Sensitization Of Sensory Neurons Research Articles

Neuroimmune Interactions in the Intestine

Recent advances have contributed to a mechanistic understanding of neuroimmune interactions in the intestine and revealed an essential role of this cross talk for gut homeostasis and modulation of inflammatory and infectious intestinal diseases. In this review, we describe the innervation of the intestine by intrinsic and extrinsic neurons and then focus on the bidirectional communication between neurons and immune cells. First, we highlight the contribution of neuronal subtypes to the development of colitis and discuss the different immune and epithelial cell types that are regulated by neurons via the release of neuropeptides and neurotransmitters. Next, we review the role of intestinal inflammation in the development of visceral hypersensitivity and summarize how inflammatory mediators induce peripheral and central sensitization of gut-innervating sensory neurons. Finally, we outline the importance of immune cells and gut microbiota for the survival and function of different neuronal populations at homeostasis and during bacterial and helminth infection.

ПРЕДПОЛАГАЕМЫЙ МЕХАНИЗМ ВЛИЯНИЯ КРАТКОВРЕМЕННОЙ ГИПОКСИИ НА ОБОНЯТЕЛЬНУЮ ЧУВСТВИТЕЛЬНОСТЬ

The article is focused on the effect of short-term hypoxia on the olfactory sensitivity. The effect of short-term hypoxia on the olfactory sensitivity in human and rats were studied by determination thresholds for n-butanol. We have shown that reduced oxygen partial pressure leads to reduction of olfactory sensitivity. As known, odorant detection thresholds characterize the functional state of receptor cells. It can be assumed that the weakening of their sensitivity may be caused by a lack of energy supply to the olfactory perception process in hypoxia. To test this hypothesis, we stimulated rat olfactory sensory neurons with n-butanol in rotenone model. The reaction of cellular respiration of olfactory sensory neurons was evaluated by changing the intensity of intrinsic fluorescence of reduced pyridine nucleotides (NADH) according to the B. Chance method. Analysis of the results showed that under normoxia, stimulation of olfactory neurons by odorant caused an increase in the intensity of NADH fluorescence, which indicates the accumulation of the reduced form of pyridine nucleotides in mitochondria, and, consequently, the activation of cellular respiration due to exposure to odorant. In hypoxia caused by rotenone, an increase in the induced change of fluorescence of NADH was either not registered or the amplitude of the reaction was halved. This means that the recovery of NADH due to the odorant either did not occur at all, or this reaction to the smell was significantly lower than with normoxia, which indicates a weakening of cellular respiration during hypoxia. This is the reason for the decrease in ATP synthesis, which affects the efficiency of the olfactory transduction process and the sensitivity of olfactory sensory neurons. Therefore, based on our results, it can be concluded that the weakening of olfactory sensitivity in rats and humans with hypoxia may be due to a weakening of cellular respiration in olfactory sensory neurons.

Knockout of OR39 reveals redundancy in the olfactory pathway regulating the acquisition of host seeking in Anopheles coluzzii.

The attraction of anthropophilic mosquitoes to human host cues, such as body odour and carbon dioxide, gradually increases during adult maturation. This acquisition of host-seeking behaviour correlates with age-dependent changes in odorant receptor (OR) transcript abundance and sensitivity of olfactory sensory neurons (OSNs). One OR gene of the human malaria vector, Anopheles coluzzii, AcolOR39, is significantly downregulated in mature females, and a cognate ligand of AcolOR39, sulcatone, a major component of human emanations, mediates the observed behavioural inhibition of newly emerged (teneral) females to human body odour. Knockout of AcolOR39, using CRISPR-Cas9 mutagenesis, selectively abolished sulcatone detection in OSNs, housed in trichoid sensilla. However, knockout of AcolOR39 altered neither the response rate nor the flight behaviour of teneral females in a wind tunnel, indicating the involvement of other genes, and thus a redundancy, in regulating the acquisition of host seeking in mosquitoes.

Bradykinin receptor expression and bradykinin-mediated sensitization of human sensory neurons.

