Browsing by Subject "Astrocytes"

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    Astrocyte-neuron signaling in the nucleus accumbens: implications for brain reward signaling
    (2019-05) Corkrum, Michelle
    Dopamine is one of the major reward signaling molecules in the brain. Dopaminergic transmission contributes to physiological states such as learning, memory encoding, movement and motivated behaviors; and, the disruption of dopamine signaling can contribute to neuropsychiatric diseases such as substance use disorders. The majority of research on reward signaling has focused on neurons; however, astrocytes are emerging as key components of brain information processing. Astrocytes are a subset of glial cell, one of the most abundant cell types in the brain. Although astrocytes are not electrically excitable, in response to brain activity, they demonstrate increases in intracellular calcium and the subsequent release of neuroactive substances, termed gliotransmitters. Therefore, my dissertation aimed to investigate the hypothesis that astrocytes respond to brain reward signaling with elevations in cytoplasmic calcium, and subsequently modulate neuronal activity in the nucleus accumbens, one of the major reward centers of the brain. Utilizing fiber photometry, I found that astrocytes in the nucleus accumbens respond to dopamine and amphetamine with cytoplasmic calcium elevations in vivo. To elucidate the cellular mechanisms of this phenomenon and the consequences of astrocyte calcium signals on neuronal activity, we conducted experiments applying multiphoton calcium imaging and whole-cell patch clamp electrophysiology in acute brain slices containing the nucleus accumbens core. We found that astrocytes respond to dopamine, amphetamine and opioids with intracellular calcium elevations and subsequently modulate neuronal activity, either through adenosine signaling in the case of dopamine and amphetamine or glutamatergic signaling in the case of opioid exposure. Furthermore, we demonstrate that astrocytes contribute to the acute psychomotor behavioral effects of amphetamine, illustrating astrocyte modulation of drug-related behaviors. Overall, the current body of work provides evidence that astrocytes actively contribute to brain reward processing via responding to dopamine and drugs of abuse with intracellular calcium increases and modulating neuronal and synaptic activity in the nucleus accumbens, one of the major nodes of the reward system.
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    Proneural transcription factor NeuroD1-mediated direct neuronal reprogramming - an AAV approach
    (2019-12) Radha, Swathi
    Direct cellular reprogramming to drive lineage switching from one differentiated cell type to another can be exploited to develop cell-based therapies for neurodegenerative diseases. In vivo reprogramming provides an attractive therapeutic strategy to circumvent the hurdles of immune rejection and ethical constraints