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PubMed2000

Cloning and expression of the human transient receptor potential 4 (TRP4) gene: localization and functional expression of human TRP4 and TRP3.

Mammalian homologues of the Drosophila transient receptor potential (TRP) protein have been proposed to function as ion channels, and in some cases as store-operated or capacitative calcium entry channels. However, for each of the mammalian TRP proteins, different laboratories have reported distinct modes of cellular regulation. In the present study we describe the cloning and functional expression of the human form of TRP4 (hTRP4), and compare its activity with another well studied protein, hTRP3. When hTRP4 was transiently expressed in human embryonic kidney (HEK)-293 cells, basal bivalent cation permeability (barium) was increased. Whole-cell patch-clamp studies of hTRP4 expressed in Chinese hamster ovary cells revealed a constitutively active non-selective cation current which probably underlies the increased bivalent cation entry. Barium entry into hTRP4-transfected HEK-293 cells was not further increased by phospholipase C (PLC)-linked receptor activation, by intracellular calcium store depletion with thapsigargin, or by a synthetic diacylglycerol, 1-oleoyl-2-acetyl-sn-glycerol (OAG). In contrast, transient expression of hTRP3 resulted in a bivalent cation influx that was markedly increased by PLC-linked receptor activation and by OAG, but not by thapsigargin. Despite the apparent differences in regulation of these two putative channel proteins, green fluorescent protein fusions of both molecules localized similarly to the plasma-membrane, notably in discrete punctate regions suggestive of specialized signalling complexes. Our findings indicate that while both hTRP4 and hTRP3 can apparently function as cation channels, their putative roles as components of capacitative calcium entry channels are not readily demonstrable by examining their behaviour when exogenously expressed in cells.

Amino Acid Sequence↗
PubMed1998

A novel capacitative calcium entry channel expressed in excitable cells.

In addition to voltage-gated calcium influx, capacitative calcium entry (CCE) represents a major pathway for calcium entry into the cell. Here we report the structure, expression and functional properties of a novel CCE channel, TRP5. This channel is a member of a new subfamily of mammalian homologues of the Drosophila transient receptor potential (TRP) protein, now comprising TRP5 (also CCE2) and the structurally related CCE1 (also TRP4). Like TRP4, TRP5 forms ion channels mainly permeable for Ca2+ which are not active under resting conditions but can be activated by manoeuvres known to deplete intracellular calcium stores. Accordingly, dialysis of TRP5-expressing cells with inositol-(1,4,5)-trisphosphate evokes inward rectifying currents which reversed polarity at potentials more positive than +30 mV. Ca2+ store depletion with thapsigargin induced TRP5-mediated calcium entry dependent on the concentration of extracellular calcium, as seen by dual wavelength fura-2 fluorescence ratio measurements. TRP5 transcripts are expressed almost exclusively in brain, where they are present in mitral cells of the olfactory bulb, in lateral cerebellar nuclei and, together with TRP4 transcripts, in CA1 pyramidal neurons of the hippocampus, indicating the presence of CCE channels in excitable cells and their participation in neuronal calcium homeostasis.

Amino Acid Sequence↗
PubMed2004

Chronic hypoxia-induced upregulation of store-operated and receptor-operated Ca2+ channels in pulmonary arterial smooth muscle cells: a novel mechanism of hypoxic pulmonary hypertension.