Bradykinin is a peptide implicated in inflammatory pain in both humans and rodents. In rodent sensory neurons, activation of B1 and B2 bradykinin receptors induces neuronal hyperexcitability. Recent evidence suggests that human and rodent dorsal root ganglia (DRG), which contain the cell bodies of sensory neurons, differ in the expression and function of key GPCRs and ion channels; whether bradykinin receptor expression and function are conserved across species has not been studied in depth. In this study, we used human DRG tissue from organ donors to provide a detailed characterization of bradykinin receptor expression and bradykinin-induced changes in the excitability of human sensory neurons. We found that B2 and, to a lesser extent, B1 receptors are expressed by human DRG neurons and satellite glial cells. B2 receptors were enriched in the nociceptor subpopulation. Using patch-clamp electrophysiology, we found that acute bradykinin increases the excitability of human sensory neurons, whereas prolonged exposure to bradykinin decreases neuronal excitability in a subpopulation of human DRG neurons. Finally, our analyses suggest that donor's history of chronic pain and age may be predictors of higher B1 receptor expression in human DRG neurons. Together, these results indicate that acute bradykinin-induced hyperexcitability, first identified in rodents, is conserved in humans and provide further evidence supporting bradykinin signaling as a potential therapeutic target for treating pain in humans.

Abstract P3015: Mechanosensitive, Piezo2 Channels Mediate The Exaggerated Exercise Pressor Reflex In A Mouse Model Of Hind Limb Ischemia.

The exercise pressor reflex (EPR) is a reflexive mechanism in which blood pressure and heart rate increase in response to afferent neuron activation by skeletal muscle contraction. Patients with peripheral artery disease (PAD) have an exaggerated EPR, contributing to exercise intolerance, morbidity and mortality. Recently, we developed a mouse model which we used to determine the effect of femoral artery ligation (FAL) on the EPR. We observed that the EPR is exaggerated 72 hrs following FAL in mice. Stimulation of metabolically active afferent neurons was not different between FAL and control. In contrast, the responses induced by static contraction (stimulation of both mechanically and metabolically sensitive afferent fibers) and passive stretch (preferential stimulation of mechanically sensitive afferent neurons) were exaggerated in FAL mice when compared to control. These exaggerated responses were not affected by SB-705498 (a TRPv1 receptor inhibitor) but were nearly abolished by Gadolinium (a mechanical sensitive group III afferent neuron inhibitor) and significantly reduced by GsMTx-4 (a selective inhibitor of the PIEZO2 receptor).. Furthermore, we observed that PIEZO2 expression was upregulated in the dorsal root ganglia (DRG) from FAL mice compared to controls. Importantly, FAL mice displayed a decreased exercise capacity when compared to control mice, verifying that this mouse model mimics human PAD and supporting the notion that an exaggerated EPR contributes to exercise intolerance. These results further indicate that the exaggerated EPR in FAL is mediated, in part, by the PIEZO2 receptor. To explore this, we generated mice with a conditional, DRG targeted, deletion of Piezo2 . In these mice, we observed that the responses to static contraction and passive stretch were significantly reduced compared to control. Moreover, we performed whole-cell voltage-clamp on dissociated DRG observing increased negative pressure induced inward currents in the setting of FAL while these currents were decreased in the Piezo2 -/- DRG. These data indicate that PIEZO2 expression may serve as a therapeutic target in the treatment of PAD.

Engineering a multicompartment in vitro model for dorsal root ganglia phenotypic assessment.

Despite the significant global prevalence of chronic pain, current methods to identify pain therapeutics often fail translation to the clinic. Phenotypic screening platforms rely on modeling and assessing key pathologies relevant to chronic pain, improving predictive capability. Patients with chronic pain often present with sensitization of primary sensory neurons (that extend from dorsal root ganglia [DRG]). During neuronal sensitization, painful nociceptors display lowered stimulation thresholds. To model neuronal excitability, it is necessary to maintain three key anatomical features of DRGs to have a physiologically relevant platform: (1) isolation between DRG cell bodies and neurons, (2) 3D platform to preserve cell-cell and cell-matrix interactions, and (3) presence of native non-neuronal support cells, including Schwann cells and satellite glial cells. Currently, no culture platforms maintain the three anatomical features of DRGs. Herein, we demonstrate an engineered 3D multicompartment device that isolates DRG cell bodies and neurites and maintains native support cells. We observed neurite growth into isolated compartments from the DRG using two formulations of collagen, hyaluronic acid, and laminin-based hydrogels. Further, we characterized the rheological, gelation and diffusivity properties of the two hydrogel formulations and found the mechanical properties mimic native neuronal tissue. Importantly, we successfully limited fluidic diffusion between the DRG and neurite compartment for up to 72 h, suggesting physiological relevance. Lastly, we developed a platform with the capability of phenotypic assessment of neuronal excitability using calcium imaging. Ultimately, our culture platform can screen neuronal excitability, providing a more translational and predictive system to identify novel pain therapeutics to treat chronic pain.

