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Cellular context determines primary characteristics of human TRPC5 as a cold-activated channel.

The human transient receptor potential canonical 5 (TRPC5) is a calcium-permeable, nonselective cation channel expressed in the central and peripheral nervous system and also in other tissues such as the kidney, synovium, and odontoblasts. TRPC5 has been recently confirmed to play a key role in spontaneous, inflammatory mechanical, and cold pain. Although TRPC5 activation is known to be cold sensitive, it is unclear whether this property is intrinsic to the channel protein and whether or to what extent it may be determined by the cellular environment. In this study, we explored the cold sensitivity of human TRPC5 at the single-channel level using transiently transfected HEK293T cells. Upon decreasing the temperature, the channel demonstrated prolonged mean open dwell times and a robust increase in the open probability (P ), whereas the amplitude of unitary currents decreased ~1.5-fold per 10°C of temperature difference. In the absence of any agonists, the temperature dependence of P was sigmoidal, with a steep slope within the temperature range of 16°C-11°C, and exhibited saturation below 8-5°C. Thermodynamic analysis revealed significant changes in enthalpy and entropy, suggesting that substantial conformational changes accompany cold-induced gating. The mutant channel T970A, in which the regulation downstream of G-protein coupled receptor signaling was abrogated, exhibited higher basal activity at room temperature and a less steep temperature response profile, with an apparent threshold below 22°C. An even more pronounced decrease in the activation threshold was observed in a mutant that disrupted the electrostatic interaction of TRPC5 with the endoplasmic reticulum calcium sensor stromal interaction molecule 1. Thus, TRPC5 exhibits features of an intrinsically cold-gated channel; its sensitivity to cold tightly depends on the phosphorylation status of the protein and intracellular calcium homeostasis.

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Distinct basolateral amygdala excitatory inputs mediate the somatosensory and aversive-affective components of pain.

Pain is a multidimensional perception that includes unpleasant somatosensory and affective experiences; however, the underlying neural circuits that mediate different components of pain remain elusive. Although hyperactivity of basolateral amygdala glutamatergic (BLA) neurons is required for the somatosensory and emotional processing of pain, the precise excitatory inputs to BLA neurons and their roles in mediating different aspects of pain are unclear. Here, we identified two discrete glutamatergic neuronal circuits in male mice: a projection from the insular cortex (IC) to BLA neurons, which modulates both the somatosensory and affective components of pain, and a projection from the mediodorsal thalamic nucleus (MD) to BLA neurons, which modulates only the aversive-affective component of pain. Using whole-cell recording and fiber photometry, we found that neurons within the IC→BLA and MD→BLA pathways were activated in mice upon inflammatory pain induced by injection of complete Freund's adjuvant (CFA) into the paw. Optical inhibition of the IC→BLA pathway increased the nociceptive threshold and induced behavioral place preference in CFA mice. In contrast, optical inhibition of the MD→BLA pathway did not affect the nociceptive threshold, but still induced place preference in CFA mice. In normal mice, optical activation of the IC→BLA pathway decreased the nociceptive threshold and induced place aversion, while optical activation of the MD→BLA pathway only evoked aversion. Taken together, our results demonstrate that discrete IC→BLA and MD→BLA pathways are involved in modulating different components of pain, and provide insights into its circuit basis and better our understanding of pain perception.

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Scorpion Neurotoxin Syb-prII-1 Exerts Analgesic Effect through Nav1.8 Channel and MAPKs Pathway.

Trigeminal neuralgia (TN) is a common type of peripheral neuralgia in clinical practice, which is usually difficult to cure. Common analgesic drugs are difficult for achieving the desired analgesic effect. Syb-prII-1 is a β-type scorpion neurotoxin isolated from the scorpion venom of It has an important influence on the voltage-gated sodium channel (VGSCs), especially closely related to Nav1.8 and Nav1.9. To explore whether Syb-prII-1 has a good analgesic effect on TN, we established the Sprague Dawley (SD) rats' chronic constriction injury of the infraorbital nerve (IoN-CCI) model. Behavioral, electrophysiological, Western blot, and other methods were used to verify the model. It was found that Syb-prII-1 could significantly relieve the pain behavior of IoN-CCI rats. After Syb-prII-1 was given, the phosphorylation level of the mitogen-activated protein kinases (MAPKs) pathway showed a dose-dependent decrease after IoN-CCI injury. Moreover, Syb-prII-1(4.0 mg/kg) could significantly change the steady-state activation and inactivation curves of Nav1.8. The steady-state activation and inactivation curves of Nav1.9 were similar to those of Nav1.8, but there was no significant difference. It was speculated that it might play an auxiliary role. The binding mode, critical residues, and specific interaction type of Syb-prII-1 and VSD2 were clarified with computational simulation methods. Our results indicated that Syb-prII-1 could provide a potential treatment for TN by acting on the Nav1.8 target.

