Les neurones nocicepteurs jouent un rôle clé dans la défense de l’organisme. Dans le cas des réactions inflammatoires, ils initient des réflexes protecteurs tels que la toux, les vomissements, où les démangeaisons, et participent à la régulation de plusieurs mécanismes physiologiques, notamment la réponse immunitaire. Ils jouent ainsi un rôle prépondérant dans l’inflammation de type 2, souvent associée aux allergies. Mais les mécanismes qui permettent l’activation de ces neurones dans ce contexte sont encore mal connus. Au cours de ce projet de recherche, nous avons exploré la capacité des neurones nocicepteurs à détecter les signaux immunitaires spécifiquement associés à l’asthme. Nous avons ainsi identifié les caractéristiques des nocicepteurs des voies aériennes. Nous avons également démontré leur sensibilité aux allergènes grâce à l’expression du récepteur aux immunoglobulines de type E, FcεR1, ainsi que leur capacité à modifier leur transcriptome en réponse aux cytokines IL-4 et IL-13. Ces travaux soutiennent l’importance de la communication entre systèmes nerveux et immunitaires, et mettent en évidence de nouvelles cibles pour limiter la contribution neuronale aux réactions allergiques. / Nociceptor neurons play a major role in organism defense. In the context of inflammation, they initiate protective reflexes such as cough, vomiting, or itch, and participate in the regulation of various physiological mechanisms, including the immune response. They notably participate in type 2 inflammation, often associated with allergies. But the mechanisms driving the activation of nociceptor neurons in this context are still elusive. During this research project, we investigated the ability of nociceptor neurons to sense immune signals specifically associated with asthma. We identified the characteristics of airway innervating nociceptors. We also demonstrated their sensitivity to allergens through the expression of the Immunoglobulin E receptor FcεR1, as well as their ability to change their transcriptome in response to IL-4 and IL-13. This work supports the importance of bidirectional communication between the nervous and immune systems and unravels new targets to regulate neuronal contribution to inflammation.

ErbB2 gehört zu den Klasse I Rezeptor-Tyrosinkinasen und funktioniert als Ko-rezeptor bei der Vermittlung des Neuregulin-Signals. Während der Embryonal-entwicklung wird ErbB2 im Herzen, in den Neuralleistenzellen, im Muskel und in den Epithelien exprimiert (Kokai et al. 1987). Embryonen mit einer Null-Mutation im ErbB2 Gen sterben am Tag 10,5 der Embryonalentwicklung. Die Mutation bewirkt eine morphogenetische Fehlbildung des Herzens, die durch das Fehlen von ventrikulären Trabekeln gekennzeichnet ist (Lee et al. 1995). Weiterhin zeigen diese Embryonen Defekte in den Kranialganglien und in der primären sympathischen Ganglien-Kette, die von Neuralleistenzellen gebildet werden (Lee et al. 1995; Erickson et al. 1997; Britsch et al. 1998). Die herzspezifische Expression von ErbB2 cDNA ermöglicht ErbB2-/- Tieren, sich bis zur Geburt zu entwickeln. Dies erlaubte mir, spätere Funktionen des Rezeptors zu untersuchen. In den geretteten ErbB2-/- Embryonen erfolgte die Bildung der ventrikulären Trabekel, der fingerähnlichen Ausstülpungen des Myokards, zwischen dem 9. und 10. Tag in der Embryonalentwicklung. In den späteren Phasen der intrauterinen Entwicklung war das Herz der geretteten Tiere normal ausgebildet. In den ErbB2-/-R Embryonen fehlten Schwann'sche Zellen entlang der peripheren Nerven. Die Abwesenheit von Schwann'schen Zellen führte zum massiven Absterben von sensorischen und motorischen Neuronen des Rückenmarkes. Dabei zeigten