Resensitization of TRPV1 channels after the P2 receptor activation in sensory neurons of spinal ganglia in rats.

TRPV1 channels are responsible for detecting noxious stimuli such as heat (>43°C), acid, and capsaicin. P2 receptors are involved in numerous functions of the nervous system, including its modulation and specific response to the application of ATP. In our experiments, we investigated the dynamics of calcium transients in DRG neurons associated with TRPV1 channel desensitization and the effect of activation of P2 receptors on this process. We used DRG neurons from rats P7-8 after 1-2 days of culture to measure calcium transients by microfluorescence calcimetry using the fluorescent dye Fura-2 AM. We have shown that DRG neurons of small (d < 22 μm) and medium (d = 24-35 μm) sizes differ in TRPV1 expression. Thus, TRPV1 channels are mainly present in small nociceptive neurons (59% of the studied neurons). Short-term sequential application of the TRPV1 channel agonist capsaicin (100nM) leads to the desensitization of TRPV1 channels by the type of tachyphylaxis. We identified three types of sensory neurons based on responses to capsaicin: (1) desensitized 37.5%, (2) non-desensitized 34.4%, and (3) insensitive 23.4% to capsaicin. It has also been shown that P2 receptors are present in all types of neurons according to their size. So, the responses to ATP were different in different-sized neurons. Applying ATP (0.1 mM) to the intact cell membrane after the onset of tachyphylaxis caused recovery of calcium transients in response to the addition of capsaicin in these neurons. The amplitude of the capsaicin response after reconstitution with ATP was 161% of the previous minimal calcium transient in response to capsaicin. Significantly, the restoration of the amplitude of calcium transients under the ATP application is not associated with changes in the cytoplasmic pool of ATP because this molecule does not cross the intact cell membrane, thus, our results show the interaction between TRPV1 channels and P2 receptors. It is important to note that the restoration of the amplitude of calcium transients through TRPV1 channels after application of ATP was observed mainly in cells of 1-2 days of cultivation. Thus, the resensitization of capsaicin transients following P2 receptor activation may be associated with the regulation of the sensitivity of sensory neurons.

Identification of key factors driving inflammation-induced sensitization of muscle sensory neurons.

Sensory neurons embedded in muscle tissue that initiate pain sensations, i.e., nociceptors, are temporarily sensitized by inflammatory mediators during musculoskeletal trauma. These neurons transduce peripheral noxious stimuli into an electrical signal [i.e., an action potential (AP)] and, when sensitized, demonstrate lower activation thresholds and a heightened AP response. We still do not understand the relative contributions of the various transmembrane proteins and intracellular signaling processes that drive the inflammation-induced hyperexcitability of nociceptors. In this study, we used computational analysis to identify key proteins that could regulate the inflammation-induced increase in the magnitude of AP firing in mechanosensitive muscle nociceptors. First, we extended a previously validated model of a mechanosensitive mouse muscle nociceptor to incorporate two inflammation-activated G protein-coupled receptor (GPCR) signaling pathways and validated the model simulations of inflammation-induced nociceptor sensitization using literature data. Then, by performing global sensitivity analyses that simulated thousands of inflammation-induced nociceptor sensitization scenarios, we identified three ion channels and four molecular processes (from the 17 modeled transmembrane proteins and 28 intracellular signaling components) as potential regulators of the inflammation-induced increase in AP firing in response to mechanical forces. Moreover, we found that simulating single knockouts of transient receptor potential ankyrin 1 (TRPA1) and reducing the rates of Gαq-coupled receptor phosphorylation and Gαq subunit activation considerably altered the excitability of nociceptors (i.e., each modification increased or decreased the inflammation-induced fold change in the number of triggered APs compared to when all channels were present). These results suggest that altering the expression of TRPA1 or the concentration of intracellular Gαq might regulate the inflammation-induced increase in AP response of mechanosensitive muscle nociceptors.

