents, such as epithelial Ca2+ reabsorption in the , peptides or catecholamines, often act by redistributing Ca2+ and hence external Ca2+ balance [8, 9]. Genetically or Mg2+ in intracellular compartments upon activation of the engineered TRPC5-knockout mice exhibit hypercalciuria appropriate receptors [1, 2]. Further levels of signalling may and skeletal defects consistent with altered distal tubular be encoded through frequency and/or amplitude of reabsorption of Ca2+ [8, 9]. This model resembles human oscillatory currents, in turn actuated by cyclic ion gating [3]. idiopathic hypercalciuria. Another epithelial cell mechanism is more of a “fine tuning” nature, in that kidney tubular or Extensive literature has been amassed in the past 10-15 hepatic biliary cells express polycystin 2 (PKD-2), a TRP- years on a large superfamily of ion channels, the so-called type Ca2+ channel in the context of cilia, sensing organelles transient receptor potential (TRP) channels, fairly likely controlling urine or bile flows in the surrounding ubiquitous heterotetrameric polypeptides, initially identified environment [10, 11]. Defects in renal tubular PKD-2-

mediated Ca2+ transport, as commonly encountered in

autosomic dominant polycystic kidney disese (ADPKD), *Address correspondence to this author at the Dipartimento di Medicina Clinica e Molecolare, Cattedra di Nefrologia, Azienda Ospedaliera may underlie dysregulated epithelial cell proliferation and Sant’Andrea, Via di Grottarossa 1035-1039, 00189 Rome, Italy; thus cyst formation [10,11,12]. Notably, ADPKD is often Tel: +39 (06) 3377-5949; Fax: +39 (06) 3377-5866; associated with arterial hypertension, not necessarily through E-mail: [email protected]

1875-6506/12 $58.00+.00 © 2012 Bentham Science Publishers 182 Current Hypertension Reviews, 2012, Vol. 8, No. 3 Menè et al. the intermediation of chronic renal failure. Obviously, already obtained from studies in spontaneously hypertensive altered Na+ transport and balance through a defective, cystic rats [13-15]. tubular may be involved in the pathogenesis of the so-called “renoparenchymal” hypertension, often seen in THE TRP SUPERFAMILY OF ION CHANNELS chronic kidney disease even though GFR is preserved. Direct + 2+ Mammalian TRP proteins are an ever-growing list of links between tubular Na and altered Ca handling may cation-permeant homologues of the Drosophila gene also be at play in hypertensive congenital disorders such as transcripts, of which at least 28 members have been so far ADPKD. identified (5-7, 12). The TRP superfamily incorporates 3 Along the same line, the hypothesis has been put forward homologous subfamilies (TRPC, TRPV, and TRPA) and 4 that dysregulated Ca2+ entry through TRP channels in more distantly related subfamilies (TRPM,TRPP, TRPML, vascular smooth muscle cells (VSMC) or endothelial cells and TRPN/NOMPC) (Table 1). TRP proteins further of the pulmonary or systemic microcirculation may be assemble into heterotetramers, combinations of any given implicated in the pathogenesis of pulmonary or arterial isoform in ratios of 1:3, 1:2:1, etc. This implies that hypertension. Data consistent with this model have been electrophysiology and selectivity of a channel are dictated by

Table 1. Overview of Known TRP Channel Subfamilies and their Proposed Functions

Species

Human Murine Insect

TRPA 1 thermal (cold) sensing pungent compound sensing thermotaxis TRPC 1 2 sperm acrosome reaction pheromone sensing 3 neurodifferentiation synaptic development 4 vasorelaxation microvascular permeability 5 6 vascular tone regulation 7

TRPV 1 nociception 2 nociception 3 nociception 4 osmo-sensation 5 Ca2+ transport Ca2+ transport, kidney tubules 6 Ca2+ transport Ca2+ transport, duodenum OSM-9

TRPM 1 melanoma tumor suppressor 2 redox-sensing 3 oxidative stress-induced apoptosis 4 cation selective channel 5 cells 6 familial hypomagnesemia 7 cell survival, proliferation 8 thermal sensing, tumor marker

