Sodium channels contribute to microglia/macrophage activation and function in EAE and MS

Loss of axons is a major contributor to nonremitting deficits in the inflammatory demyelinating disease multiple sclerosis (MS). Based on biophysical studies showing that activity of axonal sodium channels can trigger axonal degeneration, recent studies have tested sodium channel-blocking drugs in experimental autoimmune encephalomyelitis (EAE), an animal model of MS, and have demonstrated a protective effect on axons. However, it is possible that, in addition to a direct effect on axons, sodium channel blockers may also interfere with inflammatory mechanisms. We therefore examined the novel hypothesis that sodium channels contribute to activation of microglia and macrophages in EAE and acute MS lesions. In this study, we demonstrate a robust increase of sodium channel Nav1.6 expression in activated microglia and macrophages in EAE and MS. We further demonstrate that treatment with the sodium channel blocker phenytoin ameliorates the inflammatory cell infiltrate in EAE by 75%. Supporting a role for sodium channels in microglial activation, we show that tetrodotoxin, a specific sodium channel blocker, reduces the phagocytic function of activated rat microglia by 40%. To further confirm a role of Nav1.6 in microglial activation, we examined the phagocytic capacity of microglia from med mice, which lack Nav1.6 channels, and show a 65% reduction in phagocytic capacity compared with microglia from wildtype mice. Our findings indicate that sodium channels are important for activation and phagocytosis of microglia and macrophages in EAE and MS and suggest that, in addition to a direct neuroprotective effect on axons, sodium channel blockade may ameliorate neuroinflammatory disorders via anti-inflammatory mechanisms.     

NOX2 drives M1-like microglial/macrophage activation and neurodegeneration following experimental traumatic brain injury

Following traumatic brain injury (TBI), activation of microglia and peripherally derived inflammatory macrophages occurs in association with tissue damage. This neuroinflammatory response may have beneficial or detrimental effects on neuronal survival, depending on the functional polarization of these cells along a continuum from M1-like to M2-like activation states. The mechanisms that regulate M1-like and M2-like activation after TBI are not well understood, but appear in part to reflect the redox state of the lesion microenvironment. NADPH oxidase (NOX2) is a critical enzyme system that generates reactive oxygen species in microglia/macrophages. After TBI, NOX2 is strongly up-regulated in M1-like, but not in M2-like polarized cells. Therefore, we hypothesized that NOX2 drives M1-like neuroinflammation and contributes to neurodegeneration and loss of neurological function after TBI. In the present studies we inhibited NOX2 activity using NOX2-knockout mice or the selective peptide inhibitor gp91ds-tat. We show that NOX2 is highly up-regulated in infiltrating macrophages after injury, and that NOX2 deficiency reduces markers of M1-like activation, limits tissue loss and neurodegeneration, and improves motor recovery after moderate-level control cortical injury (CCI). NOX2 deficiency also promotes M2-like activation after CCI, through increased IL-4Rα signaling in infiltrating macrophages, suggesting that NOX2 acts as a critical switch between M1- and M2-like activation states after TBI. Administration of gp91ds-tat to wild-type CCI mice starting at 24 h post-injury reduces deficits in cognitive function and increased M2-like activation in the hippocampus. Collectively, our data indicate that increased NOX2 activity after TBI drives M1-like activation that contributes to inflammatory-mediated neurodegeneration, and that inhibiting this pathway provides neuroprotection, in part by altering M1-/M2-like balance towards the M2-like neuroinflammatory response.     