Cutaneous neuropeptides: the missing link between psychological stress and chronic inflammatory skin disease?

A "brain-skin" connection has been long been observed between chronic stress and chronic inflammatory skin disease including urticaria, psoriasis, atopic dermatitis, and prurigo nodularis. The relationship appears to be bidirectional. Chronic psychological stress has been shown to sustain hyperactivity of the sympathetic branch of the autonomic nervous system. Chronic stress is proinflammatory and in the context of several dermatologic disorders may be associated with an increase in dermal nerve fiber density, mast cells, nerve growth factor and calcitonin-gene-related peptide (CGRP). Furthermore, CGRP elicits a TH2-polarized T-cell response that is a hallmark of chronic pruritic conditions such as atopic dermatitis and prurigo nodularis. This TH2 response contributes directly to acute pruritus as well as the sensitization of cutaneous sensory neurons that are critical for chronic pruritus. Prurigo nodularis is a debilitating skin disorder featuring prominent nerve structural, neuropeptide, and TH2 cytokine aberrations that is a model deserving of future study.

Phenotypic screen identifies the natural product silymarin as a novel anti-inflammatory analgesic.

Sensory neuron hyperexcitability is a critical driver of pathological pain and can result from axon damage, inflammation, or neuronal stress. G-protein coupled receptor signaling can induce pain amplification by modulating the activation of Trp-family ionotropic receptors and voltage-gated ion channels. Here, we sought to use calcium imaging to identify novel inhibitors of the intracellular pathways that mediate sensory neuron sensitization and lead to hyperexcitability. We identified a novel stimulus cocktail, consisting of the SSTR2 agonist L-054,264 and the S1PR3 agonist CYM5541, that elicits calcium responses in mouse primary sensory neurons in vitro as well as pain and thermal hypersensitivity in mice in vivo. We screened a library of 906 bioactive compounds and identified 24 hits that reduced calcium flux elicited by L-054,264/CYM5541. Among these hits, silymarin, a natural product derived from milk thistle, strongly reduced activation by the stimulation cocktail, as well as by a distinct inflammatory cocktail containing bradykinin and prostaglandin E2. Silymarin had no effect on sensory neuron excitability at baseline, but reduced calcium flux via Orai channels and downstream mediators of phospholipase C signaling. In vivo, silymarin pretreatment blocked development of adjuvant-mediated thermal hypersensitivity, indicating potential use as an anti-inflammatory analgesic.

Sickle Cell Disease Patient Plasma Sensitizes iPSC-Derived Sensory Neurons from Sickle Cell Disease Patients

