Microglial response and synaptic plasticity after proximal axotomy and treatment with memantine

Abstract

Excitatory synaptic transmission in the spinal cord is mediated by the neurotransmitter glutamate acting on receptors such as NMDA and AMPA. Excessive NMDA activation, as observed in compressive root lesions, triggers excitotoxic processes that culminate in alterations to the cellular and molecular microenvironment of the spinal cord and the synaptic organization between peripheral afferents and motoneurons, causing maladaptive neuroplasticity events and consequent functional impairment that is maintained even after axonal regeneration. Thus, the therapeutic hypothesis arises of reducing the excitotoxic effect of glutamate through NMDA receptor antagonists, such as memantine. To test this hypothesis, this study aims to evaluate the microglial response and synaptic plasticity after proximal axotomy treatment with memantine. To this end, we will crush the ventral roots (L4-L6) in adult C57BL/6JUnib mice, divided into the following experimental groups: Vehicle, Memantine 30 mg/kg, Memantine 45 mg/kg and Memantine 60 mg/kg. Treatment will take place orally for 14 days after the injury. Once treatment has ceased, the animals will be kept for a further 14 days until perfusion is performed and specimens obtained. The lumbar intumescences will then be processed to assess the morphological alteration of the microglia (via morphological analysis and Sholl analysis), quantification of total synaptic coverage, differentiation of excitatory and inhibitory synapses and localization of receptors (immunohistochemistry technique for the markers synaptophysin, VGLUT-1, GAD65 and AMPA). In addition, 3 and 7 days after injury and treatment, changes in the gene expression of genes related to glutamatergic excitotoxicity will also be assessed using the RT-qPCR technique.