Chronic hypoxic pulmonary hypertension is associated with profound vascular remodeling and alterations in Ca(2+) homeostasis in pulmonary arterial smooth muscle cells (PASMCs). Recent studies show that transient receptor potential (TRPC) genes, which encode store-operated and receptor-operated cation channels, play important roles in Ca(2+) regulation and cell proliferation. However, the influence of chronic hypoxia on TRPC channels has not been determined. Here we compared TRPC expression, and store- and receptor-operated Ca(2+) entries in PASMCs of normoxic and chronic hypoxic rats. Reverse-transcription polymerase chain reaction (RT-PCR), Western blot, and immunostaining showed consistently that TRPC1, TRPC3, and TRPC6 were expressed in intralobar pulmonary arteries (PAs) and PASMCs. Application of 1-oleoyl-2-acetyl-sn-glycerol (OAG) to directly activate receptor-operated channels, or thapsigargin to deplete Ca(2+) stores, caused dramatic increase in cation entry measured by Mn(2+) quenching of fura-2 and by Ca(2+) transients. OAG-induced responses were approximately 700-fold more resistant to La(3+) inhibition than thapsigargin-induced responses. siRNA knockdown of TRPC1 and TRPC6 specifically attenuated thapsigargin- and OAG-induced cation entries, respectively, indicating that TRPC1 mediates store-operated entry and TRPC6 mediates receptor-operated entry. In hypoxic PAs, there were 2- to 3-fold increases in TRPC1 and TRPC6 expression. They were accompanied by significant increases in basal, OAG-induced, and thapsigargin-induced cation entries in hypoxic PASMCs. Moreover, removal of Ca(2+) or inhibition of store-operated Ca(2+) entry with La(3+) and SK&F-96365 reversed the elevated basal [Ca(2+)](i) in PASMCs and vascular tone in PAs of chronic hypoxic animals, but nifedipine had minimal effects. Our results for the first time to our knowledge show that both store- and receptor-operated channels of PASMCs are upregulated by chronic hypoxia and contribute to the enhanced vascular tone in hypoxic pulmonary hypertension.

Animals↗
PubMed1998

Store-operated calcium entry promotes shape change in pulmonary endothelial cells expressing Trp1.

Activation of Ca2+ entry is known to produce endothelial cell shape change, leading to increased permeability, leukocyte migration, and initiation of angiogenesis in conduit-vessel endothelial cells. The mode of Ca2+ entry regulating cell shape is unknown. We hypothesized that activation of store-operated Ca2+ channels (SOCs) is sufficient to promote cell shape change necessary for these processes. SOC activation in rat pulmonary arterial endothelial cells increased free cytosolic Ca2+ that was dependent on a membrane current having a net inward component of 5.45 +/- 0.90 pA/pF at -80 mV. Changes in endothelial cell shape accompanied SOC activation and were dependent on Ca2+ entry-induced reconfiguration of peripheral (cortical) filamentous actin (F-actin). Because the identity of pulmonary endothelial SOCs is unknown, but mammalian homologues of the Drosophila melanogaster transient receptor potential (trp) gene have been proposed to form Ca2+ entry channels in nonexcitable cells, we performed RT-PCR using Trp oligonucleotide primers in both rat and human pulmonary arterial endothelial cells. Both cell types were found to express Trp1, but neither expressed Trp3 nor Trp6. Our study indicates that 1) Ca2+ entry in pulmonary endothelial cells through SOCs produces cell shape change that is dependent on site-specific rearrangement of the microfilamentous cytoskeleton and 2) Trp1 may be a component of pulmonary endothelial SOCs.

Amino Acid Sequence↗
PubMed2003

TRPC3-like protein is involved in the capacitative cation entry induced by 1alpha,25-dihydroxy-vitamin D3 in ROS 17/2.8 osteoblastic cells.

