Muscarinic, adenosine and tropomyosin-related kinase B receptors modulate the neuromuscular developmental synapse elimination process

• Autores: Laura Nadal Magriñà
• Directores de la Tesis: Josep Tomas Ferrer (dir. tes.), Mª de les Neus García Sancho (dir. tes.), Ma. Angel Lanuza Escolano (dir. tes.)
• Lectura: En la Universitat Rovira i Virgili ( España ) en 2017
• Idioma: español
• Tribunal Calificador de la Tesis: Josep E. Esquerda Colell (presid.), Nicolau Ortiz Castellon (secret.), Teresa Victoria Obis Ibáñez (voc.)
• Programa de doctorado: Programa de Doctorado en Neurociencias por la Universidad de A Coruña; la Universidad de Castilla-La Mancha; la Universidad del País Vasco/Euskal Herriko Unibertsitatea; la Universidad Pablo de Olavide y la Universidad Rovira i Virgili
• Materias:
  • Ciencias de la vida
    • Biología celular
      • Morfología celular
  • Ciencias médicas
    • Medicina interna
      • Neurología
• Enlaces
  • Tesis en acceso abierto en:  hdl.handle.net  TDX 
• Resumen

  • The development of the nervous system involves an initially exuberant production of neurons that establish excessive synaptic contacts and the subsequent reduction in both neurons and synapses as maturation proceed. This process is called synaptic elimination and consists of an initial synapse overproduction to promote broad connectivity and a subsequent activity-dependent reduction in synapse number. This allows connectivity to be refined and specificity gained. Hebbian competition between different axons with different activities (the less active are punished) seems to be the fundamental characteristic of this process. In newborn animals, the skeletal muscle fibers are polyinnervated by several motor axons but at the end of the axonal competition, the endplates are innervated by a single axon. Because of its relative simplicity, the neuromuscular junction (NMJ) has long been used as a model for studying the general principles of synapse development in an attempt to understand the synapse elimination process.

    Various presynaptic receptors seem to play an important role in the axonal competition leading to synapse loss in the NMJ. The involvement of muscarinic acetylcholine autoreceptors (mAChRs), adenosine receptors (AR) and tropomyosin-related kinase B (TrkB) receptor in the elimination process may allow direct competitive interaction between nerve endings through a differential activity-dependent acetylcholine (ACh) release. Several signaling pathways coordinate the pre-, post-synaptic sites in the neuromuscular junction. In previous results obtained by Histology and Neurobiology Unit at URV indicated that there is a release inhibition mechanism based on a mAChR-PKC-VDCC intracellular cascade. When it is fully active in certain weak motor axons, it can depress ACh release and even disconnect synapses. This mechanism plays a central role in the elimination of redundant neonatal synapses because functional axonal withdrawal can indeed be reversed by mAChR, protein kinase C (PKC) or voltage-dependent calcium channels (VDCC) block. This seems to indicate that the involvement of these molecules in the process of synapse elimination. Is for this reason that is very important to know the cellular and molecular mechanisms that determine the plastic readjustment of the synapses to the functional requirements that determine its stabilization or its unstabilization and loss. These molecules and mechanisms have a considerable importance from the neurophysiological point of view and also as possible targets of therapeutic agents. In this thesis is shown the study of the multiple signaling pathways that converge and cooperate to determine the outcome of the synapse elimination process.

    In this thesis, it has been investigated by immunohistochemistry, confocal microscopy and quantitative morphological analysis the involvement of the individual and synergic or occlusive effect of M1-, M2- and M4-subtypes of mAChRs, A1 and A2A of ARs and TrkB in the control of the axonal elimination in developing NMJ. To perform this study, agonists and antagonists of these receptors were exposed in the Levator auris longus muscle of Thy1-YFP mice during postnatal days (P7, P9 and P15).

    The results show that mAChRs, ARs and TrkB cooperate in the elimination process and promote axonal disconnection at the beginning of the second postnatal week (P7 and P9) without affecting the postsynaptic maturation of the nicotinic receptor cluster. In summary, mAChRs, ARs and TrkB delay axonal loss at P7 (although mAChR accelerate the last phase of axonal disconnection) but accelerate it at P9. In terms of receptor cooperation, it has been demonstrated that M4 produces some occlusion of the M1 pathway and some addition to the A1 pathway, which suggests that they cooperate at P7. The cooperation between M1, A1 and A2A receptors promotes axonal loss at P9, whereas the potent axonal loss-promoting effect of M2 is largely independent of the other receptors. M1 and TrkB receptors work together to increase axonal loss rate at P9 but the effect of M2 is largely independent of the TrkB receptor.

    In conclusion, postnatal synapse elimination is a regulated multireceptor mechanism involving the cooperation of several muscarinic, adenosine and TrkB receptor subtypes that guarantees the monoinnervation of the neuromuscular synapses in the end of the process.