Estrogen receptor beta treats Alzheimer’s disease

In vitro studies have shown that estrogen receptor β can attenuate the cytotoxic effect of amyloid β protein on PC12 cells through the Akt pathway without estrogen stimulation. In this study, we aimed to observe the effect of estrogen receptor β in Alzheimer’s disease rat models established by intraventricular injection of amyloid β protein. Estrogen receptor β lentiviral particles delivered via intraventricular injection increased Akt content in the hippocampus, decreased interleukin-1β mRNA, tumor necrosis factor α mRNA and amyloid β protein levels in the hippocampus, and improved the learning and memory capacities in Alzheimer’s disease rats. Estrogen receptor β short hairpin RNA lentiviral particles delivered via intraventricular injection had none of the above impacts on Alzheimer’s disease rats. These experimental findings indicate that estrogen receptor β, independent from estrogen, can reduce inflammatory reactions and amyloid β deposition in the hippocampus of Alzheimer’s disease rats, and improve learning and memory capacities. This effect may be mediated through activation of the Akt pathway.     

Involvement of the Wnt signaling pathway and cell apoptosis in the rat hippocampus following cerebral ischemia/reperfusion injury

We investigated the role of the Wnt signaling pathway in cerebral ischemia/reperfusion injury by examining β-catenin and glycogen synthase kinase-3β protein expression in the rat hippocampal CA1 region following acute cerebral ischemia/reperfusion. Our results demonstrate that cell apoptosis increases in the CA1 region following ischemia/reperfusion. In addition, β-catenin and glycogen synthase kinase-3β protein expression gradually increases, peaking at 48 hours following reperfusion. Dickkopf-1 administration, after cerebral ischemia/reperfusion injury, results in decreased cell apoptosis, and β-catenin and glycogen synthase kinase-3β expression, in the CA1 region. This suggests that β-catenin and glycogen synthase kinase-3β, both components of the Wnt signaling pathway, participate in cell apoptosis following cerebral ischemia/reperfusion injury.     

ATP‐induced IL‐1β secretion is selectively impaired in microglia as compared to hematopoietic macrophages

Under stressful conditions nucleotides are released from dying cells into the extracellular space, where they can bind to purinergic P2X and P2Y receptors. High concentrations of extracellular ATP in particular induce P2X7‐mediated signaling, which leads to inflammasome activation. This in turn leads to the processing and secretion of pro‐inflammatory cytokines, like interleukin (IL)−1β. During neurodegenerative diseases, innate immune responses are shaped by microglia and we have previously identified microglia‐specific features of inflammasome‐mediated responses. Here, we compared ATP‐induced IL‐1β secretion in primary rhesus macaque microglia and bone marrow‐derived macrophages (BMDM). We assessed the full expression profile of P2 receptors and characterized the induction and modulation of IL‐1β secretion by extracellular nucleotides. Microglia secreted significantly lower levels of IL‐1β in response to ATP when compared to BMDM. We demonstrate that this is not due to differences in sensitivity, kinetics or expression of ATP‐processing enzymes, but rather to differences in purinergic receptor expression levels and usage. Using a combined approach of purinergic receptor agonists and antagonists, we demonstrate that ATP‐induced IL‐1β secretion in BMDM was fully dependent on P2X7 signaling, whereas in microglia multiple purinergic receptors were involved, including P2X7 and P2X4. These cell type‐specific features of conserved innate immune responses may reflect adaptations to the vulnerable CNS microenvironment. GLIA 2016;64:2231–2246     

Suppression of P2X3 receptor‐mediated currents by the activation of α2A‐adrenergic receptors in rat dorsal root ganglion neurons