In ROS 17/2.8 rat osteoblastic-like cells a capacitative Ca(2+) entry (CCE) pathway operates which is activated by either 1alpha,25-dihydroxy-vitamin D3 (1alpha,25(OH)(2)D3 or thapsigargin (Tpg)-induced depletion of Ca(2+) stores. In view of recent evidence favoring a role for transient receptor potential (TRP) proteins in mediating CCE, we investigated if channels involved in the 1alpha,25(OH)(2)D3-sensitive CCE in rat osteoblasts were related to an endogenous TRP-canonical (TRPC) isoform homologue. By reverse transcription (RT)-PCR using mRNA from ROS 17/2.8 cells and primers based on conserved regions within the mammalian TRPC3/6/7 subfamily, two fragments were amplified of 390 and 201 bp with 100 and 94% sequence identity, respectively, with human TRPC3. Northern blot analysis showed the presence of a 3.5 kb transcript and both immunobloting and immunocytochemistry using a specific anti-TRPC3 antibody confirmed endogenous expression of a TRPC3-like protein ( approximately 110 kDa) with membrane localization. In ROS 17/2.8 cells intranuclearly microinjected with anti-TRPC3 antisense oligodeoxynucleotides (ODN), both the initial rate and magnitude of CCE activated by either 1alpha,25(OH)(2)D3 or Tpg were markedly reduced, whereas no changes were detected in control-injected cells. The present findings constitute the first evidence to date suggesting that an endogenous TRPC3-like protein is functionally involved in the CCE route activated by 1alpha,25(OH)(2)D3 in a secosteroid target cell. We anticipate TRPC3 as a candidate for mediating store-operated non-selective cation entry into osteoblasts.

Animals↗
PubMed2006

Cyclin-dependent kinase 5 modulates nociceptive signaling through direct phosphorylation of transient receptor potential vanilloid 1.

Transient receptor potential vanilloid 1 (TRPV1), a ligand-gated cation channel highly expressed in small-diameter sensory neurons, is activated by heat, protons, and capsaicin. The phosphorylation of TRPV1 provides a versatile regulation of intracellular calcium levels and is critical for TRPV1 function in responding to a pain stimulus. We have previously reported that cyclin-dependent kinase 5 (Cdk5) activity regulates nociceptive signaling. In this article we report that the Cdk5-mediated phosphorylation of TRPV1 at threonine-407 can modulate agonist-induced calcium influx. Inhibition of Cdk5 activity in cultured dorsal root ganglia neurons resulted in a significant reduction of TRPV1-mediated calcium influx, and this effect could be reversed by restoring Cdk5 activity. Primary nociceptor-specific Cdk5 conditional-knockout mice showed reduced TRPV1 phosphorylation, resulting in significant hypoalgesia. Thus, the present study indicates that Cdk5-mediated TRPV1 phosphorylation is important in the regulation of pain signaling.

Amino Acid Sequence↗
PubMed2001

Regulation of melastatin, a TRP-related protein, through interaction with a cytoplasmic isoform.

The TRP (transient receptor potential) superfamily includes a group of subfamilies of channel-like proteins mediating a multitude of physiological signaling processes. The TRP-melastatin (TRPM) subfamily includes the putative tumor suppressor melastatin (MLSN) and is a poorly characterized group of TRP-related proteins. Here, we describe the identification and characterization of an additional TRPM protein TRPM4. We reveal that TRPM4 and MLSN each mediate Ca(2+) entry when expressed in HEK293 cells. Furthermore, we demonstrate that a short form of MLSN (MLSN-S) interacts directly with and suppresses the activity of full-length MLSN (MLSN-L). This suppression seems to result from the inhibition of translocation of MLSN-L to the plasma membrane. We propose that control of translocation through interaction between MLSN-S and MLSN-L represents a mode for regulating ion channel activity.

Amino Acid Sequence↗
PubMed2025

Structural Basis of Cold and Menthol Sensing by TRPM8.

The transient receptor potential melastatin member 8 (TRPM8) is a polymodal ion channel that senses cold and menthol in mammals. Despite prior structural studies, the mechanisms by which cold and menthol activate TRPM8 remain unresolved. Here, we present cryo-EM structures representing the cold and menthol-dependent activation trajectories, combined with extensive functional analyses. We captured snapshots of cooling-dependent pore opening, which involves dramatic pore rearrangement, suggesting a mechanism for cold sensing. Moreover, menthol binds dynamically to induce channel activation, which may underlie menthol specificity for TRPM8. Finally, we show how TRPM8 integrates multiple modalities (cold and menthol) through overlapping but non-identical pathways, revealing the temperature-specific "cold spot". These findings enhance our understanding of the molecular basis of physically and chemically induced cool sensation in mammals.

