In vivo astrocyte calcium activity in the somatosensory cortex during locomotion and non-convulsive seizures

Abstract

Astrocytes make up a significant portion of the total cells in the brain. Intracellular Ca2+ changes in astrocytes have been attributed to various physiological processes that directly influence neuronal activity and brain network synchrony. Moreover, altered Ca2+ patterns are indicative of astrocyte dysfunction and of diseased states.

Astrocytes are active following sensory stimulation, such as visual cues and tactile whisker stimuli. Astrocytes participate in the cortical response to sensory input and synchronously respond to locomotion with an intracellular Ca2+ increase, predominantly modulated by widespread neuromodulator release and direct activation of astrocyte α1-adrenergic receptors. However, an in-depth study focussed on astrocyte activity in the primary somatosensory cortex (S1) during locomotion is lacking. To address this gap in the field, we recorded astrocyte Ca2+ changes in head-restrained, awake mice that were free to run on a wheel. Animals were chronically implanted with ECoG electrodes and a cranial window above S1. Ca2+ changes were visualised using a virally delivered genetically encoded Ca2+ indicator, GCaMP6f, driven by a GFAP promoter for selective expression in astrocytes. Semi-automated Ca2+ analysis protocols were used to measure Ca2+ fluorescence within the different sub-cellular compartments. We show that layer 2/3 astrocytes rapidly respond to locomotion onset, within a sub-second timescale. This was most evident in the cell periphery that activated most consistently to locomotion, even when soma activation was weak or absent. The Stargazer mouse, a model of ataxia, maintained a strong astrocyte response to locomotion, which was enhanced in view of its quicker astrocyte response time, especially in the cell periphery.

Astrocytes regulate neuronal network excitability and modulate cortical state switching and network synchrony. Astrocytes are capable of modifying neuronal activity through gliotransmitter release and have been consequently shown to elicit epileptic discharges in slice models, as well as precede epileptic events in in vivo epilepsy models. During absence seizures (AS), an increased rhythmicity and synchrony among thalamic and cortical neurons is evidenced by generalised spike and wave discharges (SWD) throughout this network. fMRI studies have revealed potential astrocyte recruitment preceding seizure onset and astrocyte GABA dysfunction is known to contribute to SWD occurrence. However, astrocyte calcium activity during AS is not known. We imaged astrocyte Ca2+ changes to determine if astrocytes were indeed active prior to seizure onset and potentially drove the neuronal paroxysmal activity of AS. We showed for the first time, astrocyte Ca2+ activity in two genetic animal models of AS. Stargazer mice exhibited enhanced microdomain and endfoot activation during periods of prolonged ictal ECoG activity, but no clear widespread astrocyte activation preceded seizure onset. Astrocytes within the cortical initiation site of AS were imaged in the Genetic Absence Epilepsy Rat from Strasbourg, using the Inscopix miniature head-mounted microscope. Astrocytes within the cortical initiation site engaged in a synchronous Ca2+ pattern during a state of prolonged seizure activity. Moreover, astrocyte-specific chemogenetic activation or IP3 receptor type 2 knockout, did not influence seizure occurrence.

In conclusion, we show a diverse and complex astrocyte response to locomotion which is driven by microdomain activation with potential summation of inputs that leads to widespread soma and endfoot activation. Furthermore, evidence we obtained from two well-established models of AS further strengthens past findings of dysfunctional astrocytes within the cortico-thalamic network involved in AS initiation, and we reveal novel astrocyte Ca2+ patterns associated with spontaneous generalised seizures.

Date of Award	Sept 2021
Original language	English
Supervisor	Rhein Parri (Supervisor) & Michael Coleman (Supervisor)