AimsThe α₂‐adrenergic receptor (α₂‐AR) agonists have been shown to be effective in the treatment of various pain. For example, dexmedetomidine (DEX), a selective α_2A‐AR agonist, can be used for peripheral analgesia. However, it is not yet fully elucidated for the precise molecular mechanisms. P2X3 receptor is a major receptor processing nociceptive information in primary sensory neurons. Herein, we show that a functional interaction of α_2A‐ARs and P2X3 receptors in dorsal root ganglia (DRG) neurons could contribute to peripheral analgesia of DEX.MethodsElectrophysiological recordings were carried out on rat DRG neurons, and nociceptive behavior was quantified in rats.ResultsThe activation of α_2A‐ARs by DEX suppressed P2X3 receptor‐mediated and α,β‐methylene‐ATP (α,β‐meATP)‐evoked inward currents in a concentration‐dependent and voltage‐independent manner. Pre‐application of DEX shifted the α,β‐meATP concentration‐response curve downwards, with a decrease of 50.43 ± 4.75% in the maximal current response of P2X3 receptors to α,β‐meATP in the presence of DEX. Suppression of α,β‐meATP‐evoked currents by DEX was blocked by the α_2A‐AR antagonist BRL44408 and prevented by intracellular application of the G_i/o protein inhibitor pertussis toxin, the adenylate cyclase activator forskolin, and the cAMP analog 8‐Br‐cAMP. DEX also suppressed α,β‐meATP‐evoked action potentials through α_2A‐ARs in rat DRG neurons. Finally, the activation of peripheral α_2A‐ARs by DEX had an analgesic effect on the α,β‐meATP‐induced nociception.ConclusionsThese results suggested that activation of α_2A‐ARs by DEX suppressed P2X3 receptor‐mediated electrophysiological and behavioral activity via a G_i/o proteins and cAMP signaling pathway, which was a novel potential mechanism underlying analgesia of peripheral α_2A‐AR agonists.     

Serial expression of hypoxia inducible factor-1α and neuronal apoptosis in hippocampus of rats with chronic ischemic brain

Objective

The purpose of this study is to investigate serial changes of hypoxia-inducible factor 1α (HIF-1α), as a key regulator of hypoxic ischemia, and apoptosis of hippocampus induced by bilateral carotid arteries occlusion (BCAO) in rats.

Methods

Adult male Wistar rats were subjected to the permanent BCAO. The time points studied were 1, 2, 4, 8, and 12 weeks after occlusions, with n=6 animals subjected to BCAO, and n=2 to sham operation at each time point, and brains were fixed by intracardiac perfusion fixation with 4% neutral-buffered praraformaldehyde for brain section preparation. Immunohistochemistry (IHC), western blot and terminal uridine deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay were performed to evaluate HIF-1α expression and apoptosis.

Results

In IHC and western blot, HIF-1α levels were found to reach the peak at the 2nd week in the hippocampus, while apoptotic neurons, in TUNEL assay, were maximal at the 4th week in the hippocampus, especially in the cornu ammonis 1 (CA1) region. HIF-1α levels and apoptosis were found to fluctuate during the time course.

Conclusion

This study showed that BCAO induces acute ischemic responses for about 4 weeks then chronic ischemia in the hippocampus. These in vivo data are the first to show the temporal sequence of apoptosis and HIF-1α expression.     

Unlocking mechanisms in interleukin-1β-induced changes in hippocampal neurogenesis—a role for GSK-3β and TLX

