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An external sodium ion binding site controls allosteric gating in TRPV1 channels.

Jara-Oseguera A, Bae C, Swartz KJ - Elife (2016)

Bottom Line: Here, we show that external sodium ions stabilize the TRPV1 channel in a closed state, such that removing the external ion leads to channel activation.The binding of a tarantula toxin to the external pore also exerts control over temperature-sensor activation, whereas binding of vanilloids influences temperature-sensitivity by largely affecting the open/closed equilibrium.Our results reveal a fundamental role of the external pore in the allosteric control of TRPV1 channel gating and provide essential constraints for understanding how these channels can be tuned by diverse stimuli.

View Article: PubMed Central - PubMed

Affiliation: Molecular Physiology and Biophysics Section, Porter Neuroscience Research Center, National Institute of Neurological Disorders and Stroke, National Institutes of Health, Bethesda, United States.

ABSTRACT
TRPV1 channels in sensory neurons are integrators of painful stimuli and heat, yet how they integrate diverse stimuli and sense temperature remains elusive. Here, we show that external sodium ions stabilize the TRPV1 channel in a closed state, such that removing the external ion leads to channel activation. In studying the underlying mechanism, we find that the temperature sensors in TRPV1 activate in two steps to favor opening, and that the binding of sodium to an extracellular site exerts allosteric control over temperature-sensor activation and opening of the pore. The binding of a tarantula toxin to the external pore also exerts control over temperature-sensor activation, whereas binding of vanilloids influences temperature-sensitivity by largely affecting the open/closed equilibrium. Our results reveal a fundamental role of the external pore in the allosteric control of TRPV1 channel gating and provide essential constraints for understanding how these channels can be tuned by diverse stimuli.

No MeSH data available.


Related in: MedlinePlus

Perfusion-mediated temperature control.(A) Schematic illustration of the perfusion-based temperature-control system used for rapid temperature jumps and I-V relations at low temperatures. Solutions kept in elevated reservoirs (for gravity-driven flow) were passed through glass capillary spirals immersed in water baths at different temperatures, and recordings were performed in a small-volume (200–500 µL) chamber during constant perfusion. Temperature was measured with a thermistor located very close to the pipette tip. Separate perfusion lines were used for each solution (e.g. one for 130 mM external Na+, shown in grey in the figure, and one for external NMDG+, shown in yellow). (B) Representative current time courses obtained during rapid perfusion-induced temperature changes showing both temperature (red traces, top panels) and current (open symbols, bottom panel) recorded in the whole-cell configuration at -90 mV (triangles) and +90 mV (circles). Horizontal thick lines denote changes in extracellular solution composition, and the red dotted lines indicate the zero-current level.DOI:http://dx.doi.org/10.7554/eLife.13356.018
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fig5s3: Perfusion-mediated temperature control.(A) Schematic illustration of the perfusion-based temperature-control system used for rapid temperature jumps and I-V relations at low temperatures. Solutions kept in elevated reservoirs (for gravity-driven flow) were passed through glass capillary spirals immersed in water baths at different temperatures, and recordings were performed in a small-volume (200–500 µL) chamber during constant perfusion. Temperature was measured with a thermistor located very close to the pipette tip. Separate perfusion lines were used for each solution (e.g. one for 130 mM external Na+, shown in grey in the figure, and one for external NMDG+, shown in yellow). (B) Representative current time courses obtained during rapid perfusion-induced temperature changes showing both temperature (red traces, top panels) and current (open symbols, bottom panel) recorded in the whole-cell configuration at -90 mV (triangles) and +90 mV (circles). Horizontal thick lines denote changes in extracellular solution composition, and the red dotted lines indicate the zero-current level.DOI:http://dx.doi.org/10.7554/eLife.13356.018

