Correction: Functional human 3D microvascular networks on a chip to study the procoagulant effects of ambient fine particulate matter.

[This corrects the article DOI: 10.1039/C7RA11357A.].

Decreased calcium-activated potassium channels by hypoxia causes abnormal firing in the spontaneous firing medial vestibular nuclei neurons.

Vertebrobasilar insufficiency (VBI) presents complex varied clinical symptoms, including vertigo and hearing loss. Little is known, however, about how Ca(2+)-activated K(+) channel attributes to the medial vestibular nucleus (MVN) neural activity in VBI. To address this issue, we performed whole-cell patch clamp and quantitative polymerase chain reaction (qPCR) to examine the effects of hypoxia on neural activity and the changes of the large conductance Ca(2+) activated K(+) channels (BKCa channels) in the MVN neurons in brain slices of male C57BL/6 mice. Brief hypoxic stimuli of the brain slices containing MVN were administrated by switching the normoxic artificial cerebrospinal fluid (ACSF) equilibrated with 21% O2/5% CO2 to hypoxic ACSF equilibrated with 5% O2/5% CO2 (balance N2). 3-min hypoxia caused a depolarization in the resting membrane potential (RM) in 8/11 non-spontaneous firing MVN neurons. 60/72 spontaneous firing MVN neurons showed a dramatic increase in firing frequency and a depolarization in the RM following brief hypoxia. The amplitude of the afterhyperpolarization (AHPA) was significantly decreased in both type A and type B spontaneous firing MVN neurons. Hypoxia-induced firing response was alleviated by pretreatment with NS1619, a selective BKCa activator. Furthermore, brief hypoxia caused a decrease in the amplitude of iberiotoxin-sensitive outward currents and mRNA level of BKCa in MVN neurons. These results suggest that BKCa channels protect against abnormal MVN neuronal activity induced by hypoxia, and might be a key target for treatment of vertigo and hearing loss in VBI.

Complex regulation of capsaicin on intracellular second messengers by calcium dependent and independent mechanisms in primary sensory neurons.

Intracellular second messengers play an important role in capsaicin- and analogous-induced sensitization and desensitization in pain. Fluorescence Ca²âº imaging, enzyme immunoassay and PKC assay kit were used to determine a novel mechanism of different Ca²âº dependency in the signal transduction of capsaicin-induced desensitization. On the average, capsaicin increased cAMP, cGMP concentration and SP release in bell-shaped concentration-dependent manner, with the maximal responses at concentrations around 1 µM, suggesting acute desensitization of TRPV1 receptor activation. Capsaicin-induced intracellular Ca²âº concentration ([Ca²âº](i)) increase depended on extracellular Ca²âº influx as an initial trigger. The Ca²âº influx by capsaicin increased PKC activation and SP release. These increases were completely abolished in Ca²âº-free solution, suggesting that the modulation of capsaicin on PKC and SP are Ca²âº-dependent. Interestingly, the maximal cAMP increase by TRPV1 activation was not blocked Ca²âº removal, suggesting at least in part a Ca²âº-independent pathway is involved. Further study showed that cAMP increase was totally abolished by G-protein and adenylate cyclase (AC) antagonist, suggesting a G-protein-dependent pathway in cAMP increase. However, SP release was blocked by inhibiting PKC, but not G-protein or AC, suggesting a G-protein independent pathway in SP release. These results suggest that both Ca²âº-dependent and independent mechanisms are involved in the regulation of capsaicin on second messengers systems, which could be a novel mechanism underlying distinct desensitization of capsaicin and might provide additional opportunities in the development of effective analgesics in pain treatment.

Some subtypes of endocannabinoid/endovanilloid receptors mediate docosahexaenoic acid-induced enhanced spatial memory in rats.

ω-3 polyunsaturated fatty acid docosahexaenoic acid (DHA) enhances cognitive functions; however, the underlying molecular mechanism remains unclear. Compelling evidence suggests that the endocannabinoid/endovanilloid systems play a pivotal role in regulating cognitive function. Thus, to correlate the effect of DHA on cognitive performance with the expression of endocannabinoid and endovanilloid receptors, we supplemented the diet of rats with DHA and performed in vitro experiments that focused on the endocannabinoid/endovanilloid receptors. We found that in vivo supplementation with an appropriate dose of DHA (150 or 300mg/kg/d) significantly improved learning and memory but that a higher intake (600mg/kg/d) increased the risk of memory impairment. In addition, we found that some subtypes of endocannabinoid/endovanilloid receptors (cannabinoid [CB] and transient receptor potential vanilloid [TRPV] receptors) were regulated in vitro by different concentrations of DHA in primary hippocampal neuron culture medium. Real-time polymerase chain reaction and western blot analysis showed that expression of both CB1 and TRPV1 was upregulated in a dose-dependent manner and reached a maximum level at 30µmol/L (CB1) and 60µmol/L (TRPV1) DHA. However, TRPV2 expression was downregulated in a dose-dependent fashion, and the peak of TRPV2 suppression was observed at 60µmol/L. The dose-dependent effects of DHA on the expression of these receptors were well correlated with DHA's effect on spatial memory. Meanwhile, CB2, TRPV3, and TRPV4 expressions were not altered at diverse concentrations of DHA. We concluded that some subtypes of endocannabinoid/endovanilloid receptors might be involved in enhanced spatial memory induced by DHA supplementation.