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Alterations of monoamine neurotransmitters, HPA-axis hormones, and inflammation cytokines in reserpine-induced hyperalgesia and depression comorbidity rat model.

Pain and depression often occur simultaneously, but the mechanism of this condition is still unclear.

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PIEZO1 transduces mechanical itch in mice.

Itch triggers scratching, a behavioural defence mechanism that aids in the removal of harmful irritants and parasites. Chemical itch is triggered by many endogenous and exogenous cues, such as pro-inflammatory histamine, which is released during an allergic reaction. Mechanical itch can be triggered by light sensations such as wool fibres or a crawling insect. In contrast to chemical itch pathways, which have been extensively studied, the mechanisms that underlie the transduction of mechanical itch are largely unknown. Here we show that the mechanically activated ion channel PIEZO1 (ref. ) is selectively expressed by itch-specific sensory neurons and is required for their mechanically activated currents. Loss of PIEZO1 function in peripheral neurons greatly reduces mechanically evoked scratching behaviours and both acute and chronic itch-evoked sensitization. Finally, mice expressing a gain-of-function Piezo1 allele exhibit enhanced mechanical itch behaviours. Our studies reveal the polymodal nature of itch sensory neurons and identify a role for PIEZO1 in the sensation of itch.

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Chronic inhibition of the mitochondrial ATP synthase in skeletal muscle triggers sarcoplasmic reticulum distress and tubular aggregates.

Tubular aggregates (TA) are honeycomb-like arrays of sarcoplasmic-reticulum (SR) tubules affecting aged glycolytic fibers of male individuals and inducing severe sarcomere disorganization and muscular pain. TA develop in skeletal muscle from Tubular Aggregate Myopathy (TAM) patients as well as in other disorders including endocrine syndromes, diabetes, and ageing, being their primary cause unknown. Nowadays, there is no cure for TA. Intriguingly, both hypoxia and calcium dyshomeostasis prompt TA formation, pointing to a possible role for mitochondria in their setting. However, a functional link between mitochondrial dysfunctions and TA remains unknown. Herein, we investigate the alteration in muscle-proteome of TAM patients, the molecular mechanism of TA onset and a potential therapy in a preclinical mouse model of the disease. We show that in vivo chronic inhibition of the mitochondrial ATP synthase in muscle causes TA. Upon long-term restrained oxidative phosphorylation (OXPHOS), oxidative soleus experiments a metabolic and structural switch towards glycolytic fibers, increases mitochondrial fission, and activates mitophagy to recycle damaged mitochondria. TA result from the overresponse of the fission controller DRP1, that upregulates the Store-Operate-Calcium-Entry and increases the mitochondria-SR interaction in a futile attempt to buffer calcium overloads upon prolonged OXPHOS inhibition. Accordingly, hypoxic muscles cultured ex vivo show an increase in mitochondria/SR contact sites and autophagic/mitophagic zones, where TA clusters grow around defective mitochondria. Moreover, hypoxia triggered a stronger TA formation upon ATP synthase inhibition, and this effect was reduced by the DRP1 inhibitor mDIVI. Remarkably, the muscle proteome of TAM patients displays similar alterations in mitochondrial dynamics and in ATP synthase contents. In vivo edaravone treatment in mice with restrained OXPHOS restored a healthy phenotype by prompting mitogenesis and mitochondrial fusion. Altogether, our data provide a functional link between the ATP synthase/DRP1 axis and the setting of TA, and repurpose edaravone as a possible treatment for TA-associated disorders.

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LncRNA-UC.25 + shRNA Alleviates P2Y Receptor-Mediated Diabetic Neuropathic Pain via STAT1.

