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1.
PLoS Comput Biol ; 15(5): e1007033, 2019 05.
Artigo em Inglês | MEDLINE | ID: mdl-31107861

RESUMO

G protein-coupled receptors (GPCRs) control cellular signaling and responses. Many of these GPCRs are modulated by cholesterol and polyunsaturated fatty acids (PUFAs) which have been shown to co-exist with saturated lipids in ordered membrane domains. However, the lipid compositions of such domains extracted from the brain cortex tissue of individuals suffering from GPCR-associated neurological disorders show drastically lowered levels of PUFAs. Here, using free energy techniques and multiscale simulations of numerous membrane proteins, we show that the presence of the PUFA DHA helps helical multi-pass proteins such as GPCRs partition into ordered membrane domains. The mechanism is based on hybrid lipids, whose PUFA chains coat the rough protein surface, while the saturated chains face the raft environment, thus minimizing perturbations therein. Our findings suggest that the reduction of GPCR partitioning to their native ordered environments due to PUFA depletion might affect the function of these receptors in numerous neurodegenerative diseases, where the membrane PUFA levels in the brain are decreased. We hope that this work inspires experimental studies on the connection between membrane PUFA levels and GPCR signaling.


Assuntos
Ácidos Docosa-Hexaenoicos/metabolismo , Receptores Acoplados a Proteínas-G/metabolismo , Células Receptoras Sensoriais/metabolismo , Encéfalo/metabolismo , Colesterol/metabolismo , Biologia Computacional , Simulação por Computador , Ácidos Docosa-Hexaenoicos/química , Ácidos Graxos Insaturados/metabolismo , Humanos , Microdomínios da Membrana/química , Microdomínios da Membrana/metabolismo , Proteínas de Membrana/química , Proteínas de Membrana/metabolismo , Modelos Moleculares , Modelos Neurológicos , Conformação Proteica , Receptor A2A de Adenosina/química , Receptor A2A de Adenosina/metabolismo , Receptores Acoplados a Proteínas-G/química , Células Receptoras Sensoriais/química , Transdução de Sinais , Termodinâmica
2.
Int J Mol Sci ; 20(9)2019 Apr 30.
Artigo em Inglês | MEDLINE | ID: mdl-31052288

RESUMO

A critical aim in neuroscience is to obtain a comprehensive view of how regulated neurotransmission is achieved. Our current understanding of synapses relies mainly on data from electrophysiological recordings, imaging, and molecular biology. Based on these methodologies, proteins involved in a synaptic vesicle (SV) formation, mobility, and fusion at the active zone (AZ) membrane have been identified. In the last decade, electron tomography (ET) combined with a rapid freezing immobilization of neuronal samples opened a window for understanding the structural machinery with the highest spatial resolution in situ. ET provides significant insights into the molecular architecture of the AZ and the organelles within the presynaptic nerve terminal. The specialized sensory ribbon synapses exhibit a distinct architecture from neuronal synapses due to the presence of the electron-dense synaptic ribbon. However, both synapse types share the filamentous structures, also commonly termed as tethers that are proposed to contribute to different steps of SV recruitment and exocytosis. In this review, we discuss the emerging views on the role of filamentous structures in SV exocytosis gained from ultrastructural studies of excitatory, mainly central neuronal compared to ribbon-type synapses with a focus on inner hair cell (IHC) ribbon synapses. Moreover, we will speculate on the molecular entities that may be involved in filament formation and hence play a crucial role in the SV cycle.


Assuntos
Citoesqueleto/metabolismo , Exocitose , Células Receptoras Sensoriais/metabolismo , Membranas Sinápticas/metabolismo , Vesículas Sinápticas/metabolismo , Animais , Citoesqueleto/ultraestrutura , Humanos , Células Receptoras Sensoriais/ultraestrutura , Membranas Sinápticas/ultraestrutura , Transmissão Sináptica , Vesículas Sinápticas/ultraestrutura
3.
Genetics ; 212(1): 25-51, 2019 05.
Artigo em Inglês | MEDLINE | ID: mdl-31053616

RESUMO

Caenorhabditis elegans lives in a complex habitat in which they routinely experience large fluctuations in temperature, and encounter physical obstacles that vary in size and composition. Their habitat is shared by other nematodes, by beneficial and harmful bacteria, and nematode-trapping fungi. Not surprisingly, these nematodes can detect and discriminate among diverse environmental cues, and exhibit sensory-evoked behaviors that are readily quantifiable in the laboratory at high resolution. Their ability to perform these behaviors depends on <100 sensory neurons, and this compact sensory nervous system together with powerful molecular genetic tools has allowed individual neuron types to be linked to specific sensory responses. Here, we describe the sensory neurons and molecules that enable C. elegans to sense and respond to physical stimuli. We focus primarily on the pathways that allow sensation of mechanical and thermal stimuli, and briefly consider this animal's ability to sense magnetic and electrical fields, light, and relative humidity. As the study of sensory transduction is critically dependent upon the techniques for stimulus delivery, we also include a section on appropriate laboratory methods for such studies. This chapter summarizes current knowledge about the sensitivity and response dynamics of individual classes of C. elegans mechano- and thermosensory neurons from in vivo calcium imaging and whole-cell patch-clamp electrophysiology studies. We also describe the roles of conserved molecules and signaling pathways in mediating the remarkably sensitive responses of these nematodes to mechanical and thermal cues. These studies have shown that the protein partners that form mechanotransduction channels are drawn from multiple superfamilies of ion channel proteins, and that signal transduction pathways responsible for temperature sensing in C. elegans share many features with those responsible for phototransduction in vertebrates.