TRPN TRPL, Drosophila TRP phototransduction

TRPP 1 polycystic kidney disease 2 “ 3 “ 5 “

TRPML 1 mucolipin

TRP Channels and Hypertension Current Hypertension Reviews, 2012, Vol. 8, No. 3 183 composition of the assembled channels [4, 7]. A typical the list of proposed functions and organs expressing one or classical - or canonic - TRP protein is encoded by a gene more members of the superfamily is continuously expanding. comprised of 24 exons on 9q-21.12, yielding a The initial view of TRP as signalling receptors has evolved polypeptide chain of 1555 amino acids [4]. The general into that of multifunctional ion channels, which in structure includes 6 transmembrane domains with specialized epithelia (kidney, intestine) may even be adapted cytoplasmic N- and C-termini, incorporating a variety of for massive absorption of electrolytes [9, 23]. putative regulatory sites, including / inositol TRP CHANNELS AND PLASMA MEMBRANE Ca2+ (1,4,5)-trisphosphate (InsP ) receptor binding sites, PDZ 3 TRANSPORT binding motifs, phosphorylation sites for protein kinases, ankyrin-like repeats, proline-rich motifs, etc. [5-7, 12-19] The expression “capacitative Ca2+ influx” describes the (Fig. 1). A diacylglycerol (DAG) regulatory site has been ability of various stimuli of the discharge of intracellularly recently described on the N- terminus [18]. As a matter of stored Ca2+ (such as all agents that trigger phospholipase fact, many studies show direct regulation of TRP by DAG as C-γ breakdown of phosphoinositides, releasing InsP3) to in the case of TRPC3 expressed in Chinese hamster ovary initiate a Ca2+ current that refills the stores and accounts 2+ cells, or by related metabolites of phospholipase C, such as for a persistent elevation of [Ca ]i [1, 2, 12, 22, 24]. This polyunsaturated fatty acids or phosphatidylinositol 4,5- pathway for Ca2+ entry has often been referred to as store- 2+ 2+ bisphosphate (PIP2, 18,19). Molecular domains that are operated Ca channel (SOC) or ICRAC (intracellular Ca conserved in all members of the TRP family include release-activated Ca2+ influx channel) [1, 2, 22, 24-26]. In ankyrin-like repeats at the N- terminus and a highly the kidney, there is extensive evidence of SOC activity in conserved 25-amino acid stretch ("TRP domain") at the C- various compartments [12, 27-32]. Our own fluorimetric terminus, whose function is still unknown (Fig. 1). Despite experiments in cultured rat and human smooth muscle-like the substantial lack of positively charged amino acid residues glomerular mesangial cells (MC) have clearly shown that in transmembrane segment 4, typically found in voltage- Ca2+ influx is enhanced upon releasing intracellularly stored gated channels, where they prompt closure upon exposure to Ca2+ through receptor-operated and receptor-independent extracellular cations, at least one member of the superfamily, mechanisms [29-32]. Our lines of evidence, depicted in Fig. TRPM5, displays voltage modulation [20]. A striking feature (2), are multiple: First, known stimuli of phospholipase C of all members of the TRP superfamily is ubiquity and (angiotensin II, arginine vasopressin, thromboxane A2) flexibility: this means a highly conserved structure immediately release intracellularly stored Ca2+; this is 2+ throughout the evolutionary path, serving a remarkable followed by a sustained elevation of [Ca ]i only when the variety of functions in multiple tissues, as listed in Table 1 cells are bathed by media containing > 1 mmol/l Ca2+. [21, 22]. Most TRP channels are located in the central Second, MC equilibrated in a Ca2+-free solution display a nervous system and in peripheral sensing apparatuses, but substantial influx of Ca2+ when extracellular Ca2+ is added in

Fig. (1). A schematic diagram of the structure of two prototypical TRP channels with six transmembrane-spanning domains. TRPV6 is a duodenal epithelial Ca2+ channel (EcaC-2) / transporter (CaT1) displaying ankyrin repeats towards the N-terminus and at least 2 protein kinase C phosphorylation sites on both cytoplasmic tails. Ion fluxes occur through a “pore” between domains 5 and 6. TRPM6 (also labelled CHAK2) is a selective Mg2+ channel, incorporating a kinase domain near the C-terminus whose enzymatic activity is controlled by ATP binding (“chanzyme”, see text). Redrawn from Ref. 46. 184 Current Hypertension Reviews, 2012, Vol. 8, No. 3 Menè et al.