Glycogen synthase kinase-3β (GSK-3β) and the orphan nuclear receptor tailless homolog (TLX) are key regulators of hippocampal neurogenesis, which has been reported to be dysregulated in both neurodegenerative and psychiatric disorders. Inflammation is also implicated in the neuropathology of these disorders because of increased levels of the pro-inflammatory cytokine interleukin-1β (IL-1β) in the brain. At elevated levels, IL-1β signaling through the IL-1 receptor type 1 has been shown to be detrimental to hippocampal neurogenesis. TLX is required to maintain neural stem/progenitor cells (NSPCs) in an undifferentiated state and is involved in NSPC fate determination, while GSK-3β negatively regulates Wnt signaling, a vital pathway promoting neurogenesis. This study shows that GSK-3β inhibition using a small-molecule inhibitor and the mood stabilizer lithium restores the IL-1β-induced decrease in NSPC proliferation and neuronal differentiation of embryonic rat hippocampal NSPCs to control levels. The IL-1β-induced effect on NSPCs is paralleled by a decrease in TLX expression that can be prevented by GSK-3β inhibition. The present results suggest that GSK-3β ameliorates the anti-proliferative and pro-gliogenic effects of IL-1β, and that TLX is vulnerable to inflammatory insult. Strategies to reduce GSK-3β activity or to increase TLX expression may facilitate the restoration of hippocampal neurogenesis in neuroinflammatory conditions where neurogenesis is impaired.     

Hypoxia‐independent mechanisms of HIF‐1α expression in astrocytes after ischemic preconditioning

We recently demonstrated that ischemic tolerance was dependent on astrocytes, for which HIF‐1α had an essential role. The mild ischemia (preconditioning; PC) increased HIF‐1α in a biphasic pattern, that is, a quick and transient increase in neurons, followed by a slow and sustained increase in astrocytes. However, mechanisms underlying such temporal difference in HIF‐1α increase remain totally unknown. Here, we show that unlike a hypoxia‐dependent mechanism in neurons, astrocytes increase HIF‐1α via a novel hypoxia‐independent but P2X7‐dependent mechanism. Using a middle cerebral artery occlusion (MCAO) model of mice, we found that the PC (a 15‐min MCAO period)‐evoked increase in HIF‐1α in neurons was quick and transient (from 1 to 3 days after PC), but that in astrocytes was slow‐onset and long‐lasting (from 3 days to at least 2 weeks after PC). The neuronal HIF‐1α increase was dependent on inhibition of PHD2, an oxygen‐dependent HIF‐1α degrading enzyme, whereas astrocytic one was independent of PHD2. Astrocytes even do not possess this enzyme. Instead, they produced a sustained increase in P2X7 receptors, activation of which resulted in HIF‐1α increase. The hypoxia‐independent but P2X7‐receptor‐dependent mechanism could allow astrocytes to cause long‐lasting HIF‐1α expression, thereby leading to induction of ischemic tolerance efficiently. GLIA 2017;65:523–530     

Amyloid beta-peptide worsens cognitive impairment following cerebral ischemia-reperfusion injury

Amyloid β-peptide,a major component of senile plaques in Alzheimer’s disease,has been implicated in neuronal cell death and cognitive impairment.Recently,studies have shown that the pathogenesis of cerebral ischemia is closely linked with Alzheimer’s disease.In this study,a rat model of global cerebral ischemia-reperfusion injury was established via occlusion of four arteries;meanwhile,fibrillar amyloidβ-peptide was injected into the rat lateral ventricle.The Morris water maze test and histological staining revealed that administration of amyloidβ-peptide could further aggravate impairments to learning and memory and neuronal cell death in the hippocampus of rats subjected to cerebral ischemia-reperfusion injury.Western blot showed that phosphorylation of tau protein and the activity of glycogen synthase kinase 3βwere significantly stronger in cerebral ischemia-reperfusion injury rats subjected to amyloid β-peptide administration than those undergoing cerebral ischemia-reperfusion or amyloidβ-peptide administration alone.Conversely,the activity of protein phosphatase 2A was remarkably reduced in rats with cerebral ischemia-reperfusion injury following amyloidβ-peptide administration.These findings suggest that amyloidβ-peptide can potentiate tau phosphorylation induced by cerebral ischemia-reperfusion and thereby aggravate cognitive impairment.     

