Fig. 4. The pore-helix domain plays a critical role in camphor sensitivity of transient receptor potential vanilloid 1 (TRPV1) channel. Alignment of the pore-helix domain (627–640) for three distinct rat TRPV channels. An asterisk shows the amino acids inside the pore-helix domain at position 633 (A , a ). Alanine substitution of pore-helix threonine 633 (T633A) caused significant reduction in camphor-evoked responsiveness (A , b ). Quantification of maximal peak current density at −70 mV for camphor-evoked currents in wild type (WT) and in T633A mutant (B ). Currents evoked by 10 mM camphor (CMP10) for wild type, T633A mutant, and V538L mutant of rat TRPV1, and for human TRPV1 (hTRPV1), relative to maximal responses to 10 μM capsaicin (CAPS10), assessed as ratio CMP10/CAPS10 (C ). In B  and C  data represent mean ± SD. Number of cells is in parentheses. T633A mutant (squares ), but not the TRPV1-Δ15:Y627-C634 chimera (circles ), could be activated by depolarizing voltages. The TRPV1 chimera (Δ15-TRPV1: Y627-C634) lacked the stretch of 15 nonconserved residues between the turret and selectivity filter (T612-S626), and the pore helix (Y627-C634) was replaced with the counterpart from TRPV2, a camphor-insensitive homolog. Conductance-voltage (G–V ) relationships were obtained from steady-state whole cell currents measured at the end of voltage steps from −80 to +200 mV in increments of +20 mV (D ). TRPV1 chimera (Δ15:Y627-C634) is completely insensitive to camphor. Camphor neither induced any detectable currents nor potentiated its capsaicin-evoked responses. This construct was reported previously to uncouple proton activation from other TRPV1 activation stimuli and exhibit a slow onset and offset of capsaicin responses (E ). Control deletion mutant of TRPV1 that lacked 15 nonconserved residues between T612-S626 (situated before the pore helix) responded to camphor normally (F ). Camphor activates very poorly human TRPV1. The capsaicin-induced desensitized currents are still potentiated by camphor (G ). Camphor (CMP) induced potentiation of heat-induced currents in wild-type and N628G mutant of TRPV1. The heat-evoked currents in N628G were significantly smaller and less potentiated by 10 mM camphor. Data represent mean ± SD from four wild-type and for five mutant-expressing cells (H ).

Fig. 4. The pore-helix domain plays a critical role in camphor sensitivity of transient receptor potential vanilloid 1 (TRPV1) channel. Alignment of the pore-helix domain (627–640) for three distinct rat TRPV channels. An asterisk shows the amino acids inside the pore-helix domain at position 633 (A , a ). Alanine substitution of pore-helix threonine 633 (T633A) caused significant reduction in camphor-evoked responsiveness (A , b ). Quantification of maximal peak current density at −70 mV for camphor-evoked currents in wild type (WT) and in T633A mutant (B ). Currents evoked by 10 mM camphor (CMP10) for wild type, T633A mutant, and V538L mutant of rat TRPV1, and for human TRPV1 (hTRPV1), relative to maximal responses to 10 μM capsaicin (CAPS10), assessed as ratio CMP10/CAPS10 (C ). In B  and C  data represent mean ± SD. Number of cells is in parentheses. T633A mutant (squares ), but not the TRPV1-Δ15:Y627-C634 chimera (circles ), could be activated by depolarizing voltages. The TRPV1 chimera (Δ15-TRPV1: Y627-C634) lacked the stretch of 15 nonconserved residues between the turret and selectivity filter (T612-S626), and the pore helix (Y627-C634) was replaced with the counterpart from TRPV2, a camphor-insensitive homolog. Conductance-voltage (G–V ) relationships were obtained from steady-state whole cell currents measured at the end of voltage steps from −80 to +200 mV in increments of +20 mV (D ). TRPV1 chimera (Δ15:Y627-C634) is completely insensitive to camphor. Camphor neither induced any detectable currents nor potentiated its capsaicin-evoked responses. This construct was reported previously to uncouple proton activation from other TRPV1 activation stimuli and exhibit a slow onset and offset of capsaicin responses (E ). Control deletion mutant of TRPV1 that lacked 15 nonconserved residues between T612-S626 (situated before the pore helix) responded to camphor normally (F ). Camphor activates very poorly human TRPV1. The capsaicin-induced desensitized currents are still potentiated by camphor (G ). Camphor (CMP) induced potentiation of heat-induced currents in wild-type and N628G mutant of TRPV1. The heat-evoked currents in N628G were significantly smaller and less potentiated by 10 mM camphor. Data represent mean ± SD from four wild-type and for five mutant-expressing cells (H ).

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