Fig. (2). Capacitative Ca2+ influx through store-operated Ca2+ channels in monolayer cultures of human glomerular mesangial cells (HMC). 2+ 2+ Panel A, a [Ca ]i transient induced by 1 uM angiotensin II (ANG II) in the presence (upper trace) or in the absence of extracellular Ca in 2+ 2+ 2+ the bathing media. Note disappearance of the persistent low-amplitude [Ca ]i elevation in Ca -free solutions. B, Ca influx upon previous discharge of intracellular Ca2+ stores by switching cells to a Ca2+-free solution. C, thapsigargin (THAPSI, 0.1 mM), a SERCA inhibitor, promotes subsequent Ca2+ influx upon release of intracellularly stored Ca2+. Fluorescence monitoring of fura 2-loaded monolayers of HMC, alternate excitation/emission 340/380 and 500 nm, respectively. a stepwise fashion. On the other hand, Ca2+-free solutions are strong argument for SOC influx occurring via a non-TRP known to quickly deplete intracellular stores, due to an protein is the poor selectivity of TRP channels for Ca2+ vs. outward leak mechanism. Third, releasing intracellular stores other cations [37, 38]. Even though this channel has also by agents that bypass InsP3-coupled receptors, as in the case been described as a receptor-operated channel (ROC), 2+ of the sarcoplasmic reticulum Ca -ATPase (SERCA) inasmuch as G-protein-linked receptors releasing InsP3 inhibitor, thapsigargin, produces a similar enhancement of appear involved, there seems to exist no direct binding of Ca2+ influx (Fig. 2). extracellular ligands to a TRP channel, whose conformation is instead intracellularly regulated by calmodulin / InsP , In the past decade, TRP proteins have been proposed as 3 PIP2, DAG, or protein kinases. InsP3 itself binds to a receptor the main mechanism underlying SOC activity. A large body 2+ expressed on Ca stores, such as the endo- or sarcoplasmic of evidence is consistent with this scheme [32-36]. For reticulum, the so-called calciosomes, mitochondria, etc. [18, example, cells from TRP KO mutants display minimal or no 19, 26, 39, 40]. Thus, it seems more appropriate to consider capacitative Ca2+ influx, while transfecting Xenopus oocytes Ca2+ itself as a second messenger, perhaps in concert with with TRP homologues elicits such Ca2+ currents (34,35). another store-associated, simultaneously released, putative However, recent work on the ORAI1 subunit of a tetrameric signalling molecule [26, 39, 40]. selective Ca2+ channel and the related stromal interaction molecule 1(STIM1) channel complex has questioned the Aside from its identity with TRP channels, SOC activity identification of SOC with TRP channels, perhaps as a result seems quite suited for the “tonic” regulation of vascular cell of different techniques for the assessment of Ca2+ influx, i.e. contractility, since it grants a constant influx of Ca2+, 2+ fluorescent dye imaging vs. patch-clamping [37, 38]. A amplifying the effects of quick bursts of [Ca ]i induced by TRP Channels and Hypertension Current Hypertension Reviews, 2012, Vol. 8, No. 3 185 vasoconstrictors [1, 2, 38]. In addition, TRP channels gated Ca2+ channels, where it acts as a Ca2+ sensor, calbindins by phospholipid metabolites (InsP3, PIP2, DAG) may well could fulfill a similar function through a specific linkage 2+ 2+ contribute to persistent elevations of [Ca ]i following Ca with TRPV5 and/or TRPV6. Indeed, colocalization of store discharge. Thus, low amounts of vasoactive agents may calbindins in most TRPV5/6-expressing tissues suggests exert enduring effects on the glomerular microcirculation. As a functional relationship between the two families of an example, it is known how autoregulation of renal blood Ca2+ carriers [8, 9, 46]. flow involves “tonic” angiotensin II (ANG II)-driven Expression of TRPV6 in the medullary collecting tubule, constriction of the efferent arteriole of glomeruli. Obviously, a segment of the nephron apparently not involved in Ca2+ the issue is also relevant to systemic circulation and other reabsorption, shows that not all potential sites of vascular districts, since MC are just a specialized reabsorption are operative [8, 9]. The genetic ablation of intraglomerular type of VSMC [27, 28]. Of note, cardiac tubular TRPV5 results in failure to reabsorb Ca2+ in null (-/-) myocytes and pulmonary artery SMC are endowed with mice, with resulting hypercalciuria and polyuria, leading to TRPC isoforms [41, 