Inhibition of P2X7R in the amygdala ameliorates symptoms of neuropathic pain after spared nerve injury in rats

The amygdala circuitry and P2X7 receptor (P2X7R) have both been shown to play important roles in the modulation of neuropathic pain (NP). However, little is known about the functional role of P2X7R in the amygdala for the regulation of NP. This study aims to evaluate the alleviative effect of intra-amygdala microinfusion of a pharmacological antagonist of P2X7R (A-438079) on NP and explore its possible mechanism of action. Male Sprague-Dawley rats were used to construct the animal model of NP through spared nerve injury (SNI). The SNI rats randomly received chronic bilateral microinjection of A-438079 (100 pmol/side) or saline into the amygdalae via cannulas. Mechanical paw withdrawal threshold (MWT) and thermal withdrawal duration (TWD) were measured by von Frey monofilaments. Besides, tail suspension test (TST), forced swimming test (FST), open field test (OFT) and sucrose preference test (SPT) were performed to assess depression- and anxiety-like behaviors. Immunofluorescence assay was employed to determine the levels of glial fibrillary acidic protein (GFAP), ionized calcium binding adaptor molecule 1 (IBA-1) and connexin 43 (Cx43) in the spinal cord. In addition, the change of growth associated protein 43 (GAP43) level in the spinal cord was assessed by Western blot. Our data showed that chronic treatment with A-438079 increased MWT and decreased TWD on days 11–21 post-SNI while decreased depression-like and anxiety-like behaviors. A-438079 administration significantly attenuated the elevated immunoreactivities of IBA-1 and GFAP in microglia and astrocytes after SNI. Furthermore, the decreased expression of GAP-43 in the spinal cord due to SNI was significantly attenuated by A-438079. However, when A-438079 and a pharmacological agonist (BzATP) of P2X7R were given simultaneously, all the effects caused by A-438079 alone were reversed. In brief, our study revealed the protective role of inhibiting P2X7R in the amygdala against symptoms associated with NP, possibly attributing to its inhibitory effects on spinal microglia and astrocytes.     

Cross Talk between α7 and α3β4 Nicotinic Receptors Prevents Their Desensitization in Human Chromaffin Cells

The physical interaction and functional cross talk among the different subtypes of neuronal nicotinic acetylcholine receptors (nAChRs) expressed in the various tissues is unknown. Here, we have investigated this issue between the only two nAChRs subtypes expressed, the α7 and α3β4 subtypes, in a human native neuroendocrine cell (the chromaffin cell) using electrophysiological patch-clamp, fluorescence, and Förster resonance energy transfer (FRET) techniques. Our data show that α7 and α3β4 receptor subtypes require their mutual and maximal efficacy of activation to increase their expression, to avoid their desensitization, and therefore, to increase their activity. In this way, after repetitive stimulation with acetylcholine (ACh), α7 and α3β4 receptor subtypes do not desensitize, but they do with choline. The nicotinic current increase associated with the α3β4 subtype is dependent on Ca²⁺. In addition, both receptor subtypes physically interact. Interaction and expression of both subtypes are reversibly reduced by tyrosine and serine/threonine phosphatases inhibition, not by Ca²⁺. In addition, expression is greater in human chromaffin cells from men compared to women, but FRET efficiency is not affected. Together, our findings indicate that human α7 and α3β4 subtypes mutually modulate their expression and activity, providing a promising line of research to pharmacologically regulate their activity.SIGNIFICANCE STATEMENT Desensitization of nicotinic receptors is accepted to occur with repetitive agonist stimulation. However, here we show that human native α3β4 and α7 nicotinic acetylcholine receptor (nAChR) subtypes do not desensitize, and instead, increase their activity when they are activated by the physiological agonist acetylcholine (ACh). An indispensable requirement is the activation of the other receptor subtype with maximal efficacy, and the presence of Ca²⁺ to cooperate in the case of the α3β4 current increase. Because choline is an α3β4 partial agonist, it will act as a limiting factor of nicotinic currents enhancement in the absence of ACh, but in its presence, it will further potentiate α7 currents.     