帕金森病患者外周血Th17/Treg平衡及相关细胞因子水平测定及意义

目的探讨帕金森病(PD)患者外周血Th17/Treg平衡及相关细胞因子含量变化的意义。方法收集50例PD患者(PD组)及50例健康体检者(Control组),采集所有研究对象外周血。采用流式细胞仪检测外周血中Th17细胞及Treg细胞含量;采用ELISA检测血浆中细胞因子TGF-β、IL-6和IL-17水平。结果流式细胞计数结果显示,PD组的外周血Treg细胞和Th17细胞水平较Control组均显著增加(P均0.01),Treg细胞的增长程度明显大于大于Th17细胞,同时PD组Treg/Th17细胞比较Control组也明显增加(P均0.01)。ELISA检测结果显示,与Control组相比,PD组的血浆TGF-β1、IL-6及IL-17含量均有显著增加(P均0.01)。结论 PD患者的外周血Treg/Th17平衡存在失衡,提示PD病理过程可能与其外周血Treg/Th17失衡有关。     

Dopamine D1/D5 receptor signaling regulates synaptic cooperation and competition in hippocampal CA1 pyramidal neurons via sustained ERK1/2 activation

Synaptic cooperation and competition are important components of synaptic plasticity that tune synapses for the formation of associative long‐term plasticity, a cellular correlate of associative long‐term memory. We have recently reported that coincidental activation of weak synapses within the vicinity of potentiated synapses will alter the cooperative state of synapses to a competitive state thus leading to the slow decay of long‐term plasticity, but the molecular mechanism underlying this is still unknown. Here, using acute hippocampal slices of rats, we have examined how increasing extracellular dopamine concentrations interact and/or affect electrically induced long‐term potentiation (LTP) in the neighboring synapses. We demonstrate that D1/D5‐receptor‐mediated potentiation at the CA1 Schaffer collateral synapses differentially regulates synaptic co‐operation and competition. Further investigating the molecular players involved, we reveal an important role for extracellular signal‐regulated kinases‐1 and 2 (ERK1/2) as signal integrators and dose‐sensors. Interestingly, a sustained activation of ERK1/2 pathway seems to be involved in the differential regulation of synaptic associativity. The concentration‐dependent effects of the modulatory transmitter, as demonstrated for dopaminergic signaling in the present study, might offer additional computational power by fine tuning synaptic associativity processes for establishing long‐term associative memory in neural networks. © 2015 The Authors Hippocampus Published by Wiley Periodicals, Inc.     

The peripheral nociceptive actions of intravenously administered 5-HT in the rat requires dual activation of both 5-HT2 and 5-HT3 receptor subtypes

In 16-week-old Sprague-Dawley rats lightly anesthetized with pentobarbital, 5-HT (3–96 μg/kg, i.v.;n = 6) produced distinct pseudaffective responses and a dose-dependent (slope= 17.2 ± 6.8s/log₁₀dose) inhibition of the tail-flick (TF) reflex (ED₅₀ = 32.6 ± 9.2 μg/kg). In the same rats, a 1:1 combination of α-methyl 5-HT (a 5-HT₂ receptor selective agonist) and 2-methyl 5-HT (a 5-HT₃ receptor selective agonist) (3–192 μg/kg, i.v.), produced the same profile of pseudaffective responses and also resulted in a dose-dependent (slope= 34.0± 7.0s/log₂dose) inhibition of the TF reflex (ED₅₀ = 88.4 ± 20.5 μg/kg). In contrast, administration of α-methyl 5-HT (3–192 μg/kg, i.v.) or 2-methyl 5-HT (3–192 μg/kg, i.v.) alone did not produce any pseudaffective responses or any change in TF latency from baseline. In conscious 16-week-old male Sprague-Dawley rats, administration of 5-HT (48 μg/kg, i.v.;n = 5), or a 1:1 combination of α-methyl 5-HT and 2-methyl 5-HT (total dose= 120 μg/kg, i.v.;mn = 5), resulted in a passive avoidance behavior assessed in a step-down paradigm (slopes= 139.7 ± 58.2and154.9 ± 63.9s/trial, respectively), and the same profile of distinct pseudaffective responses exhibited by the lightly pentobarbital-anesthetized rats. However, administration of either α-methyl 5-HT (96 μg/kg, i.v.;n = 4) or 2-methyl 5-HT (96 μg/kg, i.v.;n = 4), while producing significant 5-HT receptor-mediated cardiovascular responses, produced a learned behavior not different from saline (0.25 ml, i.v.;n = 6) (slopes= 7.6 ± 2.5, 6.3 ± 1.8and7.4 ± 3.6s/trial, respectively). These results are consistent with the hypothesis that the peripheral nociceptive responses to i.v. 5-HT requires dual activation of 5-HT₂ and 5-HT₃ receptor subtypes.     