42], while gene silencing of SERCA altered bone mineral density, elevated circulating levels of pumps in cardiac myocytes (an approach similar to the 1,25-(OH) D , and skeletal abnormalities [8, 9, 46]. This is effects of thapsigargin) translates into upregulation of 2 3 clearly relevant to the pathogenesis of genetically based TRPC4 and -5 [43]. human disorders with hypercalciuria, from idiopathic TRP CHANNELS AND TUBULAR EPITHELIAL Ca2+ hypercalciuria to kidney stones, to various forms of TRANSPORT nephrocalcinosis, such as the “medullary sponge” kidney. Even osteoporosis has been linked to age- or hormonal- Recent studies have highlighted a major role of TRP dependent dysregulation of TRPV5 /-6 [47]. beyond the classical sensing/signalling functions. Transepithelial Ca2+ and Mg2+ fluxes involve TRP proteins TRP CHANNELS AND POLYCYSTIC KIDNEY in at least two major districts of pathophysiological DISEASE relevance, the duodenum and the renal distal convoluted Along with a variety of cell types across species, tubule. Cloning of TRPV5 from vitamin D-responsive rabbit epithelial cells are well endowed with TRP channels, often tubular epithelial cells, and of TRPV6 (which shares a 75% incorporated within the apical region of the plasma homology with the 5 isoform) from rat duodenum provided membrane. The PKD2 gene product, a TRP Ca2+-permeable, evidence that they are responsible for vitamin D-stimulated non selective channel protein of the P subfamily, isoforms 1- Ca2+ absorption in the distal nephron and in enterocytes, 5 [10, 11, 48-50], also termed polycystin 2, is an integral respectively [8, 9, 44, 45] (Fig. 1). Such channels have been component of the , a modified motile analogue of the previously referred to as epithelial Ca2+ channels 1 and 2 flagellum of protozoa or the tail of spermatozoa. Cilia are (EcaC1, EcaC2) [46]. It is noteworthy that TRPV5 and – 6 believed to act as sensors of luminal flow and/or solute tissue expression seems under the transcriptional control of 2+ concentration on most epithelia, including those of other 1,25 (OH) D , estrogens and plasma Ca levels, consistent 2 3 organs that develop cysts in ADPKD, such as the biliary with a candidate role in divalent ion homeostasis at the ducts of the liver. Individual epithelial cells or small clusters whole-body level [8, 9, 46]. In the distal tubule, Na+/ Ca2+ on the apical surface of renal tubules often exhibit a single exchange and Ca2+-ATPase activity at the basolateral cilium, bathed in the luminal fluid. Ciliary mechanosensation membrane of tubular cells provides the cytosolic inward may also bear relevance to embriology and biology of other gradient supporting reabsorption of filtered Ca2+ through organisms, such as C. elegans, which possesses homologues apical TRPV5 channels [46]. This apical, regulated pathway of PKD1 and 2 [48-50]. for Ca2+ reabsorption accounts for about 15% of total filtered Ca2+ recovery, whereas nearly 85% is reabsorbed at earlier Cilia are indeed very complex organelles, displaying an sites (proximal tubule, loop of Henle) through a paracellular intricated structure comprised of no less than 450 specialized basolateral electrochemical gradient mechanism [46]. proteins, organized in and around a bundle of 9 paired Intermediating cytosolic proteins link apical TRP channels to microfilaments originated from a central body, centrosome basolateral transporters.Two major subclasses of vitamin D- or centriole [49]. One defective component of cilia identified 2+ orpk dependent Ca - binding proteins, calbindin- D9K and in animal models of ADPKD, such as the Tg737 mouse or calbindin-D28K, have been identified as shuttles that could the KIF3A subunit of kinesin II KO mouse, is TRPP2. This bind Ca2+ and facilitate its cytosolic transfer [8, 9, 46]. protein shares 25% amino acid identity with TRPC3 and -6, Calbindin-D28K is highly conserved during evolution, and at the level of transmembrane spanning segments IV-VI. The expressed in kidney, small intestine, pancreas, placenta, homologues TRPP3 and -5 are reportedly not mutated in bone, and brain. The calbindins, like calmodulin (CaM), ADPKD. On the other hand, TRPP1, corresponding to parvalbumin, S100, and troponin C, belong to a group of PKD1, is a non-channel, larger polypeptide with 11 intracellular EF-hand Ca2+-binding proteins with marginal transmembrane–spanning