Electronic Thesis and Dissertation Repository

Thesis Format

Integrated Article

Degree

Master of Science

Program

Neuroscience

Supervisor

Inoue, Wataru

Abstract

The brain senses inflammatory signals and drives the release of glucocorticoids (GCs) — potent immunosuppressants — via the activation of the hypothalamic-pituitary-adrenal (HPA) axis. This inflammation-induced HPA axis activation is largely mediated by prostaglandin E2 (PGE2), acting on two subtypes of the PGE2 receptor, EP1 and EP3. Recently, our group revealed EP3 signaling mechanisms that excite HPA axis regulatory neurons. This thesis sought to tease out the remaining EP1 signaling mechanisms. Considering that the excitability of HPA axis regulatory neurons is constrained by GABAA receptor-mediated synaptic inhibition that relies on low-level intracellular Cl-. We hypothesized that PGE2-EP1 signaling impairs GABAA receptor-mediated inhibition by increasing intracellular Cl- levels. We used two electrophysiological approaches (perforated patch and whole-cell recordings) and showed that PGE2 depolarizes the reversal potential of GABAA receptor currents (EGABA), an indicator of intracellular Cl- elevation. The effect of PGE2 was mimicked by EP1 agonist and prevented by EP1 antagonist. The depolarizing shift was slow to develop but became significant by 20 min post PGE2. Our results indicate that PGE2-EP1 coupling induces a slow depolarizing shift in EGABA for the excitation of PVN-CRH neurons during inflammation.

Summary for Lay Audience

When the body encounters an injury or illness, it often goes through an immune process called inflammation. This involves the recruitment of other immune cells to the site of the illness/injury to initiate defence, cleanup, and repair. The body can regulate the inflammation process via the activation of a pathway called the hypothalamic-pituitary-adrenal axis (HPA) axis, which releases hormones called glucocorticoids (GCs) – which are potent immune regulators. The immune system activates the HPA axis by activating a subset of neurons (parvocellular corticotrophin-releasing hormone neurons; PVN-CRH neurons) in a brain region called the hypothalamus. This is achieved via an immune-signalling molecule called prostaglandin (PG)E2. PGE2 is known to act through two distinct (receptor-based) signaling pathways; EP1 & EP3. Previously our group showed how the EP3 pathway disinhibits these neurons, allowing their activation, and driving the HPA axis. This work aims to understand how the EP1 pathway modulates these neurons. We hypothesized that the EP1 pathway shifts the magnitude and direction of inhibition in the PVN-CRH neurons. Using electronic recordings (patch-clamp electrophysiology) from individual neurons, we looked at changes in induced incoming inhibitory signals in these neurons. We found that inhibition magnitude is weakened or even reversed in these neurons in the presence of both PGE2 and iloprost (a chemical mimicking PGE2 that activates the EP1 pathway). Moreover, blocking the EP1 pathway while applying PGE2 partially prevented this effect. We theorized that this weakening of inhibition would lead to disinhibition of the PVN-CRH neurons and activation of the HPA axis. We posited that this EP1 pathway works in compliment to the EP3 pathway to activate the HPA axis during periods of inflammation. In summary, we demonstrated the consequences of PGE2-EP1 signaling in PVN-CRH. In the broader scope, this contributes to our understanding of how inflammation and the brain interface.

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