Acidification of rat TRPV1 alters the kinetics of capsaicin responses

Neuroscience Research, Global Pharmaceutical Research and Development, Abbott Laboratories, R04PM, AP9A, Abbott Park, IL 60064-6123, USA.
Molecular Pain (Impact Factor: 3.53). 02/2005; 1:28. DOI: 10.1186/1744-8069-1-28
Source: PubMed

ABSTRACT TRPV1 (vanilloid receptor 1) receptors are activated by a variety of ligands such as capsaicin, as well as by acidic conditions and temperatures above 42 degrees C. These activators can enhance the potency of one another, shifting the activation curve for TRPV1 to the left. In this study, for example, we observed an approximately 10-fold shift in the capsaicin EC50 (640 nM to 45 nM) for rat TRPV1 receptors expressed in HEK-293 cells when the pH was lowered from 7.4 to 5.5. To investigate potential causes for this shift in capsaicin potency, the rates of current activation and deactivation of whole-cell currents were measured in individual cells exposed to treatments of pH 5.5, 1 microM capsaicin or in combination. Acidic pH was found to both increase the activation rate and decrease the deactivation rate of capsaicin-activated currents providing a possible mechanism for the enhanced potency of capsaicin under acidic conditions. Utilizing a paired-pulse protocol, acidic pH slowed the capsaicin deactivation rate and was readily reversible. Moreover, the effect could occur under modestly acidic conditions (pH 6.5) that did not directly activate TRPV1. When TRPV1 was maximally activated by capsaicin and acidic pH, the apparent affinity of the novel and selective capsaicin-site competitive TRPV1 antagonist, A-425619, was reduced approximately 35 fold. This shift was overcome by reducing the capsaicin concentration co-applied with acidic pH. Since inflammation is associated with tissue acidosis, these findings enhance understanding of TRPV1 receptor responses in inflammatory pain where tissue acidosis is prevalent.