regions, which may bind to and . The expression level of calbindins in regulate PKD2. The PKD2 protein may serve as a Ca2+ kidney and intestine closely correlates with the efficiency of gating mechanism activated by bending of the cilium under Ca2+ (re)absorption. Since CaM directly interacts with an the luminal flow [49]. Indeed, flow-stimulation of cilia in IQ motif at the carboxy termini of voltage-operated cultured epithelial cells results in Ca2+ entry [33]. On the 186 Current Hypertension Reviews, 2012, Vol. 8, No. 3 Menè et al. other hand, cells lacking PKD1 or the cilium itself display pathogenesis of the disorder [42, 56]. Since systemic, reduced Ca2+ responses upon exposure to a flow of culture “essential” arterial hypertension is associated in most media [21, 33]. If Ca2+-related signals control proliferation of instances with increased sensitivity to vasoconstrictors, and 2+ tubular epithelial cells, then it is easy to envision how altered [Ca ]i has often been reported in animal models and defective sensing by PKD1/PKD2 complexes may play a human hypertension, there is a potential for TRPC to be role in cyst formation and enlargement in target organs. involved in this area as well. Moreover, VSMC proliferation is regulated by [Ca2+] , inasmuch as extracellular Ca2+ or TRP CHANNELS AND HYPERTENSION i blockade of voltage-gated Ca2+ entry inhibit cell growth. Seven members of the canonical TRP channels (TRPC1- Vascular wall and myocardial hypertrophy in hypertension 7) are extensively expressed in the vasculature. TRPC are are events typically involving VSMC and/or myocardiocytes, 2+ gated by phospholipase C byproducts, with PIP2 controlling respectively. To the contrary, increased [Ca ]i appears to members of the subfamily TRPC1/4/5, and DAG acting on promote cell cycling and gene transcription [57]. The TRPC3,6,7 (2 is apparently not expressed in humans) [4, 7, correlation between TRPC6 expression and VSMC entry in 13, 51, 52]. They are both potential substrates for SOC the S and G2/M phases of the cell cycle shown by Yu et al. activity and involved in pulmonary and systemic points to store refilling via SOC as a major mechanism vasoconstriction and proliferation, particularly TRPC6. controlling hypertrophy of VSMC and perhaps myocardial Recent work links reduced TRPC channel activity to cells, of relevance to pulmonary and systemic hypertension impaired vasorelaxation in aortic rings, as in TRPC4-/- null [26, 39]. Links to operation of a Na+/Ca2+ exchanger mice or aortic endothelial cells [53]. Interestingly, the further stress the role of SOC in providing enhanced fluxes phenomenon is associated with reduced nitric oxide (NO) of Ca2+ in VSMC isolated from human pulmonary arteries synthesis and the resulting attenuation of agonist-induced [41]. vasorelaxation sensitive to L-nitroarginine. In the same Another route by which TRPC may relate to hyper- experimental model, thrombin-induced microvascular tension is the recent discovery that gain-of-function permeability is also attenuated, along with impaired mutations of TRPC6 are linked with familial forms of focal endothelial cell motility with altered stress-fiber morphology glomerulosclerosis (FSGS), a common cause of proteinuria, [54]. This lends support to the concept that TRP/SOC occasionally massive in the nephrotic range [40]. This likely operation is implicated in contractility of VSMC and/or occurs through insertion of TRPC6 into the glomerular endothelial function, in turn relevant to arterial hypertension basement membrane “slit diaphragm”, a complex intercellular [52]. meshwork of podocyte surface, electrically charged proteins. Recent work in rat carotid artery VSMC from It appears that increased Ca2+ entry through TRPC6 exerts spontaneously hypertensive rats (SHR) demonstrated profound effects on the cytoskeleton of podocytes, mostly upregulated TRPC3 channels, whereas TRPC1 activity was at the level of the intricate network calcineurin/NFAT/ reduced. As a result, contractility of denuded carotid arteries synaptopodin/Rho GTPase activity [40, 58]. As a result, from SHR was increased, as compared to normotensive collapse of the actin cytoskeleton leads to foot process WKY. It appears that TRPC3 enhanced VSMC depolarization effacement and proteinuria, a process that is typically with resulting activation of voltage-dependent Ca2+ channels