Mentions: (A) Representative whole-cell current family obtained as in Figure 4A in the absence of external Na+. The temperature vs time plot is shown on the right panel. Dotted lines denote the zero-current level. (B) Mean I-T relations in the absence (obtained from data as in (A), mean ± SEM, n = 7) and presence (same data as in Figure 4B) of external Na+ at +90 mV (circles) and -90 mV (triangles). The I-T relation in purple is from an experiment in the presence of 130 mM external Na+ with pronounced temperature-dependent inactivation at T < 50°C. The purple and yellow arrows denote the approximate onset of inactivation for data with and without external Na+, respectively. The dotted red line denotes the zero-current level. All relations are normalized to peak-current values at +90 mV. (C, left) Mean Po-T relations (+90 mV) in the presence (grey) and absence (yellow) of external Na+. Po values at room temperature for scaling Po-T relations on an absolute Po-scale were estimated from macroscopic I-V relations and noise analysis as described in Methods and Figure 5—figure supplement 2. The dotted vertical line delimits the lower range of experimentally accessible temperatures. Po-T relations from individual cells in the absence of external Na+ are shown in Figure 6—figure supplement 1A. (C, right) ΔHapp from fits of Equation 1 (see Materials and methods) to Po-T relations at +90 mV from individual cells (circles) and their mean ± SEM (squares). The mean ΔHapp for the fits to data with external Na+ at T > 25°C is shown as an open square (mean ± SEM, n = 10). The mean ΔHapp from fits to data in external Na+ at T < 25°C is shown as a closed square (mean ± SEM, n = 4). (D) Normalized I-T relations (+90 mV) from (B) plotted on a log-scale (small circles) with superimposed I-T relations obtained from rapid temperature-jumps from 8°C to higher temperatures (large circles, mean ± SEM, n = 3–8, see Materials and methods and Figure 5—figure supplement 3). The blue bars denote the increased inactivation observed in the I-T relation in the absence of external Na+ obtained from slow temperature ramps relative to that obtained with rapid temperature-jumps. (E) Schematic representation of the essential features on a log-scale of Po-T relations at +90 mV in the presence (grey) or absence (yellow) of external Na+. The dashed lines denote plateaus in which the Po does not visibly change with temperature. Arrows denote portions of the relations in which Po steeply increases with temperature. Monoexponential fits of Equation 1 correspond on a log-scale to straight lines with slope ~ΔHapp.


An external sodium ion binding site controls allosteric gating in TRPV1 channels.

Jara-Oseguera A, Bae C, Swartz KJ - Elife (2016)

Perfusion-mediated temperature control.(A) Schematic illustration of the perfusion-based temperature-control system used for rapid temperature jumps and I-V relations at low temperatures. Solutions kept in elevated reservoirs (for gravity-driven flow) were passed through glass capillary spirals immersed in water baths at different temperatures, and recordings were performed in a small-volume (200–500 µL) chamber during constant perfusion. Temperature was measured with a thermistor located very close to the pipette tip. Separate perfusion lines were used for each solution (e.g. one for 130 mM external Na+, shown in grey in the figure, and one for external NMDG+, shown in yellow). (B) Representative current time courses obtained during rapid perfusion-induced temperature changes showing both temperature (red traces, top panels) and current (open symbols, bottom panel) recorded in the whole-cell configuration at -90 mV (triangles) and +90 mV (circles). Horizontal thick lines denote changes in extracellular solution composition, and the red dotted lines indicate the zero-current level.DOI:http://dx.doi.org/10.7554/eLife.13356.018
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4764576&req=5