Assuntos
Caenorhabditis elegans/fisiologia , Mecanotransdução Celular , Células Receptoras Sensoriais/fisiologia , Transdução de Sinais , Estresse Fisiológico , Sensação Térmica , Animais , Caenorhabditis elegans/metabolismo , Fenômenos Mecânicos , Sensação , Células Receptoras Sensoriais/metabolismo , Temperatura Ambiente
4.
Int J Mol Sci ; 20(8)2019 Apr 22.
Artigo em Inglês | MEDLINE | ID: mdl-31013625

RESUMO

The neuron-specific Elav-like Hu RNA-binding proteins were described to play an important role in neuronal differentiation and plasticity by ensuring the post-transcriptional control of RNAs encoding for various proteins. Although Elav-like Hu proteins alterations were reported in diabetes or neuropathy, little is known about the regulation of neuron-specific Elav-like Hu RNA-binding proteins in sensory neurons of dorsal root ganglia (DRG) due to the diabetic condition. The goal of our study was to analyze the gene and protein expression of HuB, HuC, and HuD in DRG sensory neurons in diabetes. The diabetic condition was induced in CD-1 adult male mice with single-intraperitoneal injection of streptozotocin (STZ, 150 mg/kg), and 8-weeks (advanced diabetes) after induction was quantified the Elav-like proteins expression. Based on the glycemia values, we identified two types of responses to STZ, and mice were classified in STZ-resistant (diabetic resistant, glycemia < 260 mg/dL) and STZ-sensitive (diabetic, glycemia > 260 mg/dL). Body weight measurements indicated that 8-weeks after STZ-induction of diabetes, control mice have a higher increase in body weight compared to the diabetic and diabetic resistant mice. Moreover, after 8-weeks, diabetic mice (19.52 ± 3.52 s) have longer paw withdrawal latencies in the hot-plate test than diabetic resistant (11.36 ± 1.92 s) and control (11.03 ± 1.97 s) mice, that correlates with the installation of warm hypoalgesia due to the diabetic condition. Further on, we evidenced the decrease of Elav-like gene expression in DRG neurons of diabetic mice (Elavl2, 0.68 ± 0.05 fold; Elavl3, 0.65 ± 0.01 fold; Elavl4, 0.53 ± 0.07 fold) and diabetic resistant mice (Ealvl2, 0.56 ± 0.07 fold; Elavl3, 0.32 ± 0.09 fold) compared to control mice. Interestingly, Elav-like genes have a more accentuated downregulation in diabetic resistant than in diabetic mice, although hypoalgesia was evidenced only in diabetic mice. The Elav-like gene expression changes do not always correlate with the Hu protein expression changes. To detail, HuB is upregulated and HuD is downregulated in diabetic mice, while HuB, HuC, and HuD are downregulated in diabetic resistant mice compared to control mice. To resume, we demonstrated HuD downregulation and HuB upregulation in DRG sensory neurons induced by diabetes, which might be correlated with altered post-transcriptional control of RNAs involved in the regulation of thermal hypoalgesia condition caused by the advanced diabetic neuropathy.


Assuntos
Proteína Semelhante a ELAV 2/genética , Proteína Semelhante a ELAV 3/genética , Proteína Semelhante a ELAV 4/genética , Gânglios Espinais/citologia , Gânglios Espinais/metabolismo , Regulação da Expressão Gênica , Células Receptoras Sensoriais/metabolismo , Animais , Biomarcadores , Glicemia , Peso Corporal , Diabetes Mellitus Experimental , Proteína Semelhante a ELAV 2/metabolismo , Proteína Semelhante a ELAV 3/metabolismo , Proteína Semelhante a ELAV 4/metabolismo , Gânglios Espinais/fisiopatologia , Imuno-Histoquímica , Camundongos , Proteínas de Ligação a RNA
5.
Int J Mol Sci ; 20(6)2019 Mar 24.
Artigo em Inglês | MEDLINE | ID: mdl-30909643

RESUMO

An exaggerated exercise pressor reflex (EPR) is associated with excessive sympatho-excitation and exercise intolerance in the chronic heart failure (CHF) state. We hypothesized that brain-derived neurotrophic factor (BDNF) causes the exaggerated EPR via sensitizing muscle mechanosensitive afferents in CHF. Increased BDNF expression was observed in lumbar dorsal root ganglia (DRGs) from CHF rats compared to sham rats. Immunofluorescence data showed a greater increase in the number of BDNF-positive neurons in medium and large-sized DRG subpopulations from CHF rats. Patch clamp data showed that incubation with BDNF for 4⁻6 h, significantly decreased the current threshold-inducing action potential (AP), threshold potential and the number of APs during current injection in Dil-labeled isolectin B4 (IB4)-negative medium-sized DRG neurons (mainly mechano-sensitive) from sham rats. Compared to sham rats, CHF rats exhibited an increased number of APs during current injection in the same DRG subpopulation, which was significantly attenuated by 4-h incubation with anti-BDNF. Finally, chronic epidural delivery of anti-BDNF attenuated the exaggerated pressor response to either static contraction or passive stretch in CHF rats whereas this intervention had no effect on the pressor response to hindlimb arterial injection of capsaicin. These data suggest that increased BDNF in lumbar DRGs contributes to the exaggerated EPR in CHF.