Mutant ubiquitin-mediated ��-secretase stability via activation of caspase-3 is related to ��-amyloid accumulation in ischemic striatum in rats

Previous studies have demonstrated that ischemic stroke increases β-amyloid (Aβ) production by increasing β-secretase (BACE1) through activation of caspase-3, and stimulates generation of mutant ubiquitin (UBB⁺¹) in rat brains. In this study, we examined whether caspase-3 activation participates in the regulation of UBB⁺¹ generation and UBB⁺¹-mediated BACE1 stability in ischemic injured brains. The results showed that UBB⁺¹ and activated caspase-3-immunopositive-stained cells were time dependently increased in the ipsilateral striatum of rat brains after middle cerebral artery occlusion. UBB⁺¹-immunopositive cells could be co-stained with caspase-3, Aβ (UBB⁺¹–Aβ), and BACE1 (UBB⁺¹–BACE1). BACE1 protein could also be pulled down by immunoprecipitation with UBB⁺¹ antibody. Z-DEVD-FMK (DEVD), a caspase-3 inhibitor, significantly decreased the level of UBB⁺¹ protein and the number of UBB⁺¹–Aβ and UBB⁺¹–BACE1 double-stained cells in the ischemic striatum, as well as the level of UBB⁺¹/BACE1 protein complex. We conclude that activation of caspase-3 might be upstream of UBB⁺¹ formation and that excessive UBB⁺¹ could bind to BACE1 and increase the stability of BACE1, thereby increasing Aβ in ischemic injured brains. These results suggest new biological and pathological effects of caspases and regulation of the ubiquitin–proteasome system in the brain. Our results provide new therapeutic targets to prevent further neurodegeneration in patients after stroke.     

Microglia‐derived purines modulate mossy fibre synaptic transmission and plasticity through P2X4 and A1 receptors

Recent data have provided evidence that microglia, the brain‐resident macrophage‐like cells, modulate neuronal activity in both physiological and pathophysiological conditions, and microglia are therefore now recognized as synaptic partners. Among different neuromodulators, purines, which are produced and released by microglia, have emerged as promising candidates to mediate interactions between microglia and synapses. The cellular effects of purines are mediated through a large family of receptors for adenosine and for ATP (P2 receptors). These receptors are present at brain synapses, but it is unknown whether they can respond to microglia‐derived purines to modulate synaptic transmission and plasticity. Here, we used a simple model of adding immune‐challenged microglia to mouse hippocampal slices to investigate their impact on synaptic transmission and plasticity at hippocampal mossy fibre (MF) synapses onto CA3 pyramidal neurons. MF–CA3 synapses show prominent forms of presynaptic plasticity that are involved in the encoding and retrieval of memory. We demonstrate that microglia‐derived ATP differentially modulates synaptic transmission and short‐term plasticity at MF–CA3 synapses by acting, respectively, on presynaptic P2X₄ receptors and on adenosine A₁ receptors after conversion of extracellular ATP to adenosine. We also report that P2X₄ receptors are densely located in the mossy fibre tract in the dentate gyrus–CA3 circuitry. In conclusion, this study reveals an interplay between microglia‐derived purines and MF–CA3 synapses, and highlights microglia as potent modulators of presynaptic plasticity.     

The novel amyloid-beta peptide aptamer inhibits intracellular amyloid-beta peptide toxicity

Amyloid β peptide binding alcohol dehydrogenase (ABAD) decoy peptide (DP) can competitively antagonize binding of amyloid β peptide to ABAD and inhibit the cytotoxic effects of amyloid β peptide. Based on peptide aptamers, the present study inserted ABAD-DP into the disulfide bond of human thioredoxin (TRX) using molecular cloning technique to construct a fusion gene that can express the TRX1-ABAD-DP-TRX2 aptamer. Moreover, adeno-associated virus was used to allow its stable expression. Immunofluorescent staining revealed the co-expression of the transduced fusion gene TRX1-ABAD-DP-TRX2 and amyloid β peptide in NIH-3T3 cells, indicating that the TRX1-ABAD-DP-TRX2 aptamer can bind amyloid β peptide within cells. In addition, cell morphology and MTT results suggested that TRX1-ABAD-DP-TRX2 attenuated amyloid β peptide-induced SH-SY5Y cell injury and improved cell viability. These findings confirmed the possibility of constructing TRX-based peptide aptamer using ABAD-DP. Moreover, TRX1-ABAD-DP-TRX2 inhibited the cytotoxic effect of amyloid β peptide.     

