Knockdown of NADPH oxidase 4 reduces mitochondrial oxidative stress and neuronal pyroptosis following intracerebral hemorrhage.

Intracerebral hemorrhage is often accompanied by oxidative stress induced by reactive oxygen species, which causes abnormal mitochondrial function and secondary reactive oxygen species generation. This creates a vicious cycle leading to reactive oxygen species accumulation, resulting in progression of the pathological process. Therefore, breaking the cycle to inhibit reactive oxygen species accumulation is critical for reducing neuronal death after intracerebral hemorrhage. Our previous study found that increased expression of nicotinamide adenine dinucleotide phosphate oxidase 4 (NADPH oxidase 4, NOX4) led to neuronal apoptosis and damage to the blood-brain barrier after intracerebral hemorrhage. The purpose of this study was to investigate the role of NOX4 in the circle involving the neuronal tolerance to oxidative stress, mitochondrial reactive oxygen species and modes of neuronal death other than apoptosis after intracerebral hemorrhage. We found that NOX4 knockdown by adeno-associated virus (AAV-NOX4) in rats enhanced neuronal tolerance to oxidative stress, enabling them to better resist the oxidative stress caused by intracerebral hemorrhage. Knockdown of NOX4 also reduced the production of reactive oxygen species in the mitochondria, relieved mitochondrial damage, prevented secondary reactive oxygen species accumulation, reduced neuronal pyroptosis and contributed to relieving secondary brain injury after intracerebral hemorrhage in rats. Finally, we used a mitochondria-targeted superoxide dismutase mimetic to explore the relationship between reactive oxygen species and NOX4. The mitochondria-targeted superoxide dismutase mimetic inhibited the expression of NOX4 and neuronal pyroptosis, which is similar to the effect of AAV-NOX4. This indicates that NOX4 is likely to be an important target for inhibiting mitochondrial reactive oxygen species production, and NOX4 inhibitors can be used to alleviate oxidative stress response induced by intracerebral hemorrhage.

Monitored anesthesia care and asleep-awake-asleep techniques combined with multiple monitoring for resection of gliomas in eloquent brain areas: a retrospective analysis of 225 patients.

BACKGROUND: Awake craniotomy (AC) has become gold standard in surgical resection of gliomas located in eloquent areas. The conscious sedation techniques in AC include both monitored anesthesia care (MAC) and asleep-awake-asleep (AAA). The choice of optimal anesthetic method depends on the preferences of the surgical team (mainly anesthesiologist and neurosurgeon). The aim of this study was to compare the difference in physiological and blood gas data, dosage of different drugs, the probability of switching to endotracheal intubation, and extent of tumor resection and dysfunction after operation between AAA and MAC anesthetic management for resection of gliomas in eloquent brain areas. METHODS: Two-hundred and twenty-five patients with super-tentorial tumor located in eloquent areas underwent AC from 2009 to 2021 in Xijing Hospital. Forty-one patients underwent AAA technique, and the rest one-hundred eighty-four patients underwent MAC technique. Anesthetic management, dosage of different drugs, intraoperative complications, postoperative outcomes, adverse events, extent of resection and motor, and sensory and language dysfunction after operation were compared between MAC and AAA. RESULT: There was no significant difference in gender, KPS score, MMSE score, glioma grade, type, and growth site between the patients in the two groups, except the older age of patients in MAC group than that in AAA group. During the whole process of operation, there were greater pulse pressure difference (P = 0.046), shorter operation time (P = 0.039), less dosage of remifentanil (P = 0.000), more dosage of dexmedetomidine (P = 0.013), more use of antiemetics (81%, P = 0.0067), lower use of vasoactive agent (45.1%, P = 0.010), and lower probability of conversion to general anesthesia (GA, P = 0.027) in MAC group than that in AAA group. Blood gas analysis showed that PetCO2 (P = 0.000), Glu concentration (P = 0.000), and PaCO2 (P = 0.000) were higher, but SPO2 (P = 0.002) and PaO2 (P = 0.000) were lower in MAC group than that in AAA group. In the postoperative recovery stage, compared with that of AAA group, the probability of dysfunction in MAC group at 1, 3, 5, and 7 days after operation was lower, which were 27.8% vs 53.6% (P = 0.003), 31% vs 68.3% (P = 0.000), 28.8% vs 63.4% (P = 0.000), and 25.6% vs 58.5% (P = 0.000), respectively. CONCLUSION: Compared with AAA, it seems that MAC has more advantages in the management for resection of gliomas in eloquent brain areas, and MAC combined with multiple monitoring such as cerebral cortical mapping, neuronavigation, and ultrasonic detection is worthy of popularization for the resection of gliomas in eloquent brain areas.