Assuntos
Fator Neurotrófico Derivado do Encéfalo/metabolismo , Gânglios Espinais/metabolismo , Insuficiência Cardíaca/etiologia , Insuficiência Cardíaca/fisiopatologia , Condicionamento Físico Animal , Reflexo , Animais , Anticorpos Monoclonais/farmacologia , Peso Corporal , Fator Neurotrófico Derivado do Encéfalo/antagonistas & inibidores , Fator Neurotrófico Derivado do Encéfalo/genética , Modelos Animais de Doenças , Insuficiência Cardíaca/diagnóstico , Hemodinâmica , Imuno-Histoquímica , Região Lombossacral , Masculino , Tamanho do Órgão , Ratos , Reflexo/efeitos dos fármacos , Células Receptoras Sensoriais/metabolismo
6.
Dev Cell ; 48(6): 864-872.e7, 2019 03 25.
Artigo em Inglês | MEDLINE | ID: mdl-30827898

RESUMO

Dynamic coupling of microtubule ends to kinetochores, built on the centromeres of chromosomes, directs chromosome segregation during cell division. Here, we report that the evolutionarily ancient kinetochore-microtubule coupling machine, the KMN (Knl1/Mis12/Ndc80-complex) network, plays a critical role in neuronal morphogenesis. We show that the KMN network concentrates in microtubule-rich dendrites of developing sensory neurons that collectively extend in a multicellular morphogenetic event that occurs during C. elegans embryogenesis. Post-mitotic degradation of KMN components in sensory neurons disrupts dendritic extension, leading to patterning and functional defects in the sensory nervous system. Structure-guided mutations revealed that the molecular interface that couples kinetochores to spindle microtubules also functions in neuronal development. These results identify a cell-division-independent function for the chromosome-segregation machinery and define a microtubule-coupling-dependent event in sensory nervous system morphogenesis.


Assuntos
Cinetocoros/metabolismo , Microtúbulos/metabolismo , Morfogênese , Sistema Nervoso/embriologia , Sistema Nervoso/metabolismo , Células Receptoras Sensoriais/metabolismo , Animais , Caenorhabditis elegans/embriologia , Proteínas de Caenorhabditis elegans/metabolismo , Dendritos/metabolismo , Embrião não Mamífero/metabolismo , Desenvolvimento Embrionário , Mitose
7.
Neuron ; 102(2): 373-389.e6, 2019 04 17.
Artigo em Inglês | MEDLINE | ID: mdl-30819546

RESUMO

Neurons exhibit a limited ability of repair. Given that mechanical forces affect neuronal outgrowth, it is important to investigate whether mechanosensitive ion channels may regulate axon regeneration. Here, we show that DmPiezo, a Ca2+-permeable non-selective cation channel, functions as an intrinsic inhibitor for axon regeneration in Drosophila. DmPiezo activation during axon regeneration induces local Ca2+ transients at the growth cone, leading to activation of nitric oxide synthase and the downstream cGMP kinase Foraging or PKG to restrict axon regrowth. Loss of DmPiezo enhances axon regeneration of sensory neurons in the peripheral and CNS. Conditional knockout of its mammalian homolog Piezo1 in vivo accelerates regeneration, while its pharmacological activation in vitro modestly reduces regeneration, suggesting the role of Piezo in inhibiting regeneration may be evolutionarily conserved. These findings provide a precedent for the involvement of mechanosensitive channels in axon regeneration and add a potential target for modulating nervous system repair.


Assuntos
Axônios/fisiologia , Proteínas de Drosophila/genética , Canais Iônicos/genética , Regeneração/genética , Animais , Cálcio/metabolismo , Proteínas Quinases Dependentes de GMP Cíclico/metabolismo , Proteínas de Drosophila/metabolismo , Drosophila melanogaster , Cones de Crescimento/metabolismo , Canais Iônicos/metabolismo , Mecanotransdução Celular/genética , Camundongos , Camundongos Knockout , Regeneração Nervosa/genética , Óxido Nítrico Sintase/metabolismo , Células Receptoras Sensoriais/metabolismo , Células Receptoras Sensoriais/fisiologia
8.
Neuron ; 102(2): 420-434.e8, 2019 04 17.
Artigo em Inglês | MEDLINE | ID: mdl-30826183