Journal Article↗
PubMed2007

Transcription of rat TRPV1 utilizes a dual promoter system that is positively regulated by nerve growth factor.

The capsaicin receptor, also known as 'transient receptor potential vanilloid receptor subtype 1' (TRPV1, VR1), is an ion channel subunit expressed in primary afferent nociceptors, which plays a critical role in pain transduction and thermal hyperalgesia. Increases in nociceptor TRPV1 mRNA and protein are associated with tissue injury-inflammation. As little is understood about what controls TRPV1 RNA transcription in nociceptors, we functionally characterized the upstream portion of the rat TRPV1 gene. Two functional rTRPV1 promoter regions and their transcription initiation sites were identified. Although both promoter regions directed transcriptional activity in nerve growth factor (NGF) treated rat sensory neurons, the upstream Core promoter was the most active in cultures enriched in sensory neurons. Because NGF is a key modulator of inflammatory pain, we examined the effect of NGF on rTRPV1 transcription in PC12 cells. NGF positively regulated transcriptional activity of both rTRPV1 promoter regions in PC12 cells. We propose that the upstream regulatory region of the rTRPV1 gene is composed of a dual promoter system that is regulated by NGF. These findings support the hypothesis that NGF produced under conditions of tissue injury and/or inflammation directs an increase of TRPV1 expression in nociceptors in part through a transcription-dependent mechanism.

Animals↗
PubMed2006

TRPM8 in prostate cancer cells: a potential diagnostic and prognostic marker with a secretory function?

During the past 5 years it has emerged that the transient receptor potential (TRP) family of Ca(2+)-and Na(+)-permeable channels plays a diverse and important role in cell biology and in pathology. One member of this family, TRPM8, is highly expressed in prostate cancer cells but the physiological and pathological functions of TRPM8 in these cells are not known. Here we address these questions, and the issue of whether or not TRPM8 is an effective diagnostic and prognostic marker in prostate cancer. TRPM8 is known to be activated by cool stimuli (17-25 degrees C) and cooling compounds such as menthol. The activation mechanism(s) involves voltage sensing of membrane potential, phosphatidylinositol 4,5-bisphosphate and Ca(2+). In addition to prostate cancer cells, TRPM8 is expressed in sensory neurons where it acts as a sensor of cold. In prostate epithelial cells, expression of TRPM8 is regulated by androgen and is elevated in androgen-sensitive cancerous cells compared with normal cells. While there is some evidence that in prostate cancer cells Ca(2+) and Na(+) inflow through TRPM8 is necessary for survival and function, including secretion at the apical membrane, the function of TRPM8 in these cells is not really known. It may well differ from the role of TRPM8 as a cool sensor in sensory nerve cells. Androgen unresponsive prostate cancer is difficult to treat effectively and there are limited diagnostic and prognostic markers available. TRPM8 is a potential tissue marker in differential diagnosis and a potential prognostic marker for androgen-unresponsive and metastatic prostate cancer. As a consequence of its ability to convey Ca(2+) and Na(+) and its expression in only a limited number of cell types, TRPM8 is considered to be a promising target for pharmaceutical, immunological and genetic interventions for the treatment of prostate cancer.

Biomarkers, Tumor↗
PubMed2002

Comparison of human TRPC3 channels in receptor-activated and store-operated modes. Differential sensitivity to channel blockers suggests fundamental differences in channel composition.

