Disordered communication between cortical neurons and glia underlies many of the characteristics of Fragile X syndrome (FXS), the most common monogenic form of intellectual disability and autism spectrum disorder (ASD). Despite extensive research, no effective treatments exist to comprehensively mitigate ASD- or FXS-related cognitive and motor disabilities, sensory hyperresponsivity, seizures, and other excitation-related symptoms. Glial-glial and glial-neuronal communication can be facilitated by purinergic signalling pathways, which utilize ATP, UTP, and their metabolites to influence both short-term and longer-term activation. The overall objective of this thesis work was to establish whether purinergic signalling is dysregulated within cortical astrocytes derived from the Fmr1 KO mouse model of FXS, and furthermore, to determine whether astrocyte purinergic dysregulations contribute to aberrant Fmr1 KO neuronal-glial interactions. Collectively, these studies provide the first reported evidence that P2Y receptor-driven purinergic signalling is elevated in Fmr1 KO cortical astrocytes, and suggest that this impacts the formation and activity of neuronal circuitry in a manner consistent with FXS symptomatology. Fmr1 KO cortical astrocyte dysregulations included elevated expression of P2Y2 and P2Y6 purinergic receptors, increased intracellular calcium release following P2Y activation, aberrant levels of intracellular purinergic signalling molecules, and increased ectonucleotidase glycosylation. UTP treatment promoted excess Fmr1 KO astrocyte expression and secretion of the synaptogenic protein TSP-1 to potentially influence neuronal connectivity, as well as increased phosphorylation of transcription factor STAT3 to likely drive cortical immune responses. Both exogenous UTP and the presence of Fmr1 KO astrocyte secretions promoted neurite outgrowth, while Fmr1 KO astrocyte-neuron co-cultures demonstrated elevated neuronal burst frequency that was normalized through chronic and selective P2Y2 antagonism. Together, these findings indicate novel and significant astrocyte P2Y-mediated purinergic upregulations within the Fmr1 KO mouse cortex, and suggest that astrocyte purinergic signalling should be further investigated in the search for innovative FXS treatments. / Thesis / Doctor of Philosophy (PhD) / Autism spectrum disorders (ASDs) have become a serious health concern in recent years due to rapidly rising rates of diagnosis. Despite extensive research, there are still no effective treatments for these disorders of brain development. It is therefore important that we study the cellular events contributing to ASDs in order to design new therapeutic strategies. The most common inherited form of ASD is Fragile X syndrome (FXS), which is characterized by cognitive and motor disabilities, sensory hyperresponsivity, attention deficits, hyperactivity, and seizures. Using the Fmr1 knockout (KO) mouse model of FXS, recent research has shown that many of these symptoms are related to disordered communication between brain cells within the cerebral cortex; specifically, between neurons and the helper-like cells known as astrocytes. One form of cellular signalling that supports this communication is known as the purinergic signalling pathway. Collectively, this thesis work is the first to show that purinergic signalling is increased in Fmr1 KO mouse cortical astrocytes and that it impacts FXS neuronal connections. Specifically, Fmr1 KO cortical astrocytes demonstrated increased communication using purinergic signalling, due to greater expression of P2Y2 and P2Y6 purinergic receptors and altered levels of the molecules that stimulate these receptors. Activation of Fmr1 KO astrocyte P2Y receptors promoted expression of the neuronal connection-forming protein TSP-1 and stimulated additional astrocyte signalling pathways. As a result of these changes, when Fmr1 KO neurons were grown in the presence of Fmr1 KO astrocytes, they grew longer extensions and demonstrated greater activity than wildtype controls, in a manner consistent with the excitation-related symptoms of FXS. Selectively targeting P2Y2-driven purinergic pathways with drug treatments corrected this activity, thereby revealing a potential new therapeutic approach for FXS. Understanding excess astrocyte P2Y-driven purinergic communication within the brain may therefore provide a foundation for the future development of new FXS treatments.
| Identifer | oai:union.ndltd.org:mcmaster.ca/oai:macsphere.mcmaster.ca:11375/26991 |
| Date | January 2021 |
| Creators | Reynolds, Kathryn |
| Contributors | Scott, Angela, Neuroscience |
| Source Sets | McMaster University |
| Language | English |
| Detected Language | English |
| Type | Thesis |
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