fig5s3: Perfusion-mediated temperature control.(A) Schematic illustration of the perfusion-based temperature-control system used for rapid temperature jumps and I-V relations at low temperatures. Solutions kept in elevated reservoirs (for gravity-driven flow) were passed through glass capillary spirals immersed in water baths at different temperatures, and recordings were performed in a small-volume (200–500 µL) chamber during constant perfusion. Temperature was measured with a thermistor located very close to the pipette tip. Separate perfusion lines were used for each solution (e.g. one for 130 mM external Na+, shown in grey in the figure, and one for external NMDG+, shown in yellow). (B) Representative current time courses obtained during rapid perfusion-induced temperature changes showing both temperature (red traces, top panels) and current (open symbols, bottom panel) recorded in the whole-cell configuration at -90 mV (triangles) and +90 mV (circles). Horizontal thick lines denote changes in extracellular solution composition, and the red dotted lines indicate the zero-current level.DOI:http://dx.doi.org/10.7554/eLife.13356.018
Mentions: (A) Representative whole-cell current family obtained as in Figure 4A in the absence of external Na+. The temperature vs time plot is shown on the right panel. Dotted lines denote the zero-current level. (B) Mean I-T relations in the absence (obtained from data as in (A), mean ± SEM, n = 7) and presence (same data as in Figure 4B) of external Na+ at +90 mV (circles) and -90 mV (triangles). The I-T relation in purple is from an experiment in the presence of 130 mM external Na+ with pronounced temperature-dependent inactivation at T < 50°C. The purple and yellow arrows denote the approximate onset of inactivation for data with and without external Na+, respectively. The dotted red line denotes the zero-current level. All relations are normalized to peak-current values at +90 mV. (C, left) Mean Po-T relations (+90 mV) in the presence (grey) and absence (yellow) of external Na+. Po values at room temperature for scaling Po-T relations on an absolute Po-scale were estimated from macroscopic I-V relations and noise analysis as described in Methods and Figure 5—figure supplement 2. The dotted vertical line delimits the lower range of experimentally accessible temperatures. Po-T relations from individual cells in the absence of external Na+ are shown in Figure 6—figure supplement 1A. (C, right) ΔHapp from fits of Equation 1 (see Materials and methods) to Po-T relations at +90 mV from individual cells (circles) and their mean ± SEM (squares). The mean ΔHapp for the fits to data with external Na+ at T > 25°C is shown as an open square (mean ± SEM, n = 10). The mean ΔHapp from fits to data in external Na+ at T < 25°C is shown as a closed square (mean ± SEM, n = 4). (D) Normalized I-T relations (+90 mV) from (B) plotted on a log-scale (small circles) with superimposed I-T relations obtained from rapid temperature-jumps from 8°C to higher temperatures (large circles, mean ± SEM, n = 3–8, see Materials and methods and Figure 5—figure supplement 3). The blue bars denote the increased inactivation observed in the I-T relation in the absence of external Na+ obtained from slow temperature ramps relative to that obtained with rapid temperature-jumps. (E) Schematic representation of the essential features on a log-scale of Po-T relations at +90 mV in the presence (grey) or absence (yellow) of external Na+. The dashed lines denote plateaus in which the Po does not visibly change with temperature. Arrows denote portions of the relations in which Po steeply increases with temperature. Monoexponential fits of Equation 1 correspond on a log-scale to straight lines with slope ~ΔHapp.

Bottom Line: Here, we show that external sodium ions stabilize the TRPV1 channel in a closed state, such that removing the external ion leads to channel activation.The binding of a tarantula toxin to the external pore also exerts control over temperature-sensor activation, whereas binding of vanilloids influences temperature-sensitivity by largely affecting the open/closed equilibrium.Our results reveal a fundamental role of the external pore in the allosteric control of TRPV1 channel gating and provide essential constraints for understanding how these channels can be tuned by diverse stimuli.

View Article: PubMed Central - PubMed

Affiliation: Molecular Physiology and Biophysics Section, Porter Neuroscience Research Center, National Institute of Neurological Disorders and Stroke, National Institutes of Health, Bethesda, United States.

ABSTRACT
TRPV1 channels in sensory neurons are integrators of painful stimuli and heat, yet how they integrate diverse stimuli and sense temperature remains elusive. Here, we show that external sodium ions stabilize the TRPV1 channel in a closed state, such that removing the external ion leads to channel activation. In studying the underlying mechanism, we find that the temperature sensors in TRPV1 activate in two steps to favor opening, and that the binding of sodium to an extracellular site exerts allosteric control over temperature-sensor activation and opening of the pore. The binding of a tarantula toxin to the external pore also exerts control over temperature-sensor activation, whereas binding of vanilloids influences temperature-sensitivity by largely affecting the open/closed equilibrium. Our results reveal a fundamental role of the external pore in the allosteric control of TRPV1 channel gating and provide essential constraints for understanding how these channels can be tuned by diverse stimuli.

No MeSH data available.


Related in: MedlinePlus