Excess glutamate secreted from astrocytes drives upregulation of P-glycoprotein in endothelial cells in amyotrophic lateral sclerosis

• Published in: Experimental neurology Vol. 316; pp. 27 - 38
• Main Authors: Mohamed, Loqman A., Markandaiah, Shashirekha S., Bonanno, Silvia, Pasinelli, Piera, Trotti, Davide
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 01.06.2019
• Subjects: Amyotrophic lateral sclerosis; Amyotrophic Lateral Sclerosis - metabolism; Animals; Astrocytes - metabolism; ATP Binding Cassette Transporter, Subfamily B - biosynthesis; ATP Binding Cassette Transporter, Subfamily B - genetics; Blood brain barrier; Blood-Brain Barrier - metabolism; Blood-Brain Barrier - pathology; Capillaries - metabolism; Cells, Cultured; Culture Media, Conditioned; Dizocilpine Maleate - pharmacology; Endothelial cells; Endothelial Cells - metabolism; Excitatory Amino Acid Antagonists - pharmacology; Glutamate; Glutamic Acid - metabolism; Humans; Mutation - genetics; NMDA receptor; P-glycoprotein; Pharmacoresistance; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate - biosynthesis; Receptors, N-Methyl-D-Aspartate - genetics; Riluzole; Superoxide Dismutase-1 - genetics; Up-Regulation; BBB; P-gp; EP-I; ALS; PLA2; Amyotrophic lateral sclerosis; ECF; ACM; NFkB; Glutamate; MRPs; Blood brain barrier; Riluzole; Endothelial cells; P-glycoprotein; MDR1; ECs; Nrf2; ROS; Pharmacoresistance; COX2; NMDAR; TEER; NMDA receptor
• ISSN: 0014-4886; 1090-2430
• DOI: 10.1016/j.expneurol.2019.04.002

In amyotrophic lateral sclerosis (ALS), upregulation in expression and activity of the ABC transporter P-glycoprotein (P-gp) driven by disease advancement progressively reduces CNS penetration and efficacy of the ALS drug, riluzole. Post-mortem spinal cord tissues from ALS patients revealed elevated P-gp expression levels in endothelial cells of the blood-spinal cord barrier compared to levels measured in control, non-diseased individuals. We recently found that astrocytes expressing familial ALS-linked SOD1 mutations regulate expression levels of P-gp in endothelial cells, which also exhibit a concomitant, significant increase in reactive oxygen species production and NFκB nuclear translocation when exposed to mutant SOD1 astrocyte conditioned media. In this study, we found that glutamate, which is abnormally secreted by mutant SOD1 and sporadic ALS astrocytes, drives upregulation of P-gp expression and activity levels in endothelial cells via activation of N-Methyl-D-Aspartic acid (NMDA) receptors. Surprisingly, astrocyte-secreted glutamate regulation of endothelial P-gp levels is not a mechanism shared by all forms of ALS. C9orf72-ALS astrocytes had no effect on endothelial cell P-gp expression and did not display increased glutamate secretion. Utilizing an optimized in vitro human BBB model consisting of patient-derived induced pluripotent stem cells, we showed that co-culture of endothelial cells with patient-derived astrocytes increased P-gp expression levels and transport activity, which was significantly reduced when endothelial cells were incubated with the NMDAR antagonist, MK801. Overall, our findings unraveled a complex molecular interplay between astrocytes of different ALS genotypes and endothelial cells potentially occurring in disease that could differentially impact ALS prognosis and efficacy of pharmacotherapies. [Display omitted] •iPS-derived astrocytes from ALS patients release abnormally high glutamate levels.•iPSC-derived astrocytes from sporadic and SOD1/ALS upregulate P-gp in BBB endothelium.•Astrocyte-released glutamate upregulate P-gp via NMDAR, COX2 and NFκB mediated pathway.