PHARMACOLOGICAL MANIPULATION OF MICROGLIA TO SUPPRESS NEUROINFLAMMATION AND PROTECT NEURONS

Abstract

Microglia are the resident immune cells of the brain that survey the microenvironment, provide trophic support to neurons, and clear debris to maintain homeostasis and healthy brain function. Microglia are also drivers of neuroinflammation in several neurodegenerative diseases. Excessive pro-inflammatory activity by microglia involves the release of soluble factors that can impair neuronal function. In this thesis, I tested the hypothesis that pro-inflammatory microglia could induce secondary damage to neurons and explored multiple pharmacological strategies to prevent this process. I found that stimulation of cultured microglia with bacterial lipopolysaccharide (LPS) and interferon-gamma (IFNγ) elicited a pro-inflammatory response that was strong enough to directly kill cultured neurons and that pre-treatment of microglia with synthetic cannabinoids targeting cannabinoid type 1 (CB1) and type 2 (CB2) receptors could reduce the inflammatory response enough to reduce neuronal death (Chapter 1). I then further explored the effects of LPS and IFNγ on the capacity of microglia to synthesize, degrade, and respond to cannabinoid receptor agonists. LPS and IFNγ each stimulated the upregulation of mRNA for Cnr2 and DAGLB in a biphasic manner (Chapter 2). The use of synthetic CB2 agonists (HU-308 and HU-433) were then used to further probe the mechanism of the anti-inflammatory effects of CB2 receptor activation on microglia. Activation of microglial CB2 receptors suppressed the canonical signaling of toll-like receptor 4 (TLR4) and directly inhibited the release of nitric oxide (NO) and tumour necrosis factor (TNF) (Chapter 4). An in-depth examination of the effects of LPS and IFNγ on microglia revealed that these stimuli act in a synergistic manner that is dependent on Janus kinase (JAK)1/2 as suppression of JAK1/2 prevented the microglial response to both molecules (Chapter 5). Finally, I explored the mechanisms by which the neurons were killed by microglia. Microglial-secreted factors induced apoptosis and necroptosis in neuronal cells which could be completely prevented by neutralization of TNF (Chapter 6). These findings suggest that direct suppression of microglia is sufficient to reduce secondary neurotoxicity and highlight some potential opportunities for the treatment of neuroinflammation.