Theiler's Murine Encephalomyelitis Virus Replicates in Primary Neuron Cultures and Impairs Spine Density Formation

TOMATIS, CARLA; LEÓN, ANTONELLA; LÓPEZ ORTIZ, AÍDA O; ONETO, PAULA; FUENTES, FEDERICO; FERRER, MARÍA F; CARRERA SILVA, EUGENIO A; SCORTICATI, CAMILA; GÓMEZ, RICARDO M

NEUROSCIENCE

PERGAMON-ELSEVIER SCIENCE LTD

Año: 2023 vol. 529 p. 162 - 171

0306-4522

In this study, we examined infection with the highly neurovirulent GDVII, the less neurovirulent DA strains, and with a mutant DA, which lacks the L* protein (L*-1) involved in viral persistence and demyelinating disease, to analyze the direct effects of Theiler´s murine encephalomyelitis virus (TMEV) replication using primary cultures of mouse brain hippocampal neurons. All viruses replicate in cultured neurons, with GDVII having the highest titers and L*-1 the lowest. Accordingly, all were positive for viral antigen staining 3 days postinfection (dpi), and DA and L*-1 were also positive after 12 dpi. NeuN + immunostaining showed an early and almost complete absence of positive cells in cultures infected with GDVII, an approximately 50% reduction in cultures infected with DA, and fewer changes in L*-1 strains at 3 dpi. Accordingly, staining with chloromethyltetramethylrosamine orange (Mitotracker OrangeTM) as a parameter for cell viability showed similar results. Moreover, at 1 dpi, the strain DA induced higher transcript levels of neuroprotective genes such as IFN-Iβ, IRF7, and IRF8. At 3 dpi, strains GDVII and DA, but not the L*-1 mutant, showed lower PKR expression. In addition, confocal analysis showed that L*-1-infected neurons exhibited a decrease in spine density. Treatment with poly (I:C), which is structurally related to dsRNA and is known to trigger IFN type I synthesis, reduced spine density even more. These results confirmed the use of mouse hippocampal neuron cultures as a model to study neuronal responses after TMEV infection, particularly in the formation of spine density.