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Available from: Ping Han, Jul 19, 2015
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    • " activating effect of low pH o on TRPV1 activity . Most in vitro studies have not investigated this possibility , because they were designed to study the effects of acidosis on TRPV1 activity using a whole - cell patch - clamp configuration with a Hepes ( 5 – 10 mM ) - containing pipette solution ( Tominaga et al . 1998 ; Ryu et al . 2003 , 2007 ; Neelands et al . 2005 ) . Hepes is a potent acid – base buffer , so pH i changes induced by extracellular acidosis can be artificially prevented by a Hepes - containing pipette solution . Therefore , to determine the effects of extracellular acidosis on TRPV1 activity more precisely , it is essential to examine the effect of extracellular acidosis on TRPV1 i"
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    ABSTRACT: Transient receptor potential V1 (TRPV1) has been suggested to play an important role in detecting decreases in extracellular pH (pH(o)). Results from recent in vivo studies, however, have suggested that TRPV1 channels play less of a role in sensing a moderately acidic pH(o) (6.0 < pH < 7.0) than predicted from the in vitro experiments. A clear explanation for this discrepancy between the in vitro and in vivo data has not yet been provided. We report here that intracellular acidification induced by a moderately low pH(o) (6.4) almost completely inhibited the effect of extracellular acidosis on TRPV1 activity. In our experiments, sodium acetate (20 mm), which was used to induce intracellular acidosis, attenuated the capsaicin-evoked TRPV1 current (I(CAP)) in a reversible manner in whole-cell patch-clamp mode and shifted the concentration-response curve to the right. Likewise, the concentration-response curve was significantly shifted to the right by lowering the pH of the pipette solution from 7.2 to 6.5. In addition, application of an acidic bath solution (pH 6.4) to the intracellular side also significantly suppressed I(CAP) in inside-out patch mode. In cell-attached patch mode, the single-channel activity of i(CAP) was significantly attenuated by intracellular acidosis that was induced by a decrease in pH(o) (6.4). These results suggested that intracellular acidification induced by a low pH(o) inhibited TRPV1 activity. When studied in perforated patch mode or by acidifying the intracellular pipette solution, potentiation or activation of TRPV1 by extracellular acidosis (pH 6.4) at 37 °C was almost completely inhibited. Likewise, enhancement of neuronal excitability by a moderately acidic pH(o) (6.4) at a physiological temperature (37 °C) was attenuated by lowering the pH of the pipette solution to 6.5 or using perforated patch mode. Taken together, these results suggest that extracellular acidosis of moderate intensity may not significantly modulate TRPV1 activity in physiological conditions at which intracellular pH can be readily affected by pH(o), and this phenomenon is due to attenuation of TRPV1 channel activity by low-pH(o)-induced intracellular acidification.
    Experimental physiology 09/2011; 96(12):1270-81. DOI:10.1113/expphysiol.2011.059444 · 2.87 Impact Factor
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    • "was used in place of HEPES buffer in the external solution for the experiments conducted at acidic pH [67]. High Five cells expressing TRPV1 were plated onto polylysinecoated glass coverslips. "
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    ABSTRACT: TRPV1 is a ligand-gated cation channel that is involved in acute thermal nociception and neurogenic inflammation. By using the GP67 signal peptide, high levels of full-length human TRPV1 was expressed in High Five insect cells using the baculovirus expression system. The functional activity of the expressed TRPV1 was confirmed by whole-cell ligand-gated ion flux recordings in the presence of capsaicin and low pH and via specific ligand binding to the isolated cellular membranes. Efficient solubilization and purification protocols have resulted in milligram amounts of detergent-solubilized channel at 80-90% purity after Ni2+ IMAC chromatography and size exclusion chromatography. Western blot analysis of amino and carboxyl terminal domains and MS of tryptic digestions of purified protein confirmed the presence of the full-length human TRPV1. Specific ligand binding experiments confirmed the protein integrity of the purified human TRPV1.
    Protein Expression and Purification 05/2009; 65(1):38-50. DOI:10.1016/j.pep.2008.12.006 · 1.51 Impact Factor
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    • "The pharmacological challenge of TRPV1 P Holzer differentiate between capsaicin and protons (Seabrook et al., 2002; Gavva et al., 2004, 2005a, b; Neelands et al., 2005). On the other hand, AMG8562 does not block heat-evoked activation of rat TRPV1 (Lehto et al., 2008). "
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    ABSTRACT: The transient receptor potential vanilloid-1 (TRPV1) cation channel is a receptor that is activated by heat (>42 degrees C), acidosis (pH<6) and a variety of chemicals among which capsaicin is the best known. With these properties, TRPV1 has emerged as a polymodal nocisensor of nociceptive afferent neurones, although some non-neuronal cells and neurones in the brain also express TRPV1. The activity of TRPV1 is controlled by a multitude of regulatory mechanisms that either cause sensitization or desensitization of the channel. As many proalgesic pathways converge on TRPV1 and this nocisensor is upregulated and sensitized by inflammation and injury, TRPV1 is thought to be a central transducer of hyperalgesia and a prime target for the pharmacological control of pain. As a consequence, TRPV1 agonists causing defunctionalization of sensory neurones and a large number of TRPV1 blockers have been developed, some of which are in clinical trials. A major drawback of many TRPV1 antagonists is their potential to cause hyperthermia, and their long-term use may carry further risks because TRPV1 has important physiological functions in the peripheral and central nervous system. The challenge, therefore, is to pharmacologically differentiate between the physiological and pathological implications of TRPV1. There are several possibilities to focus therapy specifically on those TRPV1 channels that contribute to disease processes. These approaches include (i) site-specific TRPV1 antagonists, (ii) modality-specific TRPV1 antagonists, (iii) uncompetitive TRPV1 (open channel) blockers, (iv) drugs interfering with TRPV1 sensitization, (v) drugs interfering with intracellular trafficking of TRPV1 and (vi) TRPV1 agonists for local administration.
    British Journal of Pharmacology 09/2008; 155(8):1145-62. DOI:10.1038/bjp.2008.351 · 4.99 Impact Factor
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