RESUMO

Presynaptic inhibition (PSI) of primary sensory neurons is implicated in controlling gain and acuity in sensory systems. Here, we define circuit mechanisms and functions of PSI of cutaneous somatosensory neuron inputs to the spinal cord. We observed that PSI can be evoked by different sensory neuron populations and mediated through at least two distinct dorsal horn circuit mechanisms. Low-threshold cutaneous afferents evoke a GABAA-receptor-dependent form of PSI that inhibits similar afferent subtypes, whereas small-diameter afferents predominantly evoke an NMDA-receptor-dependent form of PSI that inhibits large-diameter fibers. Behaviorally, loss of either GABAA receptors (GABAARs) or NMDA receptors (NMDARs) in primary afferents leads to tactile hypersensitivity across skin types, and loss of GABAARs, but not NMDARs, leads to impaired texture discrimination. Post-weaning age loss of either GABAARs or NMDARs in somatosensory neurons causes systemic behavioral abnormalities, revealing critical roles of two distinct modes of PSI of somatosensory afferents in adolescence and throughout adulthood.


Assuntos
Comportamento Animal , Proteínas do Tecido Nervoso/genética , Inibição Neural , Terminações Pré-Sinápticas/metabolismo , Receptores de GABA-A/genética , Receptores de N-Metil-D-Aspartato/genética , Células Receptoras Sensoriais/metabolismo , Pele/inervação , Tato , Animais , Bicuculina/análogos & derivados , Bicuculina/farmacologia , Discriminação (Psicologia) , Antagonistas de Receptores de GABA-A/farmacologia , Hiperestesia/genética , Camundongos , Proteínas do Tecido Nervoso/metabolismo , Neurônios Aferentes/metabolismo , Piridazinas/farmacologia , Receptores de GABA-A/metabolismo , Receptores de N-Metil-D-Aspartato/metabolismo , Corno Dorsal da Medula Espinal
9.
Front Neural Circuits ; 13: 14, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30894803

RESUMO

Little is known about the molecular and cellular mechanisms involved in the formation of the cranial peripheral sensory system in vertebrates. To identify genes involved in the formation of these circuits, we performed a forward genetic screen utilizing a transgenic zebrafish line (p2rx3.2:gfp sl1) that expresses green fluorescent protein (gfp) in sensory neurons of the Vth, VIIth, IXth and Xth cranial ganglia. Here, we describe a novel zebrafish mutant in which a missense mutation in the adam19b gene selectively affects the epibranchial sensory circuits.


Assuntos
Proteínas ADAM/metabolismo , Orientação de Axônios/fisiologia , Rombencéfalo/citologia , Rombencéfalo/fisiologia , Células Receptoras Sensoriais/metabolismo , Proteínas ADAM/genética , Animais , Animais Geneticamente Modificados , Orientação de Axônios/genética , Proteína 9 Associada à CRISPR/genética , Proteína 9 Associada à CRISPR/metabolismo , Repetições Palindrômicas Curtas Agrupadas e Regularmente Espaçadas , Análise Mutacional de DNA , Embrião não Mamífero , Gânglios dos Invertebrados/citologia , Regulação da Expressão Gênica no Desenvolvimento/fisiologia , Proteínas de Fluorescência Verde/genética , Proteínas de Fluorescência Verde/metabolismo , Larva , Mutação/genética , Receptores Purinérgicos P2/genética , Receptores Purinérgicos P2/metabolismo , Peixe-Zebra , Proteínas de Peixe-Zebra/genética , Proteínas de Peixe-Zebra/metabolismo
10.
Can J Physiol Pharmacol ; 97(5): 400-406, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-30712368

RESUMO

Low-power (non-thermal) infrared (IR) radiation with the wavelength of 10.6 µm activates the Na,K-ATPase transducer function in sensory neurons, which is manifested in decrease of NaV1.8 channel voltage sensitivity at the cellular membrane level and in inhibition of growth of chick embryo dorsal root ganglia neurites at the tissue level. It is shown that the effect of low-power IR radiation is totally blocked by a specific Src kinase inhibitor, PP2. Upon irradiation on the background of PP2, the effective charge of NaV1.8 channel activation gating system does not differ from its control value in patch-clamp experiments, and the area index of sensory ganglia neurites growth remains unchanged as compared with the control in organotypic tissue culture. The data obtained demonstrate that Src kinase is involved in intracellular signaling pathways triggered by CO2 laser low-power IR radiation by the transducer-activated mechanism. This is the first indication that in primary sensory neuron the signals of low-power IR radiation are sensed, amplified, and transduced by the Na,K-ATPase/Src complex and not by G proteins.