The cytokine IL-1beta transiently enhances P2X7 receptor expression and function in human astrocytes

Extracellular nucleotide di- and triphosphates such as ATP and ADP mediate their effects through purinergic P2 receptors belonging to either the metabotropic P2Y or the ionotropic P2X receptor family. The P2X7R is a unique member of the P2X family, which forms a pore in response to ligand stimulation, regulating cell permeability, cytokine release, and/or apoptosis. This receptor is also unique in that its affinity for the ligand benzoyl-benzoyl ATP (BzATP) is at least 10-fold greater than that of ATP. Primary human fetal astrocytes in culture express low-levels of P2X7R mRNA and protein, and BzATP induces only a slight influx in intracellular calcium [Ca2+]i, with little demonstrable effect on gene expression or pore formation in these cells. We now show that, following treatment with the proinflammatory cytokine IL-1beta, BzATP induces a robust rise in [Ca2+]i with agonist and antagonist profiles indicative of the P2X7R. IL-1beta also induced the formation of membrane pores as evidenced by the uptake of YO-PRO-1 (375 Da). Quantitative real-time PCR demonstrated transient upregulation of P2X7R mRNA in IL-1beta-treated cells, while FACS analysis indicated a similar upregulation of P2X7R protein at the cell membrane. In multiple sclerosis lesions, immunoreactivity for the P2X7R was demonstrated on reactive astrocytes in autopsy brain tissues. In turn, P2X7R stimulation increased the production of IL-1-induced nitric oxide synthase activity by astrocytes in culture. These studies suggest that signaling via the P2X7R may modulate the astrocytic response to inflammation in the human central nervous system.     

Distribution of P2X receptors on astrocytes in juvenile rat hippocampus

Recent evidence suggested that ATP acting via ionotropic (P2X) and metabotropic (P2Y) purinergic receptors might be involved in signaling between glial cells and within glial-neuronal networks. In contrast to their neuronal counterpart, the identity of P2X receptors in CNS glial cells is largely unknown. In the present study, antibodies recognizing the subunits P2X1-P2X7 were applied together with the astroglial marker S100beta and nuclear labeling with Hoechst 33342 to investigate semiquantitatively the distribution of the whole set of P2X receptors in astrocytes of the juvenile rat hippocampus. Expression of P2X1-P2X4, P2X6, and P2X7 subunits was observed in astrocytes of various hippocampal subregions, but the cells were completely devoid of P2X5 protein. S100beta-positive cells expressing subunits P2X3-P2X7 occurred evenly in the different subfields, while P2X1- and P2X2-positive astrocytes were distributed more heterogeneously. The staining pattern of P2X subunits also differed at the subcellular level. Antibodies against P2X2 and P2X4 labeled both astroglial cell bodies and processes. Immunoreactivity for P2X1 and P2X6 was mainly confined to somatic areas of S100beta-positive cells, whereas the subunit P2X3 was primarily localized along astroglial processes. Knowledge of the distribution of P2X receptors might provide a basis for a better understanding of their specific role in cell-cell signaling.     