sensorische Neuronen eine frühe Abhängigkeit von neurotrophen Faktoren, die von Schwann'schen Zellen produziert werden, während Motoneuronen diese Faktoren in einer späteren Phase benötigen. Zusätzlich ist bekannt, daß sensorische Neuronen und Motoneuronen neurotrophe Fakten benötigen, die von den Zielorganen, z.B. den Muskeln, produziert werden. Motoneuronen im thorakalen Rückenmark sind nur minimal betroffen, während die Degeneration von Moto-neuronen in den zervikalen und lumbalen Segmenten stark ausgeprägt ist. Verschiedene Motoneuron-Typen unterscheiden sich also in ihrer Abhängigkeit von neurotrophen Signalen. Weiterhin sind die peripheren Nerven der ErbB2-/-R Tiere defaszikuliert und ungeordnet. Der N. phrenicus, der das Diaphragma innerviert, retrahiert und ist am Tag 17 der Entwicklung vollständig degeneriert. Deshalb können die mutanten Tiere bei der Geburt nicht atmen und sterben infolgedessen. Überraschenderweise erfolgt in den geretteten ErbB2-/-R Embryonen die post-synaptische Expression und Aggregation der Acetylcholin-Rezeptoren. Die Phäno-typen der ErbB2-/-R und ErbB3-/- mutanten Tieren sind sehr ähnlich. Dies zeigt, daß ErbB2 eine essentielle Korezeptor-Funktion für ErbB3 in der Vermittlung der Neuregulin-Signale übernimmt. / ErbB2 belongs to class I of receptor tyrosine kinases and functions as a co-receptor by the transduction of the neuregulin signal. During embryonic development the ErbB2 gene is expressed in the heart, neural crest, in muscle and epithelial cells (Kokai et al. 1987). Embryos with null mutation of the ErbB2 gene die at midgestation. The mutation causes a morphogenetic defect that results in the absence of trabecules (Lee et al. 1995). In addition the mutant embryos show defects in cranial ganglia and in the primary sympathetic ganglia chain (Lee et al. 1995; Erickson et al. 1997; Britsch et al. 1998). The heart specific expression of ErbB2 cDNA allowed the mutant animals to survive till birth. This enabels me to study the late function of the receptor. In rescued ErbB2-/- embryos the ventricular trabecules, which are finger-like extensions of the myocardium, form properly between E9 and E10 of embryonic development. At late stages of intrauteral development the hearts of the rescued animals showed an overall normal growth. ErbB2-/- embryos lack Schwann cells along peripheral nerves. The absence of Schwann cells leads to enormous degeneration of sensory and motoneurons. Whereas sensory neurons show an early dependency on neurotrophic factors produced by Schwann cells, motoneurons revealed requirement of these factors during the late phase of their development. Moreover it is known that sensory and motoneurons require neurotrophic factors which are produced by their target tissues such as muscle. Motoneurons at the thoracic level of the spinal cord are minimaly affected, whereas the degeneration of motoneurons at cervical and lumbar segments of the spinal cord are pronounced. This indicates that different motoneuron types differ in their dependency on neurotrophic signals. Furthermore axons of peripheral nerves in ErbB2-/-R (rescued) animals show defasciculation and desorganization. Nervous phrenicus, that innervates the diaphragm muscle retracts and degenerates entirely at E17 of embryonic development. As a result newborn animals can not breath and die shortly after birth. Surprisingly, the expression and aggregation of AchRs (Acetylcholine Receptors) take place in rescued ErbB2-/-R embryos. The overall phenotype of ErbB2-/-R embryos is very similar to that of ErbB3-/- embryos. This substantiates the essential function of ErbB2 as the functional co-receptor for ErbB3 to transmit the neuregulin signal.