antagonized by calcineurin inhibitors (cyclosporine A, and larger Ca2+ entry [15]. tacrolimus) and by antagonists of angiotensin II (ANG II). It is now being appreciated that the antiproteinuric effect of TRPC6 also seem implicated in the control of vascular ANG II blockade or CsA in FSGS, diabetic nephropathy and tone and hence of blood pressure, as shown by studies on the 2+ other hypertensive disorders of the kidney may be mediated Ca currents activated by α-adrenoceptors, and by the by interference with TRPC6-regulated Ca2+ signalling [40]. ability of antisense oligodeoxynucleotides (ODNs) to As recently shown by Eckel et al., TRPC6-null mice are TRPC6 to attenuate arterial SMC depolarization and protected from the proteinuric effects of ANG II, and TRPC6 constriction induced by elevated pressure in intact cerebral channels increase VSMC contractility [59]. Of interest, arteries [55]. A cation current involved in pressure-induced TRPC6-null mice were expected to be hypotensive, but myogenic depolarization is also significantly inhibited by instead were found by Dietrich et al. to have elevated BP and anti-TRPC6 ODNs, as well as by micromolar concentrations hypercontractile VSMC, perhaps through a compensatory of gadolinium, a blocking substitute for mono- or divalent upregulation of TRPC3 [60]. Interestingly, FSGS is one cations [56]. Other vascular isoforms include TRPC4, TRP5, of the glomerulopathies more frequently associated with and TRP7. The former is indeed present in VSMC, but hypertension, and antagonism of ANG II is a mainstay of exerts its primary activity within the endothelium, where antiproteinuric and antihypertensive therapy in all progressive it mediates vasorelaxation and vascular permeability glomerular disorders [40]. [13, 52]. TRP may also participate in other cellular functions Hyperexpression of TRPC3 and -6 in VSMC from related to vascular disorders, atherosclerosis and hyper- patients with idiopathic or familial pulmonary hypertension tension. Their known intervention in regulation of apoptosis (mean pulmonary artery pressure > 25 / 30 mmHg, rest / and cellular proliferative potential spans from upregulation exercise, respectively, with increased pulmonary vascular of TRPM members (melastatin) in tumors to promotion of resistance) points to a role of such channels in the TRP Channels and Hypertension Current Hypertension Reviews, 2012, Vol. 8, No. 3 187 apoptosis in response to reactive oxygen species (ROS), as in TRPV4 appears to modulate 5 expression in 2+ the case of TRPM2, sensitive to H2O2, ADP-ribose, and embryonic kidney cells, reducing it by a Ca -mediated TNF-α [61-64]. TRPM6 and TRPM7, fairly selective Mg2+ mechanism in settings of low osmolality [83]. In cells of the 2+ transporters controlling Mg2+ influx, appear differently cortical collecting duct, TRPV4 mediates Ca influx in regulated in VSMC cultures from SHR hypertensive/WKY response to shear stress and hypotonicity [84]. While this normotensive rats. TRPM7 expression is reportedly reduced data is preliminary and likely not unique to the kidney, the in SHR cells, resulting in lower levels of intracellular Mg2+, ability of TRP channels to regulate transepithelial ion fluxes which is internalized through the TRPM7 channel [65, 66] may have potential implications for the renal mechanisms of 2+ (Fig. 1). Reduced [Mg ]i is typically associated with hyper- arterial hypertension [50, 68, 77, 78]. tension, increased vascular sensitivity to vasoconstrictors, TRPV1 sensory channels have instead been implicated in impaired endothelium-dependent vasorelaxation. TRPM6 is the development of salt-sensitive hypertension, through upregulated by ANG II, whereas this peptide failed to affect studies in the Dahl rat model [85]. Salt-sensitive rats fed a TRPM7 levels in SHR-derived VSMC, resulting in reduced high salt diet develop hypertension and resistance to the translocation of annexin-1, a specific TRPM7 substrate [64- hypotensive effects of the TRPV1 agonist, . To the 66]. This may affect the proliferation/apoptosis balance in contrary, salt-resistant rats display enhanced sensitivity to VSMC, with implications for the cellular composition of TRPV1 channel activation and enhanced expression of vessels in hypertensive rats [65-69]. Moreover, direct TRPV1 in mesenteric arteries [85]. On the other hand, linkage between ANG II and