Individuals living with sickle cell disease (SCD) experience severe recurrent acute and chronic pain. The underlying neurobiology of the complex pain of SCD is not well understood. Limited and often ineffective treatment options and multifactorial healthcare disparities create an immediate need to develop non-opioid based SCD pain therapies. However, in order to develop novel therapies, we must better understand the neurobiological mechanisms underlying SCD pain. Although humanized mouse models of SCD do exist, growing evidence indicates there are striking differences in gene expression and functional responsiveness between human and mouse dorsal root ganglia (DRG) sensory neurons. Human DRG tissues are extremely limited in availability, postmortem tissues only provide static, endpoint molecular signatures, and obtaining DRGs from patients with SCD is untenable. These are all barriers to gaining mechanistic insight into pathogenic pain processes. Human induced pluripotent stem cell (iPSC)-derived sensory neurons (iSNs) have recently been used to model the altered function and expression of molecules in native DRG neurons in chronic pain disorders. These iPSC-derived SNs express canonical markers of human nociceptors and exhibit functional responses to noxious stimuli similar to those properties of native mammalian nociceptors. The objective of this study was to utilize iSNs differentiated from SCD patient iPSCs as well as age- and race-matched healthy controls (HCs) to characterize baseline receptor expression and function of SCD iSNs compared to HC iSNs. Our goal is to assess the intrinsic properties of SCD nociceptors that may contribute to the persistent sensitization of nociceptors in SCD patients with pain. To this end, both HC and SCD iSNs expressed transcripts for canonical markers of mature sensory neurons (NF200, CGRP), immune receptors (CCR1/2/3/5, CD4, CRLF2, CSF2Ra/b), and functional ion channels (HCN1, Nav1.7, P2X3/4, CACNA2D1, CHRNA9) (Table 1). Further, they expressed transcripts for ion channels relevant to SCD pain (Piezo1, Piezo2, TRPA1, TRPC5, TRPM3, TRPM8, TRPV1, TRPV4). We confirmed the expression of Peripherin, CGRP, TRPV1 at the protein level using immunocytochemistry. There were no significant differences in expression for any of these targets in iSNs derived from SCD patients compared to those from HCs. However, we did identify trends for higher endothelin 1 receptor type A (ETRa) and CC motif chemokine ligand 2 (CCL2) transcript production by SCD iSNs compared to HC iSNs (Table 1). Additionally, we observed robust responses with no significant differences in baseline function between HC and SCD iSNs when stimulated with known agonists of human SNs (KCl, Glutamate, ab-meATP, capsaicin) in traditional or high-throughput calcium imaging studies. To begin defining extrinsic mechanisms that may sensitize human SCD SNs, we tested the effects of SCD patient plasma (n=4 from SCD baseline [SS BL] and acute pain [SS IP]- paired samples, n=4 Black controls [HC]) on iSN hypersensitivity by exposing iSNs in vitro to human plasma diluted 1:10 in maintenance media for 30 minutes before analyzing calcium flux. Plasma samples from SCD patients experiencing acute pain (SS IP) were found to significantly increase SCD iSN response to capsaicin (p<0.05) compared to HC iSNs treated with SS IP plasma, while race-matched HC plasma or SCD patient plasma at baseline pain states (SS BL) did not induce this effect (Figure 1). Excitingly, these findings reproduce results identified in native DRG neurons from SCD mice and suggest that iSNs derived from patient iPSCs can be used in parallel with native SCD neurons from mouse models to investigate mechanisms underlying the chronic sensory neuron sensitization in SCD. They also identify potential intrinsic mechanisms of SCD pain which may extend beyond a blood-based disease pathology. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

Abstract P3134: Mechanosensitive Piezo2 Channel Mediates The Exaggerated Exercise Pressor Reflex In A Mouse Model Of Hind Limb Ischemia.

The exercise pressor reflex (EPR) is a reflexive mechanism in which blood pressure and heart rate increase in response to skeletal muscle contraction. These reflexive increases are caused by both mechanical and metabolic stimulation of group III and IV afferent neurons innervating in contracting skeletal muscle. Patients with peripheral artery disease (PAD) have an exaggerated EPR which contributes to exercise intolerance and morbidity and mortality. Recently, we developed a mouse model of EPR, and have validated that the mouse model is similar to our rat model. With this mouse model of EPR, we studied the EPR following femoral artery ligation (FAL), briefly the right hind limb common femoral artery was ligated and transected between the proximal end near the groin and the distal end near the bifurcation of the popliteal artery and the saphenous artery. The same procedure was done without femoral artery ligation and transection for sham control. We observed that the mean arterial pressure (MAP) responses to ventral root stimulation (which excite metabo- and mechano-sensitive afferent neurons) and passive stretch (mechanosensitive stimulation only) were exaggerated 72 hours following FA ligation when compared to sham treated controls. The exaggerated EPR was significantly reduced by intra-arterial administration of Gadolinium (a mechanical sensitive group III afferent neurons inhibitor), GsMTx-4 (a selective Piezo2 inhibitor), but not by SB-705498 (a TRPv1 receptor inhibitor). Additionally, selective stimulation of metabolically sensitive afferent neurons by intra-arterial capsaicin injection (an activator of metabolically sensitive afferent neurons) induced equal, dose-related increases in MAP in both FA ligation and control mice. Finally, immunofluorescence staining visualized that the mechano-sensitive channel Piezo2 is localized in most large diameter neuron cells in mouse lumbar DRG. First, these results indicate that metabolically sensitive afferent neurons are not affected by FAL. Second, we observed that the exaggerated EPR in a murine model of PAD is mediated by mechanically sensitive group III neurons, which is due, in part, to Piezo2 receptor activation. Future studies will be aimed at interrogation of the mechanosensitive mechanisms that mediate this abnormal response to exercise in a model of PAD. The goal of these studies will be to determine molecular targets through which the exaggerated EPR can be normalized to allow for the prescription of exercise in PAD patients.