Brain natriuretic peptide modulates the delayed rectifier outward K(+) current and promotes the proliferation of mouse Schwann cells.

Brain natriuretic peptide (BNP) may act as a neuromodulator via its associated receptors (natriuretic peptide receptors, NPRs) in the central nervous system (CNS), but few studies have reported its activity in the peripheral nervous system (PNS). In this study, we observed that BNP increased the tetraethylammonium chloride (TEA)-sensitive delayed rectifier outward potassium current (I(K)) in mouse Schwann cells (SCs) using whole-cell recording techniques. At concentrations of 1-100 nM, BNP reversibly activated I(K) in a dose-dependent manner, with modulating its steady-state activation and inactivation properties. The effect of BNP on I(K) was abolished by preincubation with the specific antagonist of NPR-A, and could not be mimicked by application of NPR-C agonist. These results were supported by immunocytochemical findings indicating that NPR-A was expressed in SCs. The application of 8-Br-guanosine 3',5'-monophosphate (8-Br-cGMP) mimicked the effect of BNP on I(K), but BNP was unable to further increase I(K) after the application of cyclic guanosine monophosphate (cGMP). Genistein blocked I(K) and also completely eliminated the effects of BNP and cGMP on I(K). The selective K(V)2.1 subunit blocker, Jingzhaotoxin-III (JZTX-III), reduced I(K) amplitude by 30%, but did not abolish the increase effect of BNP on I(K) amplitude. In addition, BNP significantly stimulated SCs proliferation and this effect could be partly inhibited by TEA. Together these results suggest that BNP modulated I(K) probably via cGMP- and tyrosine kinase-dependent pathways by activation of NPR-A. This effect of BNP on I(K) in SCs might partly explain its effect on cell proliferation.

Bradykinin inhibits the transient outward K+ current in mouse Schwann cells via the cAMP/PKA pathway.

Bradykinin (BK) is an endogenous peptide with diverse biological actions and is considered to be an important mediator of the inflammatory response in both the peripheral and the central nervous systems. BK has attracted recent interest as a potential mediator of K(+) conductance, Cl(-) channels, and Ca(2+)-activated K(+) channels. However, few reports have associated BK with the voltage-gated K(+) current. In this study, we demonstrated that BK suppressed the transient outward potassium current (I(A)) in mouse Schwann cells using whole cell recording techniques. At a concentration of 0.1 muM to 5 muM, BK reversibly inhibited I(A) in a dose-dependent manner with the modulation of steady-state activation and inactivation properties. The effect of BK on I(A) current was abolished after preincubation with a B(2) receptor antagonist but could not be eliminated by B(1) receptor antagonist. Intracellular application of GTP-gammaS induced an irreversible decrease in I(A), and the inhibition of G(s) using NF449 provoked a gradual augmentation in I(A) and eliminated the BK-induced effect on I(A,) while the G(i)/(o) antagonist NF023 did not. The application of forskolin or dibutyryl-cAMP mimicked the inhibitory effect of BK on I(A) and abolished the BK-induced effect on I(A). H-89, an inhibitor of PKA, augmented I(A) amplitude and completely eliminated the BK-induced inhibitory effect on I(A). In contrast, activation of PKC by PMA augmented I(A) amplitude. A cAMP assay revealed that BK significantly increased intracellular cAMP level. It is therefore concluded that BK inhibits the I(A) current in Schwann cells by cAMP/PKA-dependent pathways via activation of the B(2) receptor.

PLC-dependent intracellular Ca2+ release was associated with C6-ceramide-induced inhibition of Na+ current in rat granule cells.