expression or operation of TRP TRPV1 KO rats have normal blood pressure, but do respond channels is further evidence of their intermediary role in to DOCA-salt loading with hypertension and marked renal for vasoconstrictors, partly through the injury, as opposed to wild-type animals, confirming a persistent, “tonic” influx of Ca2+ (and perhaps Mg2+) 2+ “protective” role of TRPV1 against the pressor effects of following an initial release of stored Ca [1, 2, 70, 71]. Na+-loading, possibly relevant to human salt-sensitive Among other functions, TRPC Ca2+ channels serve as hypertension [86]. redox sensors in the vascular endothelium [61,72]. TRPC3 expression is a determinant of the NO sensitivity of SOC. CONCLUSIONS Experiments with TRPC species overexpressed in HEK293 The progressive disclosure of the complex network of cells confirmed that TRPC3 and TRPC4 encode for redox TRP cation channels in virtually all tissues is opening up sensitive cation channels [62,73]. ROS-induced protein new directions for research on ion transport and signalling. tyrosine phosphorylation and stimulation of phospholipase C These two apparently distant areas display now a linkage activity are involved in redox activation of TRPC3. In through multifunctional proteins that evolved in hundreds of addition, oxidative stress-induced disruption of caveolin 1- variants to best suit the needs of different organs. Involvment rich lipid raft domains, which interfere with functional of divalent cations, particularly Ca2+, in signalling has been TRPC channels, is likely to contribute to redox modulation known for almost two decades. What is now emerging is that of TRP proteins and to oxidative stress-induced changes in the same channels that enable nanosecond action potentials 2+ cellular Ca signalling [62]. in the sensing apparatus of insects, may serve to detect flow Depression of Ca2+ currents can reduce apoptosis, while, in the distal nephron of mammalians, and at the same time as to the contrary, increasing Ca2+ influx may promote cell carriers for mass ion absorption and excretion through death [74]. Certain pathological redox states may induce transport epithelia. This is certainly remarkable in terms of cell death through activation of Ca2+ fluxes mediated by the evolution of molecular structures, and even more striking TRP proteins, as shown for TRPM2 [74, 75]. This protein, evidence of a parallel function of ion carriers as sensors of earlier labelled LTRPC2 or TRPC7, contains an ADP-ribose the surrounding environment. pyrophosphatase domain, which catalyzes the hydrolysis of Besides helping to unravel the pathophysiology of long- nucleoside diphosphates [63, 64, 68, 75]. The unique enzyme known genetic and acquired diseases, a promising area of activity exhibited by certain TRP channels has suggested the development is represented by pharmacologic inhibition of use of the term “chanzymes”. While inactive in TRPM2, the non-voltage-gated Ca2+ entry (87,88). A novel such class of Nudix hydrolase domain at the C-terminus is believed to link drugs may prove additive or alternative to traditional Ca2+- 2+ nucleotides (ADP-ribose, NAD), which then promote Ca channel blockers in the treatment of a host of diverse entry and apoptosis of cells exposed to ROS or a high redox cardiovascular and renal disorders, including arterial microenvironment [60, 62, 66, 72-75]. These responses hypertension, arrhytmias, mineral and electrolyte appear fairly ubiquitous, and therefore likely to play a role in disturbances, stone formation and inborn cystic disorders. vascular cell biology, where oxidative stress, leukocyte infiltration and disorderly proliferation of endothelial cells DISCLOSURE are associated with atherosclerotic plaques, in turn a strong correlate of arterial hypertension [76-81]. This manuscript is an extended /updated version of our previously published manuscript entitled: “Transient Vanilloid TRP also appear to have an intriguing role as receptor potential channels, the kidney and hypertension”. sensors and regulators of flow and osmolality in various Current Hypertension Reviews 2006; 2:61-67. epithelia, among which the mammalian distal nephron [82]. 188 Current Hypertension Reviews, 2012, Vol. 8, No. 3 Menè et al.

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Received: February 24, 2012 Revised: May 29, 2012 Accepted: May 30, 2012