Fig 2: FL potentiates TRP function in vitro and in vivo. a Traces of [Ca2+]i responses to repeated bath applications of capsaicin (Cap, 2.5 µM), a selective TRPV1 activator, or capsaicin combined with FL (0.56 nM) on [Ca2+]i levels measured by real-time Ca2+ cell imaging in cultured DRG neurons. Ratio F340/F380 is the ratio of fluorescence signals measured at 340 and 380 nm excitation wavelengths. b Effects of capsaicin (Cap, 2.5 µM) or FL at the indicated concentrations, either alone or in combinations on [Ca2+]i levels, measured at the fourth application of capsaicin as in a. Results are expressed as response amplitudes normalized to capsaicin alone (?F peak). c Effects of FL on capsaicin-induced increased [Ca2+]i levels, measured as in a, in DRG neurons from wild-type (WT) or Flt3KO mice. d Potentiation by FL (0.56 nM) of increase in [Ca2+]i levels induced by capsaicin (Cap, 2.5 µM), cinnamaldehyde, Cina (50 µM), and menthol (1 µM). e Traces of voltage-clamp whole-cell recording of capsaicin (Cap, 1 µM)-induced TRPV1 currents and their potentiation by FL (10 nM). f Quantification of FL effects measured as in e. g In vivo potentiation by FL of capsaicin-induced pain-related behaviors. Animals received either 4 repeated identical injections of capsaicin (15 ng) or a combination of Cap and FL (2 ng) followed by 3 repeated capsaicin injections in the paw, at the indicated times. Results are means ± s.e.m. of data from the number of cells indicated in columns (b–d, f) or data from 8 animals (g). Unpaired Student t-test. NS non-significant; *P < 0.05; **P < 0.01; ***P < 0.001

Fig 3: Immunoflourescence of TRPV1 from aortic ECs using a fluorescein-coupled secondary antibody. The cells were treated with buffer (control) (a) or with CS (30 µM, 30 min) (b and c). The image is a confocal section that shows the arrangement of the channel in the cell (green dots) and the nucleus (blue). Image was taken on 20X objective. (c–f) represent an increase or zoom of the figures for a better observation.

Fig 4: Expression of TRPV1 in the cardiac tissue from normotensive (N) and hypertensive (H) rats, in the absence and presence of capsaicin (CS), capsazepine (CZ), and capsazepine + capsaicin (CZ + CS). (n = 3–4 rats/group). By Duncan’s post hoc test. Data are presented as means ± SEM. (* p < 0.05 of N and H rats).

Fig 5: (a) eNOS and (b) TRPV1 expression in aortic tissue. Control (C) and CS, CZ, and CZ + CS treatments. The bars represent mean ± SEM of 4 animals per group. #p < 0.05; *p < 0.05 vs. control group. Representative western blot analysis is shown.