TRPV2 is critical for the maintenance of cardiac structure and function in mice

The heart has a dynamic compensatory mechanism for haemodynamic stress. However, the molecular details of how mechanical forces are transduced in the heart are unclear. Here we show that the transient receptor potential, vanilloid family type 2 (TRPV2) cation channel is critical for the maintenance of cardiac structure and function. Within 4 days of eliminating TRPV2 from hearts of the adult mice, cardiac function declines severely, with disorganization of the intercalated discs that support mechanical coupling with neighbouring myocytes and myocardial conduction defects. After 9 days, cell shortening and Ca2+ handling by single myocytes are impaired in TRPV2-deficient hearts. TRPV2-deficient neonatal cardiomyocytes form no intercalated discs and show no extracellular Ca2+-dependent intracellular Ca2+ increase and insulin-like growth factor (IGF-1) secretion in response to stretch stimulation. We further demonstrate that IGF-1 receptor/PI3K/Akt pathway signalling is significantly downregulated in TRPV2-deficient hearts, and that IGF-1 administration partially prevents chamber dilation and impairment in cardiac pump function in these hearts. Our results improve our understanding of the molecular processes underlying the maintenance of cardiac structure and function

TRPV3 in keratinocytes transmits temperature information to sensory neurons via ATP

Transient receptor potential V3 (TRPV3) and TRPV4 are heat-activated cation channels expressed in keratinocytes. It has been proposed that heat-activation of TRPV3 and/or TRPV4 in the skin may release diffusible molecules which would then activate termini of neighboring dorsal root ganglion (DRG) neurons. Here we show that adenosine triphosphate (ATP) is such a candidate molecule released from keratinocytes upon heating in the co-culture systems. Using TRPV1-deficient DRG neurons, we found that increase in cytosolic Ca(2+)-concentration in DRG neurons upon heating was observed only when neurons were co-cultured with keratinocytes, and this increase was blocked by P2 purinoreceptor antagonists, PPADS and suramin. In a co-culture of keratinocytes with HEK293 cells (transfected with P2X(2) cDNA to serve as a bio-sensor), we observed that heat-activated keratinocytes secretes ATP, and that ATP release is compromised in keratinocytes from TRPV3-deficient mice. This study provides evidence that ATP is a messenger molecule for mainly TRPV3-mediated thermotransduction in skin. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1007/s00424-009-0703-x) contains supplementary material, which is available to authorized users

TRPV1 and TRPV4 play pivotal roles in delayed onset muscle soreness.

Unaccustomed strenuous exercise that includes lengthening contraction (LC) often causes tenderness and movement related pain after some delay (delayed-onset muscle soreness, DOMS). We previously demonstrated that nerve growth factor (NGF) and glial cell line-derived neurotrophic factor (GDNF) are up-regulated in exercised muscle through up-regulation of cyclooxygenase (COX)-2, and they sensitized nociceptors resulting in mechanical hyperalgesia. There is also a study showing that transient receptor potential (TRP) ion channels are involved in DOMS. Here we examined whether and how TRPV1 and/or TRPV4 are involved in DOMS. We firstly evaluated a method to measure the mechanical withdrawal threshold of the deep tissues in wild-type (WT) mice with a modified Randall-Selitto apparatus. WT, TRPV1-/- and TRPV4-/- mice were then subjected to LC. Another group of mice received injection of murine NGF-2.5S or GDNF to the lateral gastrocnemius (LGC) muscle. Before and after these treatments the mechanical withdrawal threshold of LGC was evaluated. The change in expression of NGF, GDNF and COX-2 mRNA in the muscle was examined using real-time RT-PCR. In WT mice, mechanical hyperalgesia was observed 6-24 h after LC and 1-24 h after NGF and GDNF injection. LC induced mechanical hyperalgesia neither in TRPV1-/- nor in TRPV4-/- mice. NGF injection induced mechanical hyperalgesia in WT and TRPV4-/- mice but not in TRPV1-/- mice. GDNF injection induced mechanical hyperalgesia in WT but neither in TRPV1-/- nor in TRPV4-/- mice. Expression of NGF and COX-2 mRNA was significantly increased 3 h after LC in all genotypes. However, GDNF mRNA did not increase in TRPV4-/- mice. These results suggest that TRPV1 contributes to DOMS downstream (possibly at nociceptors) of NGF and GDNF, while TRPV4 is located downstream of GDNF and possibly also in the process of GDNF up-regulation

Vacuolar-ATPase-mediated muscle acidification caused muscular mechanical nociceptive hypersensitivity after chronic stress in rats, which involved extracellular matrix proteoglycan and ASIC3