Assuntos
Raios Infravermelhos , Células Receptoras Sensoriais/citologia , Células Receptoras Sensoriais/efeitos da radiação , Transdução de Sinais/efeitos da radiação , Quinases da Família src/metabolismo , Animais , Relação Dose-Resposta à Radiação , Ratos , Ratos Wistar , Células Receptoras Sensoriais/metabolismo , ATPase Trocadora de Sódio-Potássio/metabolismo
11.
Proc Natl Acad Sci U S A ; 116(11): 5126-5134, 2019 03 12.
Artigo em Inglês | MEDLINE | ID: mdl-30804200

RESUMO

Sensory neurons perceive environmental cues and are important of organismal survival. Peripheral sensory neurons interact intimately with glial cells. While the function of axonal ensheathment by glia is well studied, less is known about the functional significance of glial interaction with the somatodendritic compartment of neurons. Herein, we show that three distinct glia cell types differentially wrap around the axonal and somatodendritic surface of the polymodal dendritic arborization (da) neuron of the Drosophila peripheral nervous system for detection of thermal, mechanical, and light stimuli. We find that glial cell-specific loss of the chromatin modifier gene dATRX in the subperineurial glial layer leads to selective elimination of somatodendritic glial ensheathment, thus allowing us to investigate the function of such ensheathment. We find that somatodendritic glial ensheathment regulates the morphology of the dendritic arbor, as well as the activity of the sensory neuron, in response to sensory stimuli. Additionally, glial ensheathment of the neuronal soma influences dendritic regeneration after injury.


Assuntos
Dendritos/metabolismo , Drosophila melanogaster/metabolismo , Neuroglia/metabolismo , Células Receptoras Sensoriais/citologia , Células Receptoras Sensoriais/metabolismo , Animais , Axônios/metabolismo , Axônios/efeitos da radiação , Caspases/metabolismo , DNA Helicases/metabolismo , Dendritos/efeitos da radiação , Proteínas de Drosophila/metabolismo , Ativação Enzimática/efeitos da radiação , Luz , Neuroglia/efeitos da radiação , Células Receptoras Sensoriais/efeitos da radiação
12.
Proc Natl Acad Sci U S A ; 116(11): 5135-5143, 2019 03 12.
Artigo em Inglês | MEDLINE | ID: mdl-30804203

RESUMO

Aggression is controlled by the olfactory system in many animal species. In male mice, territorial and infant-directed aggression are tightly regulated by the vomeronasal organ (VNO), but how diverse subsets of sensory neurons convey pheromonal information to limbic centers is not yet known. Here, we employ genetic strategies to show that mouse vomeronasal sensory neurons expressing the G protein subunit Gαi2 regulate male-male and infant-directed aggression through distinct circuit mechanisms. Conditional ablation of Gαi2 enhances male-male aggression and increases neural activity in the medial amygdala (MeA), bed nucleus of the stria terminalis, and lateral septum. By contrast, conditional Gαi2 ablation causes reduced infant-directed aggression and decreased activity in MeA neurons during male-infant interactions. Strikingly, these mice also display enhanced parental behavior and elevated neural activity in the medial preoptic area, whereas sexual behavior remains normal. These results identify Gαi2 as the primary G protein α-subunit mediating the detection of volatile chemosignals in the apical layer of the VNO, and they show that Gαi2+ VSNs and the brain circuits activated by these neurons play a central role in orchestrating and balancing territorial and infant-directed aggression of male mice through bidirectional activation and inhibition of different targets in the limbic system.


Assuntos
Agressão , Subunidade alfa Gi2 de Proteína de Ligação ao GTP/metabolismo , Células Receptoras Sensoriais/metabolismo , Territorialidade , Órgão Vomeronasal/metabolismo , Animais , Animais Recém-Nascidos , Biomarcadores/metabolismo , Encéfalo/fisiologia , Mapeamento Encefálico , Feminino , Deleção de Genes , Masculino , Camundongos Endogâmicos C57BL , Mutação/genética , Comportamento Sexual Animal
13.
Mol Pharmacol ; 95(5): 519-527, 2019 05.
Artigo em Inglês | MEDLINE | ID: mdl-30808671

RESUMO

Muscle ischemia, associated with peripheral artery disease (PAD), leads to the release of proinflammatory mediators that decrease extracellular pH and trigger the activation of proton-activated acid-sensing ion channels (ASIC). Claudication pain, linked with low blood flow, can be partially relieved by endogenous opioid peptide release. However, we previously reported that sustained ASIC currents in dorsal root ganglion (DRG) neurons were enhanced by naturally occurring endomorphin-1 and -2 opioid peptides, indicating a role of opioid involvement in hyperalgesia. The present study examined whether clinically employed synthetic (fentanyl, remifentanil) and the semisynthetic opioid (oxycodone) would also potentiate sustained ASIC currents, which arise from ASIC3 channel isoforms. Here, we show that exposure of each opioid to DRG neurons resulted in potentiation of the sustained ASIC currents. On the other hand, the potentiation was not observed in DRG neurons from ASIC3 knockout rats. Further, the enhancement of the ASIC currents was resistant to pertussis toxin treatment, suggesting that Gα i/Gα o G-proteins are not involved. Additionally, the potentiation of sustained ASIC currents was greater in DRG neurons isolated from rats with ligated femoral arteries (a model of PAD). The effect of all three opioids on the transient ASIC peak current was mixed (increase, decrease, no effect). The inhibitory action appears to be mediated by the presence of ASIC1 isoform, while the potentiating effect is primarily due to ASIC3 isoform expression. These findings reveal that, under certain conditions, these three opioids can increase ASIC channel activity, possibly giving rise to opioid-induced hyperalgesia.