Capacitative calcium entry or store-operated calcium entry in nonexcitable cells is a process whereby the activation of calcium influx across the plasma membrane is signaled by depletion of intracellular calcium stores. Transient receptor potential (TRP) proteins have been proposed as candidates for store-operated calcium channels. Human TRPC3 (hTRPC3), an extensively studied member of the TRP family, is activated through a phospholipase C-dependent mechanism, not by store depletion, when expressed in HEK293 cells. However, store depletion by thapsigargin is sufficient to activate hTRPC3 channels when expressed in DT40 avian B-lymphocytes. To gain further insights into the differences between hTRPC3 channels generated in these two expression systems and further understand the role of hTRPC3 in capacitative calcium entry, we examined the effect of two well characterized inhibitors of capacitative calcium entry, Gd3+ and 2-aminoethoxydiphenyl borane (2APB). We confirmed that in both DT40 cells and HEK293 cells, 1 microm Gd3+ or 30 microm 2APB completely blocked calcium entry due to receptor activation or store depletion. In HEK293 cells, 1 microm Gd3+ did not block receptor-activated hTRPC3-mediated cation entry, whereas 2APB had a partial (approximately 60%) inhibitory effect. Interestingly, store-operated hTRPC3-mediated cation entry in DT40 cells was also partially inhibited by 2APB, whereas 1 microm Gd3+ completely blocked store-operated hTRPC3 activity in these cells. Furthermore, the sensitivity of store-operated hTRPC3 channels to Gd3+ in DT40 cells was similar to the endogenous store-operated channels, with essentially 100% block of activity at concentrations as low as 0.1 microm. Finally, Gd3+ has a rapid inhibitory effect when added to fully developed hTRPC3-mediated calcium entry, suggesting a direct action of Gd3+ on hTRPC3 channels. The distinct action of these inhibitors on hTRPC3-mediated cation entry in these two cell types may result from their different modes of activation and may also reflect differences in basic channel structure.

Boron Compounds↗
PubMed2001

Dissecting receptor-mediated Ca2+ influx pathways: TRP channels and their native counterparts.

Cellular stimulation from the surrounding extracellular environment via receptors and other pathways evoke activation of Ca2+-permeable cation channels that form essential signaling pathways in controlling biological responses. An important clue to understand the molecular mechanisms underlying these cation channels (tentatively termed as receptor-mediated cation channels (RMCC)) was first provided through molecular studies of the transient receptor potential (trp) protein (TRP), which controls light-induced depolarization in Drosophila photoreceptor cells. Use of the genetic information and recombinant expression technique lead to the discovery of numerous mammalian TRP homologues revealing novel RMCCs. In this review, we focus on the dramatic progress in the molecular investigation of RMCC in mammalian systems. The recent findings should provide powerful tools for the development of novel pharmaceutical targets.

Animals↗
PubMed2000

Electrophysiological evidence for an ATP-gated ion channel in the principal cells of the frog skin epithelium.

In the present study we investigated the effects of adenosine 5'-triphosphate (ATP) on Na+ transport in frog skin epithelium. An experimental set-up was constructed to allow simultaneous measurement of Na+ transport, measured as the amiloride-sensitive short circuit current (Isc), and free cytosolic Ca2+ concentration ([Ca2+]i) measured with the Ca(2+)-sensitive dye fura-2. The cell potential (Vsc) was measured with microelectrodes. Addition of ATP (100 micrM) to the basolateral solution resulted in a fast transient decrease in Isc followed by a slower increase and a transient increase in [Ca2+]i. Microelectrode measurements showed that the primary response, i.e. the decline in Isc was accompanied by transient depolarisation, followed by a return to the control value. The decrease in current was Ca2+ independent; i.e. treatment with thapsigargin in Ca(2+)-free solutions abolished the Ca2+ transient but did not influence the current transient. The secondary response, i.e. the slow increase in current, was accompanied by slow depolarisation of the cell. Measurements of apical Na+ permeability showed that this was due to an opening or activation of apical Na+ channels. These data show that ATP causes a fast initial drop and a secondary, long-lasting increase in Na+ absorption. The ability of ATP to cause the initial decline in current is independent of Ca2+, i.e. it is not caused by secondary effects of the P2Y-type receptors present in the tissue. Measurements of intracellular potential indicate that the initial depolarisation is caused by opening of non-selective cation channels, suggesting that this decrease is due to a transient activation of P2X-type ATP receptors.