Sensitization of colonic nociceptors by TNFα is dependent on TNFR1 expression and p38 MAPK activity.

Visceral pain is a leading cause of morbidity in gastrointestinal diseases, which is exacerbated by the gut‐related side‐effects of many analgesics. New treatments are needed and further understanding of the mediators and mechanisms underpinning visceral nociception in disease states is required to facilitate this. The pro‐inflammatory cytokine TNFα is linked to pain in both patients with inflammatory bowel disease and irritable bowel syndrome, and has been shown to sensitize colonic sensory neurons. Somatic, TNFα‐triggered thermal and mechanical hypersensitivity is mediated by TRPV1 signalling and p38 MAPK activity respectively, downstream of TNFR1 receptor activation. We therefore hypothesized that TNFR1‐evoked p38 MAPK activity may also be responsible for TNFα sensitization of colonic afferent responses to the TRPV1 agonist capsaicin, and noxious distension of the bowel. Using Ca2+ imaging of dorsal root ganglion sensory neurons, we observed TNFα‐mediated increases in intracellular [Ca2+] and sensitization of capsaicin responses. The sensitizing effects of TNFα were dependent on TNFR1 expression and attenuated by p38 MAPK inhibition. Consistent with these findings, ex vivo colonic afferent fibre recordings demonstrated an enhanced response to noxious ramp distention of the bowel and bath application of capsaicin following TNFα pre‐treatment. Responses were reversed by p38 MAPK inhibition and absent in tissue from TNFR1 knockout mice. Our findings demonstrate a contribution of TNFR1, p38 MAPK and TRPV1 to TNFα‐induced sensitization of colonic afferents, highlighting the potential utility of these drug targets for the treatment of visceral pain in gastrointestinal disease. Key points The pro‐inflammatory cytokine TNFα is elevated in gastrointestinal disease and sensitizes colonic afferents via modulation of TRPA1 and NaV1.8 activity. We further develop this understanding by demonstrating a role for p38 MAPK and TRPV1 in TNFα‐mediated colonic afferent sensitization. Specifically, we show that:TNFα sensitizes sensory neurons and colonic afferents to the TRPV1 agonist capsaicin.TNFα‐mediated sensitization of sensory neurons and colonic nociceptors is dependent on TNFR1 expression.TNFα sensitization of sensory neurons and colonic afferents to capsaicin and noxious ramp distension is abolished by inhibition of p38 MAPK.Collectively these data support the utility of targeting TNFα, TNFR1 and their downstream signalling via p38 MAPK for the treatment of visceral pain in gastrointestinal disease.

Decrypting the cellular and molecular intricacies associated with COVID-19-induced chronic pain.

Pain is one of the clinical manifestations that can vary from mild to severe symptoms in COVID-19 patients. Pain symptoms can be initiated by direct viral damage to the tissue or by indirect tissue injury followed by nociceptor sensitization. The most common types of pain that are reported to occur in COVID-19 patients are headache, myalgia, and chest pain. With more and more cases coming in the hospitals, many new and unique symptoms of pain are being reported. Testicular and abdominal pain are rare cases of pain that are also being reported and are associated with COVID-19. The SARS-CoV-2 virus has a high affinity for angiotensin-converting enzyme-2 receptor (ACE-2) which acts as an entry point for the virus. ACE-2/ Ang II/AT 1 receptor also participates directly in the transmission of pain signals from the dorsal horn of the spinal cord. It induces a series of complicated responses in the human body. Among which the cytokinetic storm and hypercoagulation are the most prominent pathways that mediate the sensitization of sensory neurons facilitating pain. The elevated immune response is also responsible for the activation of inflammatory lipid mediators such as COX-1 and COX-2 enzymes for the synthesis of prostaglandins (PGs). PG molecules especially PGE2 and PGD2 are involved in the pain transmission and are found to be elevated in COVID-19 patients. Though arachidonic acid pathway is one of the lesser discussed topics in COVID-19 pathophysiology, still it can be useful for explaining the unique and rarer symptoms of pain seen in COVID-19 patients. Understanding different pain pathways is very crucial for the management of pain and can help healthcare systems to end the current pandemic situation. We herein review the role of various molecules involved in the pain pathology of COVID-19.

Estrogen modulation of the pronociceptive effects of serotonin on female rat trigeminal sensory neurons is timing dependent and dosage dependent and requires estrogen receptor alpha.

