Ultrashort nanosecond electric pulses activate a conductance in bovine adrenal chromaffin cells that involves cation entry through TRPC and NALCN channels

3区 · 生物学

Article

作者: Gould, Thomas W ; Leblanc, Normand ; Pierce, Sophia ; Craviso, Gale L ; Yang, Lisha

In whole-cell voltage clamped bovine adrenal chromaffin cells maintained at a holding potential of -70 mV, a single 5 ns, 5 MV/m pulse elicited an inward current carried mainly by Na⁺ that displayed inward rectification and a reversal potential near -3 mV, a voltage consistent with a non-selective cation current. The broad-spectrum inhibitors of transient receptor potential (TRP) channels, La³⁺ (10 μM), Gd³⁺ (10 μM), SKF-96365 (50 μM) and 2-aminoethoxydiphenyl borane (2-APB; 100 μM), inhibited the current similarly by ∼72%, ∼83%, ∼68% and ∼76%, respectively. Depleting membrane cholesterol with methyl-β-cyclodextrin (MβCD; 1-6 mg/ml) or inhibiting phosphatidylinositol 4,5-bisphosphate (PIP₂) synthesis with wortmannin (20 and 40 μM) produced a similar level of inhibition on the NEP-induced conductance as the broad spectrum TRP channel inhibitors. Moreover, no additive inhibitory effect was detected by combining MβCD (3 mg/ml), wortmannin (20 μM) and La³⁺ (10 μM), suggesting that each agent targeted different levels of the same pathway to exert a full effect. RT-PCR experiments revealed robust expression at the mRNA level of TRPC4, TRPC5 and TRPM7 channels for which specific blockers were available. Whereas the TRPM7 blocker FTY720 had no effect, the TRPC4/5 channel inhibitor M084 (20 μM) blocked the conductance by ∼50%, indicating that TRPC4 and/or TRPC5 channel(s) may be partially involved in mediating the NEP-induced current. CP-96345 (20 μM), a specific blocker of the sodium leak current channel (NALCN), also reduced the NEP-induced current. The inhibition was ∼30% and additive to that caused by the TRPC4/5 blocker M084. RT-PCR experiments confirmed the expression of this channel at the mRNA level. Taken as a whole, these data provide evidence that a large fraction of the current evoked by a 5 ns pulse in adrenal chromaffin cells may be carried by both TRPC4/5 channels and the NALCN channel. Understanding the biophysical properties of the NEP-elicited conductance in a neural-type cell will be extremely valuable for the future development of NEP stimulation approaches for neuromodulation.

2020-05-01·Biochemical and biophysical research communications3区 · 生物学

Tannic acid, a vasodilator present in wines and beverages, stimulates Ca2+ influx via TRP channels in bEND.3 endothelial cells

3区 · 生物学

Article

作者: Shiao, Lian-Ru ; Tsai, Tien-Yao ; Wong, Kar-Lok ; Leung, Yuk-Man ; Leong, Iat-Lon ; Chan, Paul ; Shao, Li

Tannic acid (TA) is a polyphenol compound present in wines and many beverages. Although previous works have shown that TA could cause vasodilation in an endothelial cell (EC)-dependent manner, there is hitherto no report showing whether TA could raise EC cytosolic Ca²⁺ concentration. In this work we examined the effects of TA on cytosolic Ca²⁺ of mouse brain bEND.3 EC. TA (1-30 μM) caused a slow elevation in cytosolic Ca²⁺ level in a concentration-dependent manner. At 30 μM, TA triggered Ca²⁺ influx without causing intracellular Ca²⁺ release. TA-triggered Ca²⁺ influx was suppressed by Ni²⁺ (a non-specific Ca²⁺ channel blocker), ruthenium red and SKF 96365 (non-specific TRP channel blockers), CBA (a selective TRPM4 inhibitor) and M 084 (a selective TRPC4/C5 blocker). However, TA-triggered Ca²⁺ influx pathway was not permeable to Mn²⁺. Our results suggest TA activated TRP channels, possibly TRPM4 and TRPC4/C5, to promote influx of Ca²⁺.

2019-06-01·Bone2区 · 医学

Changes in Dkk-1, sclerostin, and RANKL serum levels following discontinuation of long-term denosumab treatment in postmenopausal women

2区 · 医学

Article

作者: Fassio, A ; Gatti, D ; Benini, C ; Lippolis, I ; Vantaggiato, E ; Orsolini, G ; Giollo, A ; Saag, K G ; Adami, G ; Idolazzi, L ; Viapiana, O ; Rossini, M

PURPOSE:

The positive effects of denosumab (DMAb) on bone mineral density (BMD) are quickly reversible after its discontinuation. We investigated whether this rebound was associated with dysregulation of the Wnt canonical pathway and/or by the increase in the receptor-activator of nuclear factor-kappa B ligand (RANKL) serum levels.

METHODS:

The study included patients (n = 15) with postmenopausal osteoporosis to whom DMAb was administered for 78 months and then discontinued. We collected BMD data at baseline/month 0 (M0), M60, M84 (6 months after last DMAb administration, coinciding when the next DMAb dose would typically be due), and after 3 and 12 months of follow-up (FU-M3 and FU-M12, respectively). Serum C-terminal telopeptide of type 1 collagen (CTX-I), Dickkopf-1 (Dkk-1), and sclerostin were measured at M0, M60, M84, FU-M3, and FU-M12. Serum N-terminal propeptide of type 1 procollagen (PINP) and RANKL were dosed at M60, M84, FU-M3, and FU-12.

RESULTS:

We found a significant decrease in the T-score at all sites at FU-M12, when compared to M84 (-0.51 ± 0.91 at the lumbar spine; -0.72 ± 0.33 at the total hip; and -0.42 ± 0.27 at the femoral neck, p < 0.05). After DMAb discontinuation (M84 vs FU M12) CTX-I, PINP increased already at FU-M3 (+0.921 ± 0.482 ng/mL, +126.60 ± 30.36 ng/mL, respectively, p < 0.01), RANKL increased at FU-M12 (+0.041 ± 0.062 ng/mL, p < 0.05), while Dkk-1 and sclerostin decreased at FU-M12 (-10.90 ± 11.80 and - 13.00 ± 10.52 pmol/L, respectively, p < 0.01). No changes in BMD or any of the markers were found between M60 and M84.

CONCLUSIONS:

RANKL serum levels progressively increased after discontinuation of long-term DMAb while Dkk-1 and sclerostin serum levels decreased. The increase in RANKL serum levels supports the hypothesis of a sudden loss of inhibition of the resting osteoclast line after DMAb clearance, with a hyperactivation of these cells. Our results suggest that the changes in serum Wnt inhibitors after DMAb suspension might represent a mere feedback response to the increased bone turnover.

100 项与 M-084 相关的药物交易

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