Astrocytes are ubiquitous in the central nervous system. They possess thousands of individual processes, interacting with neurons, other glia and blood vessels.
Paralleling the wide diversity of their interactions, astrocytes have general roles in supporting central nervous system structure, metabolism, blood-brain-barrier formation and control of vascular blood flow, axon guidance, synapse formation and modulation of synaptic transmission. Effectively all central nervous system diseases show astrocyte involvement, and accumulating evidencen suggests that reactive astrocytes are directly neurotoxic and present a unique therapeutic target.
By determining the molecular mechanisms that underlie astrocyte function, we hope to gain a better understanding of the disease mechanims underlying many neurological disorders, opening up exciting new therapeutic strategies.
Our research aims are to understand the basis of astrocyte-neuron interactions contributing to both the formation and function of the so-called ‘tripartite’ synapse.
We have a particular interest in how astrocyte heterogeneity differentially affects these processes. By providing a mechanistic understanding of how the tripartite synapse functions, our overall objective is to provide an integrated view of central nervous system circuit function in healthy tissue.
This will help us understand how aberrant astrocyte function, and its impact on local neuronal activity, contributes to diseases such as epilepsy, trisomy 21 and Alzheimer’s disease.
The tripartite synapse: from transcripts to function
Our main experimental tissue is the mouse primary visual cortex, as most cortical synapses are ‘tripartite’. However, some experiments are also performed in hippocampus. Projects combine a wide range of techniques, allowing a full characterization of the tripartite synapse from transcripts to function:
- We use structural imaging using iterative expansion microscopy to increase resolution.
- Molecular profiling using cell-type specific transcriptomics (single cell sequencing, translating ribosome affinity purification (TRAP) methodologies and large-scale in situ hybridization).
- We do functional studies in acute tissue slices, using 2-photon imaging of Ca2+ indicators and electrophysiology (patch-clamp and microelectrode arrays: MEAs). A number of our experiments use in vivo imaging through a cranial window.
- In addition to classical approaches to gene manipulation, based on mouse genetics, we rely heavily on the flexibility of adeno-associated virus (AAV)-based vectors to manipulate cells, following local (AAV9) or systemic (AAV-PHP.B) injection. Examples include the expression of genetically encoded Ca2+ indicators (GECIs: GCaMP6 and RCaMP1), as well as proteins allowing cell-type specific activation or inhibition (OptoXR and Designer Receptors Activated by Designer Drugs: DREADDs). NA, noradrenaline, NAR, noradrenaline receptor.
How do astrocytes contribute to the specificity of synapse formation?
Physical formation of synapses is a multi-step process, requiring neuronal arborization and precise registration of pre- and post-synaptic elements. Several protein superfamilies are known to mediate these processes, but in comparison, relatively little is known about the astrocyte-specific mechanisms that contribute to formation of the tripartite synapse.
We recently reported that molecularly distinct astrocytes reside in specific layers of both the mouse and human cortex (Bayraktar et al., Nat Neurosci, 2020) and now want to explore how differential expression of synapse mediators contribute to the formation of tripartite synapses.
How do astrocytes influence synaptic activity?
A principle pillar of the tripartite synapse concept is that astrocytes respond to local synaptic activity with elevations in intracellular Ca2+. This increase triggers the release of small neuroactive molecules – so-called gliotransmitters (such as glutamate, GABA and ATP) – which in turn modulate synaptic activity. However, astrocytes can also respond to neuromodulators, such as noradrenaline.
Neuromodulators are released globally throughout the neuropil, as a result of changes in arousal, exerting widespread effects on neuronal circuits. Interestingly, our transcriptomics data has consistently pointed towards a role for astrocytes in mediating the effects of noradrenaline on local circuits. A recent in vivo imaging study from our lab (Slezak et al., Curr Biol, 2019) demonstrated that astrocytes in mouse visual cortex respond reliably and robustly to local neuronal activity (triggered by visual stimuli), but that these responses are critically dependent on noradrenaline. This suggests that astrocytes act as signal integrators, operating across multiple signaling modalities, with potential consequences for synaptic activity and ultimately behavior.
How do astrocytes contribute to plasticity?
Neuronal circuits can change at both the structural and functional level – so-called plasticity. In the adult brain, plasticity in the visual cortex occurs following monocular enucleation, which deprives the contralateral visual cortex of its principal afferent input, resulting in a massive drop in neuronal activity. In the ensuing weeks, the unused visual cortex undergoes two plasticity processes. The first involves the expansion of the neighboring binocular cortical territory into the sensory input-deprived monocular visual cortex, and is driven by the potentiation of neuronal responses to inputs from the spared eye. The second type of reactivation depends on somatosensory inputs from the whiskers, and models the reallocation of cortical circuits to a new type of sensory stimulus. Our recent work suggests that this form of adult plasticity is critically dependent on astrocytes (Hennes et al., Glia, 2020).
Astrocyte numbers increase immediately following monocular enucleation and persist for months. We aim to fully elucidate the mechanisms driving astrocyte-mediated recovery of neuronal activity, studying plasticity at all levels: from the molecular and structural changes underlying the process through to modifications of cellular activity in vivo. We collaborate with the labs of Lut Arckens (KU Leuven) and Vincent Bonin (VIB NERF), who bring specific expertise in visual cortex physiology and in vivo imaging.
Using single cell sequencing approaches, we want to shed light on the molecular mechanisms by which astrocytes instruct the plasticity process - for example, modulation of signaling pathways associated with synapse removal and remodeling. A major interest in the lab is also whether these ‘new’ astrocytes represent ‘juvenile’ or ‘adult’ cells and whether transcriptome state is dynamic and causally involved in visual cortex plasticity.