The role of the major estrogen estradiol (E2) on orofacial pain conditions remains controversial with studies reporting both a pronociceptive and antinociceptive role of E2. E2 modulation of peripheral serotonergic activity may be one mechanism underlying the female prevalence of orofacial pain disorders. We recently reported that female rats in proestrus and estrus exhibit greater serotonin (5HT)-evoked orofacial nocifensive behaviors compared with diestrus and male rats. Further coexpression of 5HT 2A receptor mRNA in nociceptive trigeminal sensory neurons that express transient receptor potential vanilloid 1 ion channels contributes to pain sensitization. E2 may exacerbate orofacial pain through 5HT-sensitive trigeminal nociceptors, but whether low or high E2 contributes to orofacial pain and by what mechanism remains unclear. We hypothesized that steady-state exposure to a proestrus level of E2 exacerbates 5HT-evoked orofacial nocifensive behaviors in female rats, explored the transcriptome of E2-treated female rats, and determined which E2 receptor contributes to sensitization of female trigeminal sensory neurons. We report that a diestrus level of E2 is protective against 5HT-evoked orofacial pain behaviors, which increase with increasing E2 concentrations, and that E2 differentially alters several pain genes in the trigeminal ganglia. Furthermore, E2 receptors coexpressed with 5HT 2A and transient receptor potential vanilloid 1 and enhanced capsaicin-evoked signaling in the trigeminal ganglia through estrogen receptor α. Overall, our data indicate that low, but not high, physiological levels of E2 protect against orofacial pain, and we provide evidence that estrogen receptor α receptor activation, but not others, contributes to sensitization of nociceptive signaling in trigeminal sensory neurons.

Bladder infection with uropathogenic Escherichia coli increases the excitability of afferent neurons.

Urinary tract infections (UTIs) cause bladder hyperactivity and pelvic pain, but the underlying causes of these symptoms remain unknown. We investigated whether afferent sensitization contributes to the bladder overactivity and pain observed in mice suffering from experimentally induced bacterial cystitis. Inoculation of mouse bladders with the uropathogenic Escherichia coli strain UTI89 caused pelvic allodynia, increased voiding frequency, and prompted an acute inflammatory process marked by leukocytic infiltration and edema of the mucosa. Compared with controls, isolated bladder sensory neurons from UTI-treated mice exhibited a depolarized resting membrane potential, lower action potential threshold and rheobase, and increased firing in response to suprathreshold stimulation. To determine whether bacterial virulence factors can contribute to the sensitization of bladder afferents, neurons isolated from naïve mice were incubated with supernatants collected from bacterial cultures with or depleted of lipopolysaccharide (LPS). Supernatants containing LPS prompted the sensitization of bladder sensory neurons with both tetrodotoxin (TTX)-resistant and TTX-sensitive action potentials. However, bladder sensory neurons with TTX-sensitive action potentials were not affected by bacterial supernatants depleted of LPS. Unexpectedly, ultrapure LPS increased the excitability only of bladder sensory neurons with TTX-resistant action potentials, but the supplementation of supernatants depleted of LPS with ultrapure LPS resulted in the sensitization of both population of bladder sensory neurons. In summary, the results of our study indicate that multiple virulence factors released from UTI89 act on bladder sensory neurons to prompt their sensitization. These sensitized bladder sensory neurons mediate, at least in part, the bladder hyperactivity and pelvic pain seen in mice inoculated with UTI89.NEW & NOTEWORTHY Urinary tract infection (UTI) produced by uropathogenic Escherichia coli (UPEC) promotes sensitization of bladder afferent sensory neurons with tetrodotoxin-resistant and tetrodotoxin-sensitive action potentials. Lipopolysaccharide and other virulence factors produced by UPEC contribute to the sensitization of bladder afferents in UTI. In conclusion, sensitized afferents contribute to the voiding symptoms and pelvic pain present in mice bladder inoculated with UPEC.

Investigating the Function of Adult DRG Neuron Axons Using an In Vitro Microfluidic Culture System.