Effects of endogenous and exogenous D1/D5 dopamine receptor activation on LTP in ventral and dorsal CA1 hippocampal synapses

Hippocampus is importantly involved in dopamine‐dependent behaviors and dopamine is a significant modulator of synaptic plasticity in the hippocampus. Moreover, the dopaminergic innervation appears to be disproportionally segregated along the hippocampal longitudinal (dorsoventral) axis with unknown consequences for synaptic plasticity. In this study we examined the actions of endogenously released dopamine and the effects of exogenous D1/D5 dopamine receptor agonists on theta‐burst stimulation‐induced long‐term potentiation (LTP) of field excitatory synaptic potential (fEPSP) at Schaffer collateral‐CA1 synapses in slices from dorsal (DH) and ventral hippocampus (VH). Furthermore, we quantified D1 receptor mRNA and protein expression levels in DH and VH. We found that blockade of D1/D5 receptors by SCH 23390 (20 μM) significantly reduced the magnitude of LTP in both DH and VH similarly suggesting that dopamine endogenously released during TBS, presumably mimicking low activity of DA neurons, exerts a homogeneous modulation of LTP along the hippocampal long axis. Moderate to high concentrations of the selective partial D1/D5 receptor agonist SKF 38393 (50‐150 μM) did not significantly change LTP in either hippocampal segment. However, the full D1 receptor selective agonist SKF 82958 (10 μM) significantly enhanced LTP in VH but not DH. Furthermore, the expression of D1 receptor mRNA and protein was considerably higher in VH compared with DH. These results suggest that the dynamic range of D1/D5 receptor‐mediated dopamine effects on LTP may be higher in VH than DH and that VH may be specialized to acquire information about behaviorally relevant strong stimuli signaled by the dopamine system.     

Effects of adrenalectomy and type I or type II glucocorticoid receptor activation on 5-HT1A and 5-HT2 receptor binding and 5-HT transporter mRNA expression in rat brain

We evaluated the effects of adrenalectomy (ADX) and replacement with glucocorticoid receptor agonists on serotonin (5-HT) 5-HT_1A and 5-HT₂ receptor binding in rat brain. 5-HT_1A receptor binding was increased in the CA2–CA4 and the dentate gyrus of the hippocampus 1 week after ADX. This effect was prevented by the systemic administration of aldosterone (10 μg/μl/h) but not by RU28362 (10 μg/μl/h). No significant effect was observed on 5-HT₂ receptor binding in rat cortex. The expression of 5-HT transporter mRNA was unchanged in the raphe nucleus as measured by in situ hybridization.     

The effects of manipulation of presynaptic 5-HT nerve terminals on postsynaptic 5-HT1 and 5-HT2 binding sites of the rat brain

Summary The effects of long-term treatment of rats with alaproclate and amiflamine on the number and kinetics of 5-HT₁ and 5-HT₂ binding sites were investigated usingin vitro receptor binding techniques. Some other studies have reported down-regulatory effects of alaproclate and amiflamine on 5-HT₂ binding sites in certain regions of the rat forebrain, but no such effects could be detected in the present study. Induction of a high-affinity binding site for³H-5-HT after long-term antidepressant treatment, as has been reported elsewhere, was not obtained in the present study. The results are compared to the effects obtained by treatment of rats with para-chloroarnphetamine (PCA), which depletes the presynaptic neurons of monoamines. These different types of treatment do not cause any change in the binding properties of the specific 5-HT binding sites. It is thus concluded that such manipulations of the presynaptic 5-HT neurons do not affect the postsynaptic 5-HT₁ and 5-HT₂ binding sites.     