Abstract Although widespread pain, such as fibromyalgia, is considered to have a central cause, peripheral input is important. We used a rat repeated cold stress (RCS) model with many characteristics common to fibromyalgia and studied the possible involvement of decreased muscle pH in muscle mechanical hyperalgesia. After a 5-day RCS, the muscle pH and the muscular mechanical withdrawal threshold (MMWT) decreased significantly. Subcutaneously injected specific inhibitor of vacuolar ATPase (V-ATPase), bafilomycin A1, reversed both changes almost completely. It also reversed the increased mechanical response of muscle thin-fibre afferents after RCS. These results show that V-ATPase activation caused muscle pH drop, which led to mechanical hypersensitivity after RCS. Since extracellular matrix proteoglycan and acid sensitive ion channels (TRPV1 and ASIC3) have been considered as possible mechanisms for sensitizing/activating nociceptors by protons, we investigated their involvement. Manipulating the extracellular matrix proteoglycan with chondroitin sulfate and chondroitinase ABC reversed the MMWT decrease after RCS, supporting the involvement of the extracellular mechanism. Inhibiting ASIC3, but not TRPV1, reversed the decreased MMWT after RCS, and ASIC3 mRNA and protein in the dorsal root ganglia were upregulated, indicating ASIC3 involvement. These findings suggest that extracellular mechanism and ASIC3 play essential roles in proton-induced mechanical hyperalgesia after RCS

NGF-β mRNA was up-regulated in all three genotypes.

<p>(<b>A</b>) Time course of NGF-β mRNA expression in LC-exercised LGC muscle in WT mice. (<b>B</b>) Up-regulation of NGF-β mRNA 3 h after LC in the muscle of three genotypes. Median and interquartile range (IQR). All values were normalized with β-actin mRNA. n = 3–8 for each group (shown in the parentheses under each column). * <i>p</i>≤0.05 compared with pre, and n.s. not different from WT (Kruskal-Wallis one-way analysis of variance on ranks test followed by the Dunn's test).</p

Measurement of muscular mechanical withdrawal threshold with Randall-Selitto apparatus in mice.

<p>(<b>A</b>) Schedule for testing effects of EMLA cream treatment on the withdrawal threshold. More than 12 h before these measurements, inflammation was induced by injecting carrageenan into the LGC muscle. VFT: von Frey hair test, RST: Randall-Sellito test. (<b>B</b>) Change in VFT threshold (tip diameter: 0.25 mm) by surface anesthesia. (i) Vehicle cream (n = 9) did not change the threshold. (ii) EMLA treatment (n = 10) significantly raised the threshold compared with before the treatment. Median and interquartile range (IQR) are shown. *** <i>p</i><0.001 for pre- and post-cream treatment comparison by Mann-Whitney test. Note that pre values are decreased ones after induction of inflammation (same in C). (<b>C</b>) Change after surface anesthesia by EMLA cream treatment in withdrawal threshold measured by RST with a self-made larger probe (tip diameter: 2.6 mm). Filled circles: EMLA treatment (n = 10), open square: vehicle cream treatment (n = 9). EMLA cream did not significantly change the threshold, the same as vehicle cream. Mean ± S.E.M. (n = 6–10 for each group). S.E.M.s are hardly seen because they are small.</p

Muscular mechanical hyperalgesia induced by lengthening contraction in mice.

<p>(<b>A</b>) Schema of lengthening contraction (LC) application to the lower hindleg flexors, mainly the lateral gastrocnemius (LGC) muscle. LC was induced by electrical stimulation through a pair of needle electrodes inserted near the tibial and sciatic nerves. The ankle joint was dorsi-flexed in synchrony with muscle contraction, and then returned to the starting position over a 3 s resting period. This cycle was repeated 300 times. (<b>B</b>) Change in withdrawal thresholds by RST in WT mice that received LC or sham (stretch only) exercise (n = 9 for each group, mean ± S.E.M.). Vertical axis: withdrawal threshold in mN, horizontal axis: time after exercise. There was a significant difference between the groups, and the threshold decreased 6 to 36 h after exercise in LC group, but not in sham group. ** <i>p</i><0.01, *** <i>p</i><0.001 compared with −1 day in LC group; # <i>p</i><0.05, ### <i>p</i><0.001 compared with sham group on each time point, two-way repeated measures ANOVA with Bonferroni t-test.</p

Muscular mechanical hyperalgesia did not develop after LC in TRPV1−/− and TRPV4−/− mice.

<p>(<b>A</b>) Change in the withdrawal thresholds after LC measured by RST in TRPV1−/− (crosses) and TRPV4−/− (open squares) mice. Vertical axis: difference in the threshold from −1 d in mN, horizontal axis: time after LC. (<b>B</b>) Changes in the mechanical hyperalgesia by intramuscular injection of HC-067047, a TRPV4 selective antagonist (100 mg/kg; crosses) or DMSO (open squares) in WT mice. Mean ± S.E.M. (n = 6–10 for each group). ** <i>p</i><0.01, *** <i>p</i><0.001 compared with −1 d in WT,+<i>p</i><0.05 compared with −1 d in TRPV4−/−, ## <i>p</i><0.01, ### <i>p</i><0.001 compared with 14 h in HC-067047 group; two-way repeated measures ANOVA with Bonferroni t-test.</p