A critical role of the peripheral axons of nociceptors of the dorsal root ganglion (DRG) is the conduction of all-or-nothing action potentials from peripheral nerve endings to the central nervous system for the perception of noxious stimuli. Plasticity along multiple sites along the pain axis has now been widely implicated in the maladaptive changes that occur in pathological pain states such as neuropathic and inflammatory pain. Notably, increasing evidence suggests that nociceptive axons actively participate through the local expression of ion channels, receptors, and signal transduction molecules through axonal mRNA translation machinery that is independent of the soma component. In this report, we explore the sensitization of sensory neurons through the treatment of compartmentalized axon-like structures spanning microchannels that have been treated with the cytokine IL-6 and, subsequently, capsaicin. These data demonstrate the utility of isolating DRG axon-like structures using microfluidic systems, laying the groundwork for constructing the complex in vitro models of cellular networks that are involved in pain signaling for targeted pharmacological and genetic perturbations.

The Calcium Channel α2δ1 Subunit: Interactional Targets in Primary Sensory Neurons and Role in Neuropathic Pain.

Neuropathic pain is mainly triggered after nerve injury and associated with plasticity of the nociceptive pathway in primary sensory neurons. Currently, the treatment remains a challenge. In order to identify specific therapeutic targets, it is necessary to clarify the underlying mechanisms of neuropathic pain. It is well established that primary sensory neuron sensitization (peripheral sensitization) is one of the main components of neuropathic pain. Calcium channels act as key mediators in peripheral sensitization. As the target of gabapentin, the calcium channel subunit α2δ1 (Cavα2δ1) is a potential entry point in neuropathic pain research. Numerous studies have demonstrated that the upstream and downstream targets of Cavα2δ1 of the peripheral primary neurons, including thrombospondins, N-methyl-D-aspartate receptors, transient receptor potential ankyrin 1 (TRPA1), transient receptor potential vanilloid family 1 (TRPV1), and protein kinase C (PKC), are involved in neuropathic pain. Thus, we reviewed and discussed the role of Cavα2δ1 and the associated signaling axis in neuropathic pain conditions.

Sensitivity of olfactory sensory neurons to food cues is tuned to nutritional states by Neuropeptide Y signaling

Modulation of sensory perception by homeostatic feedback from physiological states is central to innate purposive behaviors. Olfaction is an important predictive modality for feeding-related behaviors and its modulation has been associated with hunger-satiety states. However, the mechanisms mapping internal states to chemosensory processing in order to modify behavior are poorly understood. In the zebrafish olfactory epithelium, a subset of olfactory sensory neurons (OSNs) and the terminal nerve projections express neuropeptide Y (NPY). Using a combination of neuronal activity and behavioral evaluation, we find that NPY signaling in the peripheral olfactory system of zebrafish is correlated with its nutritional state and is both necessary and sufficient for the olfactory perception of food-related odorants. NPY activity dynamically modulates the microvillar OSN activation thresholds and acts cooperatively with amino acid signaling resulting in a switch-like increase in OSN sensitivity in starved animals. We suggest that cooperative activation of phospholipase C by convergent signaling from NPY and amino acid receptors is central to this heightened sensitivity. This study provides ethologically relevant, physiological evidence for NPY signaling in the modulation of OSN sensitivity to food-associated amino acid cues. We demonstrate sensory gating directly at the level of OSNs and identify a novel mechanistic framework for tuning olfactory sensitivity to prevailing energy states. Cover Image for this issue:https://doi.org/10.1111/jnc.15091.

Holdemanella biformis improves glucose tolerance and regulates GLP-1 signaling in obese mice.

Impaired glucose homeostasis in obesity is mitigated by enhancing the glucoregulatory actions of glucagon-like peptide 1 (GLP-1), and thus, strategies that improve GLP-1 sensitivity and secretion have therapeutic potential for the treatment of type 2 diabetes. This study shows that Holdemanella biformis, isolated from the feces of a metabolically healthy volunteer, ameliorates hyperglycemia, improves oral glucose tolerance and restores gluconeogenesis and insulin signaling in the liver of obese mice. These effects were associated with the ability of H. biformis to restore GLP-1 levels, enhancing GLP-1 neural signaling in the proximal and distal small intestine and GLP-1 sensitivity of vagal sensory neurons, and to modify the cecal abundance of unsaturated fatty acids and the bacterial species associated with metabolic health. Our findings overall suggest the potential use of H biformis in the management of type 2 diabetes in obesity to optimize the sensitivity and function of the GLP-1 system, through direct and indirect mechanisms.