Neuronal Conditional Knockout of Collapsin Response Mediator Protein 2 Ameliorates Disease Severity in a Mouse Model of Multiple Sclerosis

Abstract

We previously showed that treatment with lanthionine ketimine ethyl ester (LKE) reduced disease severity and axonal damage in an experimental autoimmune encephalomyelitis (EAE) mouse model of multiple sclerosis and increased neuronal maturation and survival in vitro. A major target of LKE is collapsin response mediator protein 2 (CRMP2), suggesting this protein may mediate LKE actions. We now show that conditional knockout of CRMP2 from neurons using a CamK2a promoter to drive Cre recombinase expression reduces disease severity in the myelin oligodendrocyte glycoprotein (MOG) 35-55 EAE model, associated with decreased spinal cord axonal damage, and less glial activation in the cerebellum, but not the spinal cord. Immunohistochemical staining and quantitative polymerase chain reaction show CRMP2 depletion from descending motor neurons in the motor cortex, but not from spinal cord neurons, suggesting that the benefits of CRMP2 depletion on EAE may stem from effects on upper motor neurons. In addition, mice in which CRMP2 S522 phosphorylation was prevented by substitution for an alanine residue also showed reduced EAE severity. These results show that modification of CRMP2 expression and phosphorylation can influence the course of EAE and suggests that treatment with CRMP2 modulators such as LKE act in part by reducing CRMP2 S522 phosphorylation.

Conditional knockout of CRMP2 in neurons, but not astrocytes, disrupts spinal nociceptive neurotransmission to control the initiation and maintenance of chronic neuropathic pain

“…Congruent with this observation is our previous finding that conditional knockout of CRMP2 from excitatory neurons (CaMKIIα + ) prevented gliosis in a model of experimental autoimmune encephalomyelitis. 43 Glial activation in the spinal cord is a marker for chronic neuropathic pain. 34 Increased excitatory synaptic activity in the spinal cord triggers astrogliosis.…”

“…To investigate the cell-specific contribution of CRMP2, we used a transgenic mouse line where LoxP sequences flank the exon 3 of the dpysl2 gene (designated as CRMP2 f/f in this article). 43 Mice were injected intrathecally with an adeno-associated virus serotype 9 (AAV9) expressing the recombinase Cre under the control of the calcium/calmodulin-dependent protein kinase II alpha (CaMKIIa) promoter; CaMKIIa is a marker of glutamatergic neurons in the central nervous system 16,25,31 and is expressed in SDH's superficial laminae and DRG neurons involved in nociceptive pathways. 11 Using this CaMKIIa-GFP-CRE-AAV9, we tested whether conditional knockout of CRMP2 from glutamatergic neurons could recapitulate our observations in rats injected with a CRMP2 siRNA (Fig.…”

“…Pathogen-free, male Sprague-Dawley rat pups (postnatal 10-15 days; Envigo, Placentia, CA) and pups (8-15 days old) from male mice homozygous for the floxed Crmp2 allele (CRMP2 f/f ) mouse 43 were used for electrophysiological experiments. Adult male (21-28 g) and female (16-20 g) CRMP2 f/f mice were used for behavioral experiments.…”

“…38,40 However, what role, if any, CRMP2 plays in aberrant excitatory synaptic transmission underlying neuropathic pain processing after peripheral nerve injury has not been previously investigated. The availability of a transgenic mouse homozygous for the floxed Crmp2 allele (CRMP2 f/f ) 43 enables selective Cre recombinase-mediated knockout of CRMP2 expression at the spinal gate for nociceptive neurotransmission and allows us to address this question.…”

Collapsin Response Mediator Proteins: Their Biological Functions and Pathophysiology in Neuronal Development and Regeneration

“…MS is a chronic inflammatory, demyelinating, and neurodegenerative disorder of the CNS. In an EAE mouse model of MS, the importance of CRMP2 Ser522 phosphorylation was demonstrated using CRMP2 KI/KI mice (Moutal et al, 2019). CRMP2 is also expressed in the immune system and plays a critical role in T lymphocyte polarization and migration (Vincent et al, 2005).…”