In this report, the effects of C(6)-ceramide on the voltage-gated inward Na(+) currents (I(Na)), two types of main K(+) current [outward rectifier delayed K(+) current (I(K)) and outward transient K(+) current (I(A))], and cell death in cultured rat cerebellar granule cells were investigated. At concentrations of 0.01-100 microM, ceramide produced a dose-dependent and reversible inhibition of I(Na) without alteration of the steady-state activation and inactivation properties. Treatment with C(2)-ceramide caused a similar inhibitory effect on I(Na). However, dihydro-C(6)-ceramide failed to modulate I(Na). The effect of C(6)-ceramide on I(Na) was abolished by intracellular infusion of the Ca(2+)-chelating agent, 1,2-bis (2-aminophenoxy) ethane-N, N, N9, N9-tetraacetic acid, but was mimicked by application of caffeine. Blocking the release of Ca(2+) from the sarcoplasmic reticulum with ryanodine receptor blocker induced a gradual increase in I(Na) amplitude and eliminated the effect of ceramide on I(Na). In contrast, the blocker of the inositol 1,4,5-trisphosphate-sensitive Ca(2+) receptor did not affect the action of C(6)-ceramide. Intracellular application of GTPgammaS also induced a gradual decrease in I(Na) amplitude, while GDPbetaS eliminated the effect of C(6)-ceramide on I(Na). Furthermore, the C(6)-ceramide effect on I(Na) was abolished after application of the phospholipase C (PLC) blockers and was greatly reduced by the calmodulin inhibitors. Fluorescence staining showed that C(6)-ceramide decreased cell viability and blocking I(Na) by tetrodotoxin did not mimic the effect of C(6)-ceramide, and inhibiting intracellular Ca(2+) release by dantrolene could not decrease the C(6)-ceramide-induced cell death. We therefore suggest that increased PLC-dependent Ca(2+) release through the ryanodine-sensitive Ca(2+) receptor may be responsible for the C(6)-ceramide-induced inhibition of I(Na), which does not seem to be associated with C(6)-ceramide-induced granule neuron death.

Mefenamic acid bi-directionally modulates the transient outward K+ current in rat cerebellar granule cells.

The effect of non-steroidal anti-inflammatory drugs (NSAIDs) on ion channels has been widely studied in several cell models, but less is known about their modulatory mechanisms. In this report, the effect of mefenamic acid on voltage-activated transient outward K(+) current (I(A)) in cultured rat cerebellar granule cells was investigated. At a concentration of 5 microM to 100 microM, mefenamic acid reversibly inhibited I(A) in a dose-dependent manner. However, mefenamic acid at a concentration of 1 microM significantly increased the amplitude of I(A) to 113+/-1.5% of the control. At more than 10 microM, mefenamic acid inhibited the amplitude of I(A) without any effect on activation or inactivation. In addition, a higher concentration of mefenamic acid induced a significant acceleration of recovery from inactivation with an increase of the peak amplitude elicited by the second test pulse. Intracellular application of mefenamic acid could significantly increase the amplitude of I(A), but had no effect on the inhibition induced by extracellular mefenamic acid, implying that mefenamic acid may exert its effect from both inside and outside the ion channel. Furthermore, the activation of current induced by intracellular application of mefenamic acid was mimicked by other cyclooxygenase inhibitors and arachidonic acid. Our data demonstrate that mefenamic acid is able to bi-directionally modulate I(A) channels in neurons at different concentrations and by different methods of application, and two different mechanisms may be involved.

Delayed rectifier outward K+ current mediates the migration of rat cerebellar granule cells stimulated by melatonin.

Melatonin (MT) may work as a neuromodulator through the associated MT receptors in the central nervous system. Previously, our studies have shown that MT increased the I(K) current via a G protein-related pathway. In the present study, patch-clamp whole-cell recording, transwell migration assays and organotypic cerebellar slice cultures were used to examine the effect of MT on granule cell migration. MT increased the I(K) current amplitude and migration of granule cells. Meanwhile, TEA, the I(K) channel blocker, decreased the I(K) current and slowed the migration of granule cells. Furthermore, the effects of MT on the I(K) current and cell migration were not abolished by pre-incubation with P7791, a specific antagonist of MT(3)R, but were eliminated by the application of the MT(2)R antagonists K185 and 4-P-PDOT. I(K) current and cell migration were decreased by the application of dibutyryl cyclic AMP (dbcAMP), which was in contrast to the MT effect on the I(K) current and cell migration. Incubation with dbcAMP essentially blocked the MT-induced increasing effect. Moreover, incubation of isolated cell cultures in the MT-containing medium also decreased the cAMP immunoreactivity in the granule cells. It is concluded, therefore, that I(K) current, downstream of a cAMP transduction pathway, mediates the migration of rat cerebellar granule cells stimulated by MT.

