Two-photon laser scanning microscopy imaging of intact spinal cord and cerebral cortex reveals requirement for CXCR6 and neuroinflammation in immune cell infiltration of cortical injury sites.
The mouse spinal cord is an important site for autoimmune and injury models. Skull thinning surgery provides a minimally invasive window for microscopy of the mouse cerebral cortex, but there are no parallel methods for the spinal cord. We introduce a novel, facile and inexpensive method for two-photon laser scanning microscopy of the intact spinal cord in the mouse by taking advantage of the naturally accessible intervertebral space. These are powerful methods when combined with gene-targeted mice in which endogenous immune cells are labeled with green fluorescent protein (GFP). We first demonstrate that generation of the intervertebral window does not elicit a reaction of GFP(+) microglial cells in CX3CR1(gfp/+) mice. We next demonstrate a distinct rostrocaudal migration of GFP(+) immune cells in the spinal cord of CXCR6(gfp/+) mice during active experimental autoimmune encephalomyelitis (EAE). Interestingly, infiltration of the cerebral cortex by GFP(+) cells in these mice required three conditions: EAE induction, cortical injury and expression of CXCR6 on immune cells.
Helen L. and Martin S. Kimmel Center for Biology and Medicine of the Skirball Institute of Biomolecular Medicine, New York University School of Medicine, 540 First Avenue, New York, NY 10016, USA. jiyun.kim@med.nyu.edu
Jiyun V
JV
Kim
Ning
N
Jiang
Carlos E
CE
Tadokoro
Liping
L
Liu
Richard M
RM
Ransohoff
Juan J
JJ
Lafaille
Michael L
ML
Dustin
D000818:Animals; D002465:Cell Movement; D002540:Cerebral Cortex / Q000276:immunology* / Q000293:injuries / Q000473:pathology; D000075743:Chemokine CXCL16; D054427:Chemokine CXCL6 / Q000096:biosynthesis / Q000235:genetics; D004681:Encephalomyelitis, Autoimmune, Experimental / Q000175:diagnosis / Q000276:immunology* / Q000473:pathology; D049452:Green Fluorescent Proteins / Q000235:genetics; D051379:Mice; D008810:Mice, Inbred C57BL; D008822:Mice, Transgenic; D018613:Microscopy, Confocal; D019060:Minimally Invasive Surgical Procedures; D017785:Photons; D054387:Receptors, CXCR / Q000096:biosynthesis* / Q000235:genetics; D000075744:Receptors, CXCR6; D013116:Spinal Cord / Q000276:immunology* / Q000473:pathology; D013601:T-Lymphocytes / Q000276:immunology / Q000378:metabolism* / Q000473:pathology
D000075743:Chemokine CXCL16; D054427:Chemokine CXCL6; C498926:Cxcl16 protein, mouse; C482906:Cxcr6 protein, mouse; D054387:Receptors, CXCR; D000075744:Receptors, CXCR6; D049452:Green Fluorescent Proteins