We have previously demonstrated the presence of TRPV1 in human platelets (Harper et al., 2009). The TRPV1 agonist, capsaicin, was shown to evoke dose-dependent rises in cytosolic calcium concentration ([Ca2+]c) and capsaicin-evoked responses were inhibited by the TRPV1 antagonists 5′-iodo-resiniferatoxin or AMG 9810. We also demonstrated the presence of TRPV1 by Western blotting, a result since independently confirmed by Savini et al (2010) using a different antibody. Further, we demonstrated that TRPV1 contributes to platelet calcium signals evoked by the physiological agonists ADP and thrombin (Harper et al., 2009). However, the endogenous activators of TRPV1 in platelets are unknown. A recent report has suggested that skeletal muscle hypertrophy involves the activation of TRPV1 by peroxynitrite (Ito et al., 2013). It has been suggested that platelets generate peroxynitrite and that this plays a role in the abnormal Ca2+ signalling observed in platelets from type II diabetic patients (Redondo et al., 2005). Here we have investigated whether peroxynitrite generates calcium signals in human platelets in a TRPV1-dependent manner. Platelets were isolated from blood obtained by venepuncture of healthy, drug-free volunteers under informed consent with local ethical committee approval. Platelet-rich plasma was prepared and platelets loaded with the calcium indicator Fura-2 as previously described (Harper et al., 2009). The cells were collected by centrifugation and resuspended in a Hepes-buffered saline as previously described except that bovine-serum albumin was omitted from the medium. Fura-2 fluorescence was recorded from 1.5 ml stirred aliquots of platelet suspension in a Cairn Research spectrofluorimeter with excitation at 340 and 380 nm and emission at 500 nm. 1 mM CaCl2 was added immediately before experiments and the Fura-2 340/380 nm ratio was calibrated in terms of [Ca2+]c using the method of Grynkiewicz et al (1985). Calcium signals were quantified as the integrals of the rise in [Ca2+]c above basal for 120 s after peroxyinitrite addition.The addition of peroxynitrite (180 µM, from Merck Millipore) evoked a rapid rise in [Ca2+]c of 1309 ± 340 nM above basal levels (n = 4) whereas the vehicle (4.7% NaOH) alone was without effect. Rises in [Ca2+]c evoked by peroxynitrite (180 µM) were inhibited in a dose-dependent manner by preincubation of the cells with 5′-iodo-resiniferatoxin or AMG 9810, with responses being reduced to 43.3 ± 5.0% or 35.6 ± 10.4% of control after preincubation with 30 µM 5′-iodo-resiniferatoxin or AMG 9810 respectively (both n= 4, Student’s paired t-test P < 0.05).These data are compatible with peroxynitrite being a potential endogenous activator of TRPV1 in human platelets.