Flufenamic acid bi-directionally modulates the transient outward K(+) current in rat cerebellar granule cells.

In this report, the effect of flufenamic acid on voltage-activated transient outward K(+) current (I(A)) in cultured rat cerebellar granule cells was investigated. At a concentration of 20 microM to 1 mM, flufenamic acid reversibly inhibited I(A) in a dose-dependent manner. However, flufenamic acid at a concentration of 0.1 to 10 microM significantly increased the current amplitude of I(A). In addition to the current amplitude of I(A), a higher concentration of flufenamic acid had a significant effect on the kinetic parameters of the steady-state activation and inactivation process, suggesting that the binding affinity of flufenamic acid to I(A) channels may be state-dependent. Silencing the K(v)4.2, K(v)4.3, and K(v)1.1 genes of I(A) channels using small interfering RNA did not change the inhibitory effect of flufenamic on I(A), indicating that flufenamic acid did not act specifically on any of the subunits of the I(A)-channel protein. Intracellular application of flufenamic acid could significantly increase the I(A) amplitude but did not alter the inhibited effect induced by extracellular application of flufenamic acid, implying that flufenamic acid may exert its effect from both the inside and outside sites of the channel. Furthermore, the activation of current induced by intracellular application of flufenamic acid could mimic other cyclooxygenase inhibitors and arachidonic acid. Our data are the first that demonstrate how flufenamic acid is able to bidirectionally modulate I(A) channels in neurons at different concentrations and by different methods of application and that two different mechanisms may be involved.

The non-steroidal anti-inflammatory drug, diclofenac, inhibits Na(+) current in rat myoblasts.

The inhibitory effect of diclofenac, a non-steroidal anti-inflammatory drug (NSAID), on the voltage-gated inward Na+ current (I(Na)) in cultured rat myoblasts was investigated using the whole-cell voltage-clamp technique. At concentrations of 10 nM-100 microM, diclofenac produced a dose-dependent and reversible inhibition of I(Na) with an IC50 of 8.51 microM, without modulating the fast activation and inactivation process. The inhibitory effect of diclofenac took place at resting channels and increased with more depolarizing holding potential. In addition to inhibiting the Na+ current amplitude, diclofenac significantly modulated the steady-state inactivation properties of the Na+ channels, but did not alter the steady-state activation. The steady-state inactivation curve was significantly shifted towards the hyperpolarizing potential in the presence of diclofenac. Furthermore, diclofenac treatment resulted in a fairly slow recovery from inactivation of the Na+ channel. The inhibitory effect of diclofenac was enhanced by repetitive pulses and was inflected by changing frequency; the blocking effect at higher frequency was significantly greater than at lower frequency. Both intracellular and extracellular application of diclofenac could inhibit I(Na), indicating that diclofenac may exert its channel inhibitory action both inside and outside the channel sites. Our data directly demonstrate that diclofenac can inhibit the inward Na+ channels in rat myoblasts. Some different inhibitory mechanisms from that in neuronal Na+ channels are discussed.

Diclofenac, a nonsteroidal anti-inflammatory drug, activates the transient outward K+ current in rat cerebellar granule cells.

Diclofenac, a nonsteroidal anti-inflammatory drug (NSAID), has been widely investigated in terms of its pharmacological action, but less is known about its direct effect on ion channels. Here, the effect of diclofenac on voltage-dependent transient outward K+ currents (I(A)) in cultured rat cerebellar granule cells was investigated using the whole-cell voltage-clamp technique. At concentrations of 10(-5)-10(-3) M, diclofenac reversibly increased the I(A) amplitude in a dose-dependent manner and significantly modulated the steady-state inactivation properties of the I(A) channels, but did not alter the steady-state activation properties. Furthermore, diclofenac treatment resulted in a slightly accelerated recovery from I(A) channel inactivation. Intracellular application of diclofenac could mimic the effects induced by extracellular application, although once the intracellular response reached a plateau, extracellular application of diclofenac could induce further increases in the current. These observations indicate that diclofenac might exert its effects on the channel protein at both the inner and outer sides of the cell membrane. Our data provide the first evidence that diclofenac is able to activate transient outward potassium channels in neurons. Although further work will be necessary to define the exact mechanism of diclofenac-induced I(A) channel activation, this study provides evidence that the nonsteroidal anti-inflammatory drug, diclofenac, may play a novel neuronal role that is worthy of future study.