Assuntos
Canais Iônicos Sensíveis a Ácido/metabolismo , Potenciais de Ação/efeitos dos fármacos , Analgésicos Opioides/farmacologia , Células Receptoras Sensoriais/efeitos dos fármacos , Animais , Artéria Femoral/efeitos dos fármacos , Artéria Femoral/metabolismo , Proteínas de Ligação ao GTP/metabolismo , Gânglios Espinais/efeitos dos fármacos , Gânglios Espinais/metabolismo , Masculino , Oligopeptídeos/metabolismo , Dor/tratamento farmacológico , Dor/metabolismo , Isoformas de Proteínas/metabolismo , Ratos , Ratos Sprague-Dawley , Células Receptoras Sensoriais/metabolismo
14.
PLoS Genet ; 15(2): e1007940, 2019 02.
Artigo em Inglês | MEDLINE | ID: mdl-30730884

RESUMO

Members of the Ski/Sno protein family are classified as proto-oncogenes and act as negative regulators of the TGF-ß/BMP-pathways in vertebrates and invertebrates. A newly identified member of this protein family is fussel (fuss), the Drosophila homologue of the human functional Smad suppressing elements (fussel-15 and fussel-18). We and others have shown that Fuss interacts with SMAD4 and that overexpression leads to a strong inhibition of Dpp signaling. However, to be able to characterize the endogenous Fuss function in Drosophila melanogaster, we have generated a number of state of the art tools including anti-Fuss antibodies, specific fuss-Gal4 lines and fuss mutant fly lines via the CRISPR/Cas9 system. Fuss is a predominantly nuclear, postmitotic protein, mainly expressed in interneurons and fuss mutants are fully viable without any obvious developmental phenotype. To identify potential target genes or cells affected in fuss mutants, we conducted targeted DamID experiments in adult flies, which revealed the function of fuss in bitter gustatory neurons. We fully characterized fuss expression in the adult proboscis and by using food choice assays we were able to show that fuss mutants display defects in detecting bitter compounds. This correlated with a reduction of gustatory receptor gene expression (Gr33a, Gr66a, Gr93a) providing a molecular link to the behavioral phenotype. In addition, Fuss interacts with Rpd3, and downregulation of rpd3 in gustatory neurons phenocopies the loss of Fuss expression. Surprisingly, there is no colocalization of Fuss with phosphorylated Mad in the larval central nervous system, excluding a direct involvement of Fuss in Dpp/BMP signaling. Here we provide a first and exciting link of Fuss function in gustatory bitter neurons. Although gustatory receptors have been well characterized, little is known regarding the differentiation and maturation of gustatory neurons. This work therefore reveals Fuss as a pivotal element for the proper differentiation of bitter gustatory neurons acting within a chromatin modifying complex.


Assuntos
Proteínas de Drosophila/genética , Drosophila melanogaster/genética , Histona Desacetilase 1/genética , Proteínas do Tecido Nervoso/genética , Proteínas Proto-Oncogênicas/genética , Receptores de Superfície Celular/genética , Animais , Animais Geneticamente Modificados , Proteínas Morfogenéticas Ósseas/metabolismo , Diferenciação Celular , Cromatina/genética , Cromatina/metabolismo , Proteínas de Drosophila/metabolismo , Drosophila melanogaster/crescimento & desenvolvimento , Drosophila melanogaster/metabolismo , Feminino , Genes de Insetos , Histona Desacetilase 1/metabolismo , Masculino , Mutação , Proteínas do Tecido Nervoso/metabolismo , Neurônios/citologia , Neurônios/metabolismo , Proteínas Proto-Oncogênicas/metabolismo , Receptores de Superfície Celular/metabolismo , Células Receptoras Sensoriais/citologia , Células Receptoras Sensoriais/metabolismo , Transdução de Sinais , Paladar/genética
15.
J Biol Chem ; 294(14): 5496-5507, 2019 04 05.
Artigo em Inglês | MEDLINE | ID: mdl-30745360

RESUMO

α-Melanocyte-stimulating hormone (α-MSH) has been shown to be involved in nociception, but the underlying molecular mechanisms remain largely unknown. In this study, we report that α-MSH suppresses the transient outward A-type K+ current (I A) in trigeminal ganglion (TG) neurons and thereby modulates neuronal excitability and peripheral pain sensitivity in rats. Exposing small-diameter TG neurons to α-MSH concentration-dependently decreased I A This α-MSH-induced I A decrease was dependent on the melanocortin type 4 receptor (MC4R) and associated with a hyperpolarizing shift in the voltage dependence of A-type K+ channel inactivation. Chemical inhibition of phosphatidylinositol 3-kinase (PI3K) with wortmannin or of class I PI3Ks with the selective inhibitor CH5132799 prevented the MC4R-mediated I A response. Blocking Gi/o-protein signaling with pertussis toxin or by dialysis of TG neurons with the Gßγ-blocking synthetic peptide QEHA abolished the α-MSH-mediated decrease in I A Further, α-MSH increased the expression levels of phospho-p38 mitogen-activated protein kinase, and pharmacological or genetic inhibition of p38α abrogated the α-MSH-induced I A response. Additionally, α-MSH significantly increased the action potential firing rate of TG neurons and increased the sensitivity of rats to mechanical stimuli applied to the buccal pad area, and both effects were abrogated by I A blockade. Taken together, our findings suggest that α-MSH suppresses I A by activating MC4R, which is coupled sequentially to the Gßγ complex of the Gi/o-protein and downstream class I PI3K-dependent p38α signaling, thereby increasing TG neuronal excitability and mechanical pain sensitivity in rats.