Effects of WIN55,212-2 on voltage-gated sodium channels in trigeminal ganglion neurons of rats.

OBJECTIVES: This study was carried out to investigate the effects of WIN55,212-2, a potential cannabinoid receptor agonist, on voltage-gated sodium currents I(Na) in cultured trigeminal ganglion neurons of rats, and to investigate whether the anti-nociceptive effects of cannabinoid receptor subtype 1 (CB1) were produced through its modulation on I(Na). METHODS: Whole cell patch clamp techniques were used to record I(Na) before and after WIN55,212-2 was perfused in cultured trigeminal ganglion neurons of rats. RESULTS: WIN55,212-2 (0.01 micromol/l) could enhance I(Na) slightly by 11.5 +/- 4.7% (n=7, p<0.05), and this effect could not be blocked by AM251, the CB1 receptor antagonist. However, WIN55,212-2 could inhibit I(Na) in concentration dependent manner at concentrations from 0.1 to 100 micromol/l. The inhibitory rates were 17.4 +/- 6.0, 22.5 +/- 7.8, 43.9 +/- 9.4 and 73.9 +/- 6.7% respectively by 0.1, 1, 10, 100 micromol/l WIN55,212-2, and the EC(50) was 17.8 micromol/l (n=7, p<0.05 or p<0.01). This inhibitory effect could be blocked partly by 1 micromol/l AM251 (n=7, p<0.05). WIN55,212-2 (0.01 micromol/l) shifted the active curve of I(Na) leftward slightly (n=7, p<0.05), but had no effect on its stable inactive curve (n=7, p>0.05). WIN55,212-2 (10 micromol/l) did not affect the active and stable inactive curves of I(Na) (n=7, p>0.05). CONCLUSION: WIN55,212-2 had bidirectional (two phases) effects on I(Na) in trigeminal ganglion neurons. It might act on different receptors, and the CB1 receptor participated in its modulation on I(Na).

Cannabinoids inhibit ATP-activated currents in rat trigeminal ganglionic neurons.

The present study aimed to investigate whether cannabinoids could modulate the response mediated by ATP receptor (P2X purinoceptor). Whole-cell patch-clamp recording was performed on cultured rat trigeminal ganglionic (TG) neurons. The majority of TG neurons were sensitive to ATP (67/75, 89.33%). Extracellular pretreatment with WIN55212-2, a cannabinoid receptor 1 (CB1 receptor) agonist, reduced ATP-activated current (I(ATP)) significantly. This inhibitory effect was concentration-dependent and was blocked by AM281, a specific CB1 receptor antagonist. Pretreatment with WIN55212-2 at 1×10(-13), 1×10(-12), 1×10(-11), 1×10(-10), 1×10(-9) and 1×10(-8) mol/L reduced I(ATP) (induced by 1×10(-4) mol/L ATP) by (8.14±3.14)%, (20.11±2.72)%, (46.62±3.51)%, (72.16±5.64)%, (80.21±2.80)% and (80.59±3.55)%, respectively. The concentration-response curves for I(ATP) pretreated with and without WIN55212-2 showed that WIN55212-2 shifted the curve downward, and decreased the maximal amplitude of I(ATP) by (58.02±4.21)%. But the threshold value and EC(50) (1.15×10(-4) mol/L vs 1.27×10(-4) mol/L) remained unchanged. The inhibition of I(ATP) by WIN55212-2 was reversed by AM281, suggesting that the inhibition was mediated via the CB1 receptor. Pretreatment with forskolin [an agonist of adenylyl cyclase (AC)] or 8-Br-cAMP reversed the inhibition of I(ATP) by WIN55212-2. These results suggest that the inhibitory effect of cannabinoids on I(ATP) is mediated via the CB1 receptors, that lead to inhibition of the AC-cAMP-PKA signaling pathway.

[Effects of capsaicin on IA and IK in cultured trigeminal ganglion neurons of rat].

AIM: To investigate the effect of capsaicin on IA and IK in cultured rat trigeminal ganglion (TG) neurons. METHODS: Whole-cell patch clamp technique was used to record the IA and IK before and after capsaicin perfusion at different concentrations. RESULTS: In capsaicin-sensitive (CS) neurons, capsaicin was shown to selectively inhibit IA in dose-dependent manner, the IC50 was 0.99 micromol x L(-1). Yet capsaicin showed no inhibitory effect on IK, capsaicin (10 micromol x L(-1)) only slightly inhibited IK by 13.2%. In capsaicin-insensitive (CIS) neurons, capsaicin (1 micromol x L(-1)) showed no significant inhibitory effect on IA and IK, capsaicin (10 micromol x L(-1)) only slightly inhibited IA and IK by 16.8% and 15.3%, respectively. Neither 1 micromol x L(-1) nor 10 micromol x L(-1) capsaicin showed effect on the G-V curve of IA and IK. CONCLUSION: Capsaicin was found to selectively inhibit the IA current in CS neurons, and this effect may contribute to hyperalgesia when capsaicin was first used.