Diabetic neuropathic pain (DNP) is a common complication of diabetes, and its complicated pathogenesis, as well as clinical manifestations, has brought great trouble to clinical treatment. The spinal cord is an important part of regulating the occurrence and development of DNP. Spinal microglia can regulate the activity of spinal cord neurons and have a regulatory effect on chronic pain. P2Y receptor is involved in DNP. P2Y and P2Y receptors belong to the Gi subtype of P2Y receptors, but there is no report that the P2Y receptor is involved in DNP. Closely related to many human diseases, the dysregulation of long noncoding RNA (lncRNA) has the effect of promoting or inhibiting the occurrence and development of diseases. The aim of this research is to investigate the function of the spinal cord P2Y receptor in type 2 DNP and to understand the function as well as the possible mechanism of lncRNA-UC.25 + (UC.25 +) in rat spinal cord P2Y receptor-mediated DNP. Our results showed that P2Y shRNA can reduce the expression of P2Y in DNP rats, thereby restraining the activation of microglia, decreasing the expression of inflammatory factors and the level of p38 mitogen-activated protein kinase (p38 MAPK) phosphorylation. At the same time, UC.25 + shRNA can downregulate the expression of the P2Y receptor, reduce the release of inflammatory factors, and diminish the p38 MAPK phosphorylation, indicating that UC.25 + can alleviate spinal cord P2Y receptor-mediated DNP. The RNA immunoprecipitation result showed that UC.25 + enriched signal transducers and activators of transcription1 (STAT1) and positively regulated its expression. The chromatin immunoprecipitation result indicated that STAT1 combined with the promoter region of the P2Y receptor and positively regulated the expression of the P2Y receptor. Therefore, we infer that UC.25 + may alleviate DNP in rats by regulating the expression of the P2Y receptor in spinal microglia via STAT1.

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L-Glutamine alleviates osteoarthritis by regulating lncRNA-NKILA expression through the TGF-β1/SMAD2/3 signalling pathway.

Osteoarthritis (OA) is a heterogeneous condition characterised by cartilage degradation, subchondral sclerosis, and osteophyte formation, and accompanied by the generation of pro-inflammatory mediators and degradation of extracellular matrix. The current treatment for early OA is focused on the relief of symptoms, such as pain, but this treatment cannot delay the pathological process. L-Glutamine (L-Gln), which has anti-inflammatory and anti-apoptotic effects, is the most abundant amino acid in human blood. However, its role in OA has not been systematically studied. Therefore, the objective of this work was to explore the therapeutic effect and molecular mechanism of L-Gln on OA. In vitro, we found that L-Gln could upregulate the expression of the long non-coding RNA NKILA, which is regulated by the transforming growth factor-β1/SMAD2/3 pathway, and inhibit the activity of nuclear factor-κB, thereby decreasing the expression of nitric oxide synthase, cyclooxygenase-2, and matrix metalloproteinase-13 (MMP-13). This led to a reduction in the generation of nitrous oxide, prostaglandin E-2, tumour necrosis factor-α, and degradation of the extracellular matrix (i.e. aggrecan and collagen II) in rat OA chondrocytes. Moreover, intragastric administration of L-Gln reduced the degradation of cartilage tissue and expression of MMP-13 in a rat OA model. L-Gln also relieved the clinical symptoms in some patients with early knee joint OA. These findings highlight that L-Gln is a potential therapeutic drug to delay the occurrence and development of OA.

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Positive interaction between GPER and β-alanine in the dorsal root ganglion uncovers potential mechanisms: mediating continuous neuronal sensitization and neuroinflammation responses in neuropathic pain.

The pathogenesis of neuropathic pain and the reasons for the prolonged unhealing remain unknown. Increasing evidence suggests that sex oestrogen differences play a role in pain sensitivity, but few studies have focused on the oestrogen receptor which may be an important molecular component contributing to peripheral pain transduction. We aimed to investigate the impact of oestrogen receptors on the nociceptive neuronal response in the dorsal root ganglion (DRG) and spinal dorsal horn using a spared nerve injury (SNI) rat model of chronic pain.

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Phenotypes of Motor Deficit and Pain after Experimental Spinal Cord Injury.

Motor disability is a common outcome of spinal cord injury (SCI). The recovery of motor function after injury depends on the severity of neurotrauma; motor deficit can be reversible, at least partially, due to the innate tissue capability to recover, which, however, deteriorates with age. Pain is often a comorbidity of injury, although its prediction remains poor. It is largely unknown whether pain can attend motor dysfunction. Here, we implemented SCI for modelling severe and moderate neurotrauma and monitored SCI rats for up to 5 months post-injury to determine the profiles of both motor deficit and nociceptive sensitivity. Our data showed that motor dysfunction remained persistent after a moderate SCI in older animals (5-month-old); however, there were two populations among young SCI rats (1 month-old) whose motor deficit either declined or exacerbated even more over 4-5 weeks after identical injury. All young SCI rats displayed changed nociceptive sensitivity in thermal and mechanical modalities. The regression analysis of the changes revealed a population trend with respect to hyper- or hyposensitivity/motor deficit. Together, our data describe the phenotypes of motor deficit and pain, the two severe complications of neurotrauma. Our findings also suggest the predictability of motor dysfunction and pain syndromes following SCI that can be a hallmark for long-term rehabilitation and recovery after injury.

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