Indiana University-Purdue University Indianapolis (IUPUI) / Aromatase inhibitors (AIs) are the current standard of care for the treatment of hormone receptor positive breast cancer in postmenopausal women. Nearly one-half of patients receiving AI therapy develop musculoskeletal toxicity that is characterized by joint and/or muscle pain and approximately one-fourth of patients discontinue their therapy as a result of musculoskeletal pain. Since there are no effective strategies for prevention or treatment, insight into the mechanisms of AI-induced pain is critical to improve treatment. However, there are few studies of AI effects in animal models of nociception. To determine whether AIs produce hypersensitivity in animal models of pain, I examined the effects of AI administration on mechanical, thermal, and chemical sensitivity in rats. The results demonstrate that (1) repeated injection of 5 mg/kg letrozole in male rats produces mechanical, but not thermal, hypersensitivity that extinguishes when drug dosing is stopped; (2) administering a single dose of 1 or 5 mg/kg letrozole in ovariectomized (OVX) rats also induces mechanical hypersensitivity, without altering thermal sensitivity and (3) a single dose of 5 mg/kg letrozole or daily dosing of letrozole or exemestane in male rats augments flinching behavior induced by intraplantar ATP injection. To determine whether the effects of AIs on nociceptive behaviors are mediated by activation or sensitization of peptidergic sensory neurons, I determined whether letrozole exposure alters release of calcitonin gene-related peptide (CGRP) from isolated rat sensory neurons and from sensory nerve endings in rat spinal cord slices. No changes in basal, capsaicin-evoked or high extracellular potassium-evoked CGRP release were observed in sensory neuronal cultures acutely or chronically exposed to letrozole. Furthermore, letrozole exposure did not alter the ability of ATP to augment CGRP release from sensory neurons in culture. Finally, chronic letrozole treatment did not augment neuropeptide release from spinal cord slices. Taken together, these results do not support altered release of this neuropeptide into the spinal cord as mediator of letrozole-induced mechanical hypersensitivity and suggest the involvement of other mechanisms. Results from this dissertation provide a new experimental model for AI-induced hypersensitivity that could be beneficial in delineating mechanisms mediating pain during AI therapy.

Indiana University-Purdue University Indianapolis (IUPUI) / Chronic inflammatory disorders are characterized by an increase in excitability of small diameter sensory neurons located in dorsal root ganglia (DRGs). This sensitization of neurons is a mechanism for chronic inflammatory pain and available therapies have poor efficacy and severe adverse effects when used chronically. Prostaglandin E₂ (PGE₂) is an inflammatory mediator that plays an important role in sensitization by activating G-protein coupled receptors (GPCRs) known as E-series prostaglandin receptors (EPs) coupled to the protein kinase A (PKA) pathway. EPs are known to downregulate upon prolonged exposure to PGE₂ or in chronic inflammation, however, sensitization persists and the mechanism for this is unknown. I hypothesized that persistence of PGE₂-induced hypersensitivity is associated with a switch in signaling caused by prolonged exposure to PGE₂ or the neurotrophin nerve growth factor (NGF), also a crucial inflammatory mediator. DRG cultures grown in the presence or absence of either PGE₂ or NGF were used to study whether re-exposure to the eicosanoid is able to cause sensitization and activate PKA. When cultures were grown in the presence of NGF, PGE₂-induced sensitization was not attenuated by inhibitors of PKA. Activation of PKA by PGE₂ was similar in DRG cultures grown in the presence or absence of NGF when phosphatase inhibitors were added to the lysis and assay buffers, but significantly less in cultures grown in the presence of NGF when phosphatase inhibitors were not added. In DRG cultures exposed to PGE₂ for 12 hours-5 days, sensitization after re-exposure to PGE₂ is maintained and resistant to PKA inhibition. Prolonged exposure to the eicosanoid caused complete loss of PKA activation after PGE₂ re-exposure. This desensitization was homologous, time dependent, reversible, and insurmountable by a higher concentration of PGE₂. Desensitization was attenuated by reduction of expression of G-protein receptor kinase 2 and was not mediated by PKA or protein kinase C. The presented work provides evidence for persistence of sensitization by PGE₂ as well as switch from the signaling pathway mediating this sensitization after long-term exposure to NFG or PGE₂.