Microglial reactivity and neuroinflammation-driven changes in motivational behaviors are regulated by Orai1 calcium channels, 2026, DeMeulenaere et al

[jnmaciuch]

Microglial reactivity and neuroinflammation-driven changes in motivational behaviors are regulated by Orai1 calcium channels

Spoiler: Authors

DeMeulenaere, Kaitlyn E.; Grant, Rogan A.; Martin, Megan E.; Valencia, Hiam A.; Radulovic, Jelena; Salter, Michael W.; Prakriya, Murali

Abstract
Microglia are the brain’s resident immune cells that respond to injury and disease by transitioning between homeostatic and reactive states. These cell state transitions determine whether microglia promote or resolve inflammation in the central nervous system (CNS). In this study, we explored the role of Ca 2+ signaling in regulating broader microglial cell state transitions and identified Orai1 Ca 2+ channels as critical regulators of microglial plasticity and neuroinflammatory signaling. Conditional deletion of Orai1 in microglia impaired their ability to adopt reactive, proinflammatory states.

Transcriptomic and metabolomic profiling revealed that Orai1 deletion suppressed the expression of proinflammatory genes linked to immunity, inflammation, and cell metabolism. Conversely, Orai1-deficient microglia generated greater amounts of neuroprotective and anti-inflammatory mediators, including BDNF, ARG1, and the mitochondrial metabolite itaconate. In a model of CNS inflammation induced by peripheral lipopolysaccharide (LPS) challenge, microglial Orai1 deletion attenuated microglial and astrocyte reactivity and reduced hippocampal amounts of the proinflammatory cytokines IL-1β and IL-6.

Consistent with these cellular changes, microglial Orai1 knockout mice were protected against LPS-induced decreases in motivational behaviors, including impaired reward-seeking and escape behaviors. These findings establish Orai1 channels as key regulators of microglial cell state transitions, linking Ca 2+ signaling to neuroinflammation and inflammation-driven behavioral dysfunction.

[Paragraph breaks added]

Web | DOI | Science Signaling

 

[jnmaciuch]

Originally posted by @Nightsong here:

Post in thread 'Miscellaneous Research Thread'

Yesterday at 4:02 PM

Curious paper in Science Signalling from a couple of weeks ago exploring LPS (bacterial endotoxin) induced sickness responses in mice. The authors found that a certain type of calcium selective ion channel (Orai1) was associated with aspects of the observed induced sickness response behaviours:

In this study, we show that Orai1 Ca2+ channels are central regulators of microglial reactivity, microglia-driven neuroinflammation, and the development of depression-like behaviors after systemic inflammatory challenge. Microglia are highly plastic and reversibly interconvert between different...

• Nightsong

• 

Making a separate thread because I wanted to do a deep dive during my breaks today (bear with me, posts will be sporadic)

I know a bit about the lab that this came from, they've been interested in this gene in other contexts (e.g. neuropathic pain) for a while.

 

[jnmaciuch]

Starting off, they created an Orai1fl/fl Cx3CR1-Cre/ERT2 mouse model--what that means is that the mouse is engineered so that the gene Orai1 is knocked out only in cells that also express Cx3cr1, and only when you inject tamoxifen that binds to ERT2 (this is often done when you want to knock out a gene only temporarily, or for genes that are critical for development that can't be knocked out from birth).

I've mentioned before that Cx3cr1-Cre is a common way of targetting genes in microglia. However, the gene is also expressed in other cells outside the brain, so using this model alone you can't always assume that any changes are solely due to knocking out that gene in microglia.

From Fig. 1, this is just confirming that knocking out Orai1 in this mouse model affects calcium signaling in isolated mouse microglia as expected.

 

[jnmaciuch]

To do more detailed characterization, they created a similar knockout but only in cultured microglia to see if those cells react differently to stimulation.

Fig 2:


From from more familiarity with immunology work, the stimulation that they're using here is a bit unusual. Instead of directly using immune signals, they're using a combination of agents that directly affect intracellular calcium signaling in a way that mimics what happens downstream of several microglial receptors. What they found is that the Orai1 knockout cells have a suppressed response for several classic immune pathways (NFkB and STAT3, and IFN related signaling mostly). I don't think there's anything wrong per se in how this was done, but it adds a degree of separation since we are relying on this kind of stimulation being relevant to infection response

 

[jnmaciuch]

Fig 3:


Main takeaway is just that knocking out Orai1 substantially changes how microglia respond metabolically to that same stimulation as in the previous figure.

 

[jnmaciuch]

Fig 4:



Here we're doing direct stimulation of microglia with LPS, a bacterial fragment that induces an immune response via TLR4. What's encouraging is that we see very similar changes as in the earlier stimulation, with Orai1 knockouts having a blunted response across multiple classic immune pathways.

 

[jnmaciuch]

Fig 5:


Now they're going back to the mouse model, and seeing what happens when LPS is injected peripherally. This would be more relevant, because in the vast majority of infections, fragments of the pathogen aren't actually making it into the brain to activate TLRs on microglia. As in the in-vitro model, microglia collected from mice after peripheral exposure to LPS have a lowered interferon and NFkB response. In figures D and G, they're also showing that there are different sets of genes that are differential between the knock out and wild type at baseline (with saline), 6 hrs after injection, and 24 hours after injection.

So it's interesting that even if we don't expect LPS to be directly stimulating microglia, they respond similarly as if they were being directly stimulated. An exception is some immune pathways like the inflammasome, which were actually higher in the Orai1 knockouts.

 

[jnmaciuch]

Skipping ahead to the behavioral data, they wanted to see if their Orai1 knockout exhibits different "sickness behaviors" compared to the wild type mouse.

Fig. 6:

They have a couple tests, checking things like food motivation and avoidance of unpleasant stimuli. I'll note that these tests are usually used to measure depressive behaviors, so might not be as relevant for pathways related to flu-like malaise and exhaustion in humans.

What they found is that the Orai1 knockouts behaved very similarly to control mice regardless of whether they were injected with LPS or not. The authors interpret this to mean that depression-like behaviors in response to infection are dependent on Orai1.

 

[jnmaciuch]

Overall takeaway, I think the authors have done a very thorough job showing that this gene is pretty critical to microglial responsiveness to stimuli. the main piece this study is missing is definitive proof that this gene in microglia mediate depressive-like behavior. It seems likely based on their data, but unfortunately the mouse model cannot rule out the possibility that other immune cells, or even other non-immune cells that express CXC3R1, are responsible.