Assuntos
Potenciais de Ação/efeitos dos fármacos , Dor/metabolismo , Canais de Potássio/metabolismo , Receptor Tipo 4 de Melanocortina/metabolismo , Células Receptoras Sensoriais/metabolismo , Transdução de Sinais/efeitos dos fármacos , Gânglio Trigeminal/metabolismo , alfa-MSH/farmacologia , Animais , Proteínas de Ligação ao GTP/metabolismo , Dor/patologia , Fosfatidilinositol 3-Quinases/antagonistas & inibidores , Fosfatidilinositol 3-Quinases/metabolismo , Ratos , Ratos Sprague-Dawley , Receptor Tipo 4 de Melanocortina/agonistas , Células Receptoras Sensoriais/patologia , Gânglio Trigeminal/patologia , Wortmanina/farmacologia
16.
Pulm Pharmacol Ther ; 55: 62-66, 2019 04.
Artigo em Inglês | MEDLINE | ID: mdl-30763726

RESUMO

Cough is an important protective mechanism for clearing the airways but becomes a troublesome, and often difficult to treat, symptom in respiratory disease. Although cough can be produced as a reflex in response to the presence of irritants within the airways, emerging research demonstrates an unappreciated complexity in the peripheral and central neural systems that regulate cough. This complexity includes multiple primary sensory neurons that can induce or facilitate reflex coughing, different ascending central circuits in the brain that contribute to cough sensory discrimination and the perception of the urge-to-cough, and several descending brain systems for inducing, facilitating and inhibiting cough responses. Consequently, the mechanisms responsible for cough becoming dysregulated in disease are not likely homogeneous across all patients with chronic cough. The available data suggests that changes in primary sensory neuron excitability, altered central nervous system integration of sensory inputs and changes in descending control mechanisms may each contribute to the development of cough hypersensitivity.


Assuntos
Encéfalo/metabolismo , Tosse/fisiopatologia , Reflexo/fisiologia , Animais , Doença Crônica , Humanos , Células Receptoras Sensoriais/metabolismo
17.
Proc Natl Acad Sci U S A ; 116(9): 3811-3816, 2019 02 26.
Artigo em Inglês | MEDLINE | ID: mdl-30755524

RESUMO

The ability to detect environmental cold serves as an important survival tool. The sodium channels NaV1.8 and NaV1.9, as well as the TRP channel Trpm8, have been shown to contribute to cold sensation in mice. Surprisingly, transcriptional profiling shows that NaV1.8/NaV1.9 and Trpm8 are expressed in nonoverlapping neuronal populations. Here we have used in vivo GCaMP3 imaging to identify cold-sensing populations of sensory neurons in live mice. We find that ∼80% of neurons responsive to cold down to 1 °C do not express NaV1.8, and that the genetic deletion of NaV1.8 does not affect the relative number, distribution, or maximal response of cold-sensitive neurons. Furthermore, the deletion of NaV1.8 had no observable effect on transient cold-induced (≥5 °C) behaviors in mice, as measured by the cold-plantar, cold-plate (5 and 10 °C), or acetone tests. In contrast, nocifensive-like behavior to extreme cold-plate stimulation (-5 °C) was completely absent in mice lacking NaV1.8. Fluorescence-activated cell sorting (FACS) and subsequent microarray analysis of sensory neurons activated at 4 °C identified an enriched repertoire of ion channels, which include the Trp channel Trpm8 and potassium channel Kcnk9, that are potentially required for cold sensing above freezing temperatures in mouse DRG neurons. These data demonstrate the complexity of cold-sensing mechanisms in mouse sensory neurons, revealing a principal role for NaV1.8-negative neurons in sensing both innocuous and acute noxious cooling down to 1 °C, while NaV1.8-positive neurons are likely responsible for the transduction of prolonged extreme cold temperatures, where tissue damage causes pan-nociceptor activation.