Adenosine Triphosphate↗
PubMed2000

Functional significance of human trp1 and trp3 in store-operated Ca(2+) entry in HEK-293 cells.

The Drosophila trp (transient receptor potential) gene appears to encode the Drosophila store-operated channel (SOC), and some mammalian trp homologues have been proposed to encode mammalian SOCs. This study provides evidence for the expression of three trp homologues (Mtrp2, Mtrp3, and Mtrp4) in fibroblasts from wild-type and src knockout mice, and four trp homologues (Htrp1, Htrp3, Htrp4, and Htrp6) in human embryonic kidney (HEK-293) cells based on RT-PCR techniques. In HEK-293 cells stably transfected with a 323-bp Htrp3 antisense construct (Htrp3AS), Northern blot analysis revealed that the expression of a 4-kb transcript was dramatically suppressed in comparison to that observed in cells stably transfected with a short Htrp3 sense construct (Htrp3S). Activity of SOCs, monitored as Ba(2+) influx following Ca(2+) store depletion with thapsigargin, was reduced by 32% in Htrp3AS cells in comparison with Htrp3S cells. Transient transfection of a 369-bp Htrp1 antisense construct in cells stably expressing Htrp3AS induced a higher level of inhibition (55%) of store-operated Ca(2+) entry. These data suggest that Htrp1 and Htrp3 may be functional subunits of SOCs.

Animals↗
PubMed2005

Emerging roles of TRPM6/TRPM7 channel kinase signal transduction complexes.

Investigations into Drosophila mutants with impaired vision due to mutations in the transient receptor potential gene (trp) initiated a systematic search for TRP homologs in other species, finally leading to the discovery of a whole new family of plasma membrane cation channels involved in multiple physiological processes. Among the recently discovered TRP cation channels two homologous proteins, TRPM6 and TRPM7, display unique domain compositions and biophysical properties. These remarkable genes are vital for Mg(2+) homeostasis in vertebrates and, if disrupted, lead to cell death or human disease.

Animals↗
PubMed2002

Expression and role of TRPC proteins in human platelets: evidence that TRPC6 forms the store-independent calcium entry channel.

Store-operated Ca(++) entry (SOCE) is thought to comprise the major pathway for Ca(++) entry in platelets. Recently, a number of transient receptor potential (TRP) proteins, which have been divided into 3 groups (TRPC, TRPM, and TRPV), have been suggested as SOCE channels. We report the expression and function of TRPC proteins in human platelets. TRPC6 is found at high levels and TRPC1 at low levels. Using purified plasma (PM) and intracellular membranes (IM), TRPC6 is found in the PM, but TRPC1 is localized to the IM. Using Fura-2-loaded platelets, we report that, in line with TRPC6 expression, 1-oleoyl-2-acetyl-sn-glycerol (OAG) stimulated the entry of Ca(++) and Ba(2+) independently of protein kinase C. Thrombin also induced the entry of Ca(++) and Ba(2+), but thapsigargin, which depletes the stores, induced the entry of only Ca(++). Thus, thrombin activated TRPC6 via a SOCE-independent mechanism. In phosphorylation studies, we report that neither TRPC6 nor TRPC1 was a substrate for tyrosine kinases. TRPC6 was phosphorylated by cAMP-dependent protein kinase (cAMP-PK) and associated with other cAMP-PK substrates. TRPC1 was not phosphorylated by cAMP-PK but also associated with other substrates. Activation of cAMP-PK inhibited Ca(++) but not Ba(2+) entry induced by thrombin and neither Ca(++) nor Ba(2+) entry stimulated by OAG. These results suggest that TRPC6 is a SOCE-independent, nonselective cation entry channel stimulated by thrombin and OAG. TRPC6 is a substrate for cAMP-PK, although phosphorylation appears to not affect cation permeation. TRPC1 is located in IM, suggesting a role at the level of the stores.