Effects of 8-OH-DPAT, a 5-HT1A receptor agonist, and DOI, a 5-HT2A/2C agonist, on monosynaptic transmission in spinalized rats

We investigated the effect of the 5-HT_1A receptor agonist (±)-8-hydroxy-2-(di-N-propylamino)tetralin (8-OH-DPAT) and the 5-HT_2A/2C receptor agonist (±)-1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane (DOI) on monosynaptic transmission in spinalized rats. 8-OH-DPAT significantly inhibited the excitation of α-motoneurons evoked by monosynaptic transmission without a direct effect on α-motoneuron excitation. DOI potentiated the excitation of α-motoneurons by both direct stimulation and monosynaptic transmission. These results indicate that activation of 5-HT_1A receptors inhibits monosynaptic transmission, whereas activation of 5-HT_2A/2C receptors enhances it.     

Sodium appetite elicited by low‐sodium diet is dependent on p44/42 mitogen‐activated protein kinase (extracellular signal‐regulated kinase 1/2) activation in the brain

Sodium appetite is regulated by several signalling molecules, among which angiotensin II (Ang II) serves as a key driver of robust salt intake by binding to Ang II type 1 receptors (AT1R) in several regions in the brain. The activation of these receptors recruits the mitogen‐activated protein kinase (MAPK) pathway, which has previously been linked to Ang II‐induced increases in sodium appetite. Thus, we addressed the involvement of MAPK signalling in the induction of sodium appetite after 4 days of low‐sodium diet consumption. An increase in extracellular signal‐regulated kinase (ERK) phosphorylation in the laminae terminalis and mediobasal hypothalamus was observed after low‐sodium diet consumption. This response was reduced by i.c.v. microinjection of an AT1R antagonist into the laminae terminalis but not the hypothalamus. This result indicates that low‐sodium diet consumption activates the MAPK pathway via Ang II/AT1R signalling on the laminae terminalis. On the other hand, activation of the MAPK pathway in the mediobasal hypothalamus after low‐sodium diet consumption appears to involve another extracellular mediator. We also evaluated whether a low‐sodium diet could increase the sensitivity for Ang II in the brain and activate the MAPK pathway. However, i.c.v. injection of Ang II increased ERK phosphorylation on the laminae terminalis and mediobasal hypothalamus; this increase achieved a response magnitude similar to those observed in both the normal and low‐sodium diet groups. These data indicate that low‐sodium diet consumption for 4 days is insufficient to change the ERK phosphorylation response to Ang II in the brain. To investigate whether the MAPK pathway is involved in sodium appetite after low‐sodium diet consumption, we performed i.c.v. microinjections of a MAPK pathway inhibitor (PD98059). PD98059 inhibited both saline and water intake after low‐sodium diet consumption. Thus, the MAPK pathway is involved in promoting the sodium appetite after low‐sodium diet consumption.     

The influence of 5-HT(2C) and MDR1 genetic polymorphisms on antipsychotic-induced weight gain in female schizophrenic patients

We investigated the relationships between functional genetic variants of the 5-HT(2C) receptor and multidrug-resistant protein (MDR1), coding for P-glycoprotein, and second generation antipsychotic (SDA)-induced weight gain among 108 female schizophrenic patients treated with olanzapine or risperidone for up to 4 months. No significant differences in -759C/T allelic and genotype variants of 5-HT(2C) were found between patients who gained more than 7% of their initial weight compared with those who gained less. Haplotype-based analysis of two MDR1 loci, exon 21 G2677T and exon 26 C3435T, revealed a slightly lower representation of the G2677/C3435 haplotype in the >/=7% group. In the subgroup of patients treated with risperidone, we found borderline overrepresentation of 2677T, significant overrepresentation of 3435T variant and borderline overrepresentation of 2677T/3435T haplotype the >/=7% group, whereas G2677/C3435 haplotype was found to be less represented in the >/=7% group. Our data indicate a nonsignificant role of 759C/T 5-HT(2C) in SDA-induced weight gain, and a stronger influence of the MDR1 G2677T and C3435T polymorphisms on risperidone-induced weight gain in female schizophrenic patients. 3435T and 2677T MDR1 variants, both associated with lower P-gp function, might predispose to higher risperidone accessibility to the brain that would lead to stronger effects, including weight gain.