Assuntos
Canal de Sódio Disparado por Voltagem NAV1.8/genética , Canais de Potássio/genética , Células Receptoras Sensoriais/fisiologia , Canais de Cátion TRPM/genética , Animais , Temperatura Baixa , Gânglios Espinais/diagnóstico por imagem , Gânglios Espinais/metabolismo , Gânglios Espinais/fisiologia , Camundongos , Nociceptores/metabolismo , Nociceptores/fisiologia , Células Receptoras Sensoriais/metabolismo , Sensação Térmica/genética
18.
Nat Commun ; 10(1): 643, 2019 02 07.
Artigo em Inglês | MEDLINE | ID: mdl-30733440

RESUMO

Nervous systems exhibit myriad cell types, but understanding how this diversity arises is hampered by the difficulty to visualize and genetically-probe specific lineages, especially at early developmental stages prior to expression of unique molecular markers. Here, we use a genetic immortalization method to analyze the development of sensory neuron lineages in the Drosophila olfactory system, from their origin to terminal differentiation. We apply this approach to define a fate map of nearly all olfactory lineages and refine the model of temporal patterns of lineage divisions. Taking advantage of a selective marker for the lineage that gives rise to Or67d pheromone-sensing neurons and a genome-wide transcription factor RNAi screen, we identify the spatial and temporal requirements for Pointed, an ETS family member, in this developmental pathway. Transcriptomic analysis of wild-type and Pointed-depleted olfactory tissue reveals a universal requirement for this factor as a switch-like determinant of fates in these sensory lineages.


Assuntos
Bulbo Olfatório/citologia , Bulbo Olfatório/metabolismo , Células Receptoras Sensoriais/citologia , Células Receptoras Sensoriais/metabolismo , Animais , Drosophila , Proteínas de Drosophila/metabolismo
19.
Mol Pharmacol ; 95(4): 433-441, 2019 04.
Artigo em Inglês | MEDLINE | ID: mdl-30679204

RESUMO

Transient receptor potential (TRP) cation channels are molecular targets of various natural products. TRPA1, a member of TRP channel family, is specifically activated by natural products such as allyl isothiocyanate (mustard oil), cinnamaldehyde (cinnamon), and allicin (garlic). In this study, we demonstrated that TRPA1 is also a target of trans-anethole in fennel oil (FO) and fennel seed extract. Similar to FO, trans-anethole selectively elicited calcium influx in TRPA1-expressing mouse sensory neurons of the dorsal root and trigeminal ganglia. These FO- and anethole-induced calcium responses were blocked by a selective TRPA1 channel antagonist, HC-030031. Moreover, both FO and trans-anethole induced calcium influx and transmembrane currents in HEK293 cells stably overexpressing human TRPA1 channels, but not in regular HEK293 cells. Mutation of the amino acids S873 and T874 binding site of human TRPA1 significantly attenuated channel activation by trans-anethole, whereas pretreating with glutathione, a nucleophile, did not. Conversely, activation of TRPA1 by the electrophile allyl isothiocyanate was abolished by glutathione, but was ostensibly unaffected by mutation of the ST binding site. Finally, it was found that trans-anethole was capable of desensitizing TRPA1, and unlike allyl isothiocyanate, it failed to induce nocifensive behaviors in mice. We conclude that trans-anethole is a selective, nonelectrophilic, and seemingly less-irritating agonist of TRPA1.


Assuntos
Anisóis/farmacologia , Óleos Voláteis/farmacologia , Canal de Cátion TRPA1/agonistas , Animais , Canais de Cálcio/metabolismo , Foeniculum/química , Células HEK293 , Humanos , Isotiocianatos/farmacologia , Camundongos , Camundongos Endogâmicos C57BL , Células Receptoras Sensoriais/efeitos dos fármacos , Células Receptoras Sensoriais/metabolismo , Raízes Nervosas Espinhais/efeitos dos fármacos , Raízes Nervosas Espinhais/metabolismo , Canais de Receptores Transientes de Potencial/metabolismo , Gânglio Trigeminal/efeitos dos fármacos , Gânglio Trigeminal/metabolismo
20.
Neuron ; 101(4): 738-747.e3, 2019 02 20.
Artigo em Inglês | MEDLINE | ID: mdl-30654923

RESUMO

Thermosensation is critical for avoiding thermal extremes and regulating body temperature. While thermosensors activated by noxious temperatures respond to hot or cold, many innocuous thermosensors exhibit robust baseline activity and lack discrete temperature thresholds, suggesting they are not simply warm and cool detectors. Here, we investigate how the aristal Cold Cells encode innocuous temperatures in Drosophila. We find they are not cold sensors but cooling-activated and warming-inhibited phasic thermosensors that operate similarly at warm and cool temperatures; we propose renaming them "Cooling Cells." Unexpectedly, Cooling Cell thermosensing does not require the previously reported Brivido Transient Receptor Potential (TRP) channels. Instead, three Ionotropic Receptors (IRs), IR21a, IR25a, and IR93a, specify both the unique structure of Cooling Cell cilia endings and their thermosensitivity. Behaviorally, Cooling Cells promote both warm and cool avoidance. These findings reveal a morphogenetic role for IRs and demonstrate the central role of phasic thermosensing in innocuous thermosensation. VIDEO ABSTRACT.


Assuntos
Proteínas de Drosophila/metabolismo , Neurogênese , Receptores Ionotrópicos de Glutamato/metabolismo , Células Receptoras Sensoriais/metabolismo , Sensação Térmica , Animais , Proteínas de Drosophila/genética , Drosophila melanogaster , Receptores Ionotrópicos de Glutamato/genética , Células Receptoras Sensoriais/citologia , Células Receptoras Sensoriais/fisiologia , Termotolerância
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