Amino Acid Sequence↗
PubMed2005

TRPC1 functions as a store-operated Ca2+ channel in intestinal epithelial cells and regulates early mucosal restitution after wounding.

An increase in cytosolic free Ca(2+) concentration ([Ca(2+)](cyt)) results from Ca(2+) release from intracellular stores and extracellular Ca(2+) influx through Ca(2+)-permeable ion channels and is crucial for initiating intestinal epithelial restitution to reseal superficial wounds after mucosal injury. Capacitative Ca(2+) entry (CCE) induced by Ca(2+) store depletion represents a major Ca(2+) influx mechanism, but the exact molecular components constituting this process remain elusive. This study determined whether canonical transient receptor potential (TRPC)1 served as a candidate protein for Ca(2+)-permeable channels mediating CCE in intestinal epithelial cells and played an important role in early epithelial restitution. Normal intestinal epithelial cells (the IEC-6 cell line) expressed TRPC1 and TPRC5 and displayed typical records of whole cell store-operated Ca(2+) currents and CCE generated by Ca(2+) influx after depletion of intracellular stores. Induced TRPC1 expression by stable transfection with the TRPC1 gene increased CCE and enhanced cell migration during restitution. Differentiated IEC-Cdx2L1 cells induced by forced expression of the Cdx2 gene highly expressed endogenous TRPC1 and TRPC5 and exhibited increased CCE and cell migration. Inhibition of TRPC1 expression by small interfering RNA specially targeting TRPC1 not only reduced CCE but also inhibited cell migration after wounding. These findings strongly suggest that TRPC1 functions as store-operated Ca(2+) channels and plays a critical role in intestinal epithelial restitution by regulating CCE and intracellular [Ca(2+)](cyt).

Animals↗
PubMed2006

Capsaicin inhibits catecholamine secretion and synthesis by blocking Na+ and Ca2+ influx through a vanilloid receptor-independent pathway in bovine adrenal medullary cells.

We report here the effects of capsaicin, a flavoring ingredient in the hot pepper Capsicum family, on catecholamine secretion and synthesis in cultured bovine adrenal medullary cells. Capsaicin inhibited catecholamine secretion (IC(50)=9.5, 11.8, and 62 microM) stimulated by carbachol, an agonist of the nicotinic acetylcholine receptor, by veratridine, an activator of voltage-dependent Na(+) channels, and by high K(+), an activator of voltage-dependent Ca(2+) channels, respectively. Capsaicin also suppressed carbachol-induced (22)Na(+) influx (IC(50)=5.0 microM) and (45)Ca(2+) influx (IC(50)=24.4 muM), veratridine-induced (22)Na(+) influx (IC(50)=2.4 microM) and (45)Ca(2+) influx (IC(50)=1.1 microM), and high K(+)-induced (45)Ca(2+) influx (IC(50)=5.8 microM). The reduction in catecholamine secretion caused by capsaicin was not overcome by increasing the concentration of carbachol. Furthermore, capsazepine (10 microM), a competitive antagonist for the transient receptor potential vanilloid 1, and ruthenium red (30 microM), a nonselective cation channel antagonist, did not block the inhibition by capsaicin of catecholamine secretion. Capsaicin also suppressed both basal and carbachol-stimulated (14)C-catecholamine synthesis (IC(50)=10.6 and 26.4 microM, respectively) from [(14)C] tyrosine but not from L: -3, 4-dihydroxyphenyl [3-(14)C] alanine ([(14)C] DOPA) as well as tyrosine hydroxylase activity (IC(50)=8.4 and 39.0 microM, respectively). The present findings suggest that capsaicin inhibits catecholamine secretion and synthesis via suppression of Na(+) and Ca(2+) influx through a vanilloid receptor-independent pathway.

Adrenal Medulla↗