PMC

Modulation of the piperine-activated TRPV1 current by acid. (a) A typical recording trace showing the dramatic potentiation of a capsaicin-(1 μM, black bar)-mediated TRPV1 response by extracellular acidification from pH 7.3 to pH 6.5 (grey bar). In these experiments, pH 6.5 was selected for study as it is a subagonist pH versus TRPV1 (data not shown). pH 6.5 potentiated the capsaicin response by 284±32% (n=8). (b) A similar protocol was employed to study the effects of acidification on the piperine response. Switching from pH 7.30 to pH 6.5 potentiated the piperine-(30 μM gated response, white bar) by 230±20% (n=11) (c) Pooled data showing that capsaicin and piperine are similarly modulated by acid (pH 6.5). The data are expressed as a percentage of the mean current recorded in response to the agonist at pH 7.3.

Piperine produces a greater degree of desensitisation than capsaicin. The effects of piperine and capsaicin on TRPV1 desensitisation and tachyphylaxis were compared using a repeated applications protocol. Representative traces showing the typical profile of responses obtained during eight repeated applications of (a) 1 μM capsaicin (closed bar) or (b) 100 μM piperine (open bar) are shown. (b) Agonists were applied for 20 s with intervening washout periods of 1 min (indicated by the gaps). (c) The pooled data from the experiments described in (a and b) are given. Agonist responses are expressed as a percentage of the initial current for each of eight repeated applications of capsaicin (n=7) or piperine (n=8). ^*P<0.05; ^**P<0.01 versus the corresponding capsaicin-mediated response.

Piperine activates human TRPV1. (a) The chemical structures of the vanilloid capsaicin and the related nonvanilloid congener piperine, which lacks the 4-hydroxy-3-methoxybenzyl or ‘vanillyl' group of capsaicin, are shown. (b) Capsaicin (1 μM; a concentration close to Emax, see ) activated robust inward currents in hTRPV1.HEK293 cells demonstrating the clear expression of human TRPV1 in these cells. In the same cells, piperine (100 μM) typically activated larger currents which had a distinct kinetic profile due to the effects of desensitisation. The traces shown are from one experiment and are typical of five others. Mean data for these experiments were: I_piperine=1600±360 pA and I_capsaicin=610±140 pA; P<0.01, n=6, Student's paired test). Capsaicin and piperine were without effect on parental wild-type HEK293 cells (data not shown).

Rectification and pharmacological properties of piperine-gated currents. (a) The current–voltage relationship for piperine- and capsaicin-activated currents was established using a voltage-ramp protocol (see top inset) ranging from −70 to +70 mV. A series of voltage-ramps (15 × −70 mV to +70 mV at 0.14 mV ms⁻¹) was used to capture data prior to, during and following the recovery of the agonist-induced response. The net agonist-evoked current was calculated by subtracting the mean background current from the agonist-evoked current obtained from ramps, which coincided with the steady-state phase of the response. The example shown is for a piperine-evoked response where the agonist was applied for the duration indicated by the bar. Similar experiments were conducted for capsaicin. (b) A plot of the current–voltage relationship obtained for piperine (30 μM dotted line; n=5) and capsaicin (1 μM, solid line; n=5) from pooled data generated from experiments similar to those in (a). Currents were normalised to the steady-state current observed at −70 mV and then averaged across cells. Occasional error bars (±1s.e.m.) are shown at 10 mV intervals. Piperine responses show clear outward rectification (I_+70 mV/I_−70 mV=25±4 compared to 24±5 for capsaicin; P=0.87, unpaired Student's t-test) and exhibit a reversal potential close to zero (E_rev=0.0±0.4 mV compared to −1.0±0.8 mV for capsaicin; P=0.29, unpaired Student's t-test). (c) Piperine-evoked currents (clear bar; 30 μM) were inhibited by coapplication of capsazepine 10 μM or ruthenium red 10 μM (grey bars). (d) Pooled data showing the % block of the piperine-evoked current expressed by capsazepine, (n=3) and ruthenium red, (n=4).

Piperine exhibits greater efficacy than capsaicin at the human TRPV1. (a) The concentration-response relationship for capsaicin (0.01–10 μM) and piperine (10–100 μM) are shown. These normalised data were generated by measuring the net currents evoked in response to a test concentration of agonist and are expressed as a percentage of a preceding 1 μM capsaicin control response recorded in the same cell. Data are expressed as the mean±s.e.m., where n=3–10 individual cells. The EC₅₀ for capsaicin was 292±54 nM (Hill coefficient of 1.2±0.2; n=3–10 per concentration). The concentration–response profile for piperine clearly indicates the less potent nature of this compound with an EC₅₀ of 37.9 μM±1.9 (Hill coefficient 3.7±0.5; n=5–9 per concentration) indicating a greater efficacy and degree of cooperativity for piperine than for capsaicin. (b) A representative trace from an experiment designed to quantify the relative difference in efficacy between a 1 μM capsaicin and 100 μM piperine activated response (n=4). A TRPV1 response was first-evoked by capsaicin (black bar) before subsequent addition of piperine. Upon switching to the solution containing piperine the peak response was significantly increased and a greater degree of receptor desensitisation was evident. (c) Mean data from the experiments described in (b) showing a 3.9±0.4 fold increase in TRPV1 current by addition of piperine. Control experiments in which the solution was switched from 1 μM capsaicin to a second identical 1 μM capsaicin solution yielded no change in peak current.