The antihyperalgesic activity of a selective P2X7 receptor antagonist, A-839977, is lost in IL-1αβ knockout mice

The pro-inflammatory cytokine interleukin-1β (IL-1β) has been implicated in both inflammatory processes and nociceptive neurotransmission. Activation of P2X7 receptors is the mechanism by which ATP stimulates the rapid maturation and release of IL-1β from macrophages and microglial cells. Recently, selective P2X7 receptor antagonists have been shown to reduce inflammatory and neuropathic pain in animal models. However, the mechanisms underlying these analgesic effects are unknown. The present studies characterize the pharmacology and antinociceptive effects of a structurally novel P2X7 antagonist. A-839977 potently (IC₅₀ = 20–150 nM) blocked BzATP-evoked calcium influx at recombinant human, rat and mouse P2X7 receptors. A-839977 also potently blocked agonist-evoked YO-PRO uptake and IL-1β release from differentiated human THP-1 cells. Systemic administration of A-839977 dose-dependently reduced thermal hyperalgesia produced by intraplantar administration of complete Freund's adjuvant (CFA) (ED₅₀ = 100 μmol/kg, i.p.) in rats. A-839977 also produced robust antihyperalgesia in the CFA model of inflammatory pain in wild-type mice (ED₅₀ = 40 μmol/kg, i.p.), but the antihyperalgesic effects of A-839977 were completely absent in IL-1αβ knockout mice. These data demonstrate that selective blockade of P2X7 receptors in vivo produces significant antinociception in animal models of inflammatory pain and suggest that the antihyperalgesic effects of P2X7 receptor blockade in an inflammatory pain model in mice are mediated by blocking the release of IL-1β.     

P2X4 receptors control the fate and survival of activated microglia

Microglia, the resident immune cells of the central nervous system, responds to brain disarrangements by becoming activated to contend with brain damage. Here we show that the expression of P2X4 receptors is upregulated in inflammatory foci and in activated microglia in the spinal cord of rats with experimental autoimmune encephalomyelitis (EAE) as well as in the optic nerve of multiple sclerosis patients. To study the role of P2X4 receptors in microgliosis, we activated microglia with LPS in vitro and in vivo. We observed that P2X4 receptor activity in vitro was increased in LPS‐activated microglia as assessed by patch‐clamp recordings. In addition, P2X4 receptor blockade significantly reduced microglial membrane ruffling, TNFα secretion and morphological changes, as well as LPS‐induced microglial cell death. Accordingly, neuroinflammation provoked by LPS injection in vivo induced a rapid microglial loss in the spinal cord that was totally prevented or potentiated by P2X4 receptor blockade or facilitation, respectively. Within the brain, microglia in the hippocampal dentate gyrus showed particular vulnerability to LPS‐induced neuroinflammation. Thus, microglia processes in this region retracted as early as 2 h after injection of LPS and died around 24 h later, two features which were prevented by blocking P2X4 receptors. Together, these data suggest that P2X4 receptors contribute to controlling the fate of activated microglia and its survival.GLIA 2014;62:171–184     

Inhibition of P2X receptor-mediated inward current by protein kinase C in small-diameter dorsal root ganglion neurons of adult rats

Objective

The aim of the present study is to verify the ATP-induced varied responses in isolated dorsal root ganglion (DRG) neurons of the adult rat, and investigate the modulatory effects of specific P2X receptor agonist β, γ-me-ATP and protein kinase C (PKC) on P2X receptor-mediated inward current in DRG neurons.

Methods

Whole cell patch-clamp was employed to record the currents on acutely isolated DRG neurons in the adult rats.

Results

β, γ-me-ATP, similar as ATP, evoked 2 distinct subtypes of P2X receptor-mediated inward currents in a dose-dependent manner in DRG neurons. Activation of PKC by phorbol 12,13-dibutyrate (PDBu) significantly inhibited both subtypes of inward currents mediated by P2X receptors in a dose-dependent manner.

Conclusion

Activation of PKC negatively modulated the P2X receptor-mediated currents in rat DRG neurons, which may be of benefit to preventing the over-excitation of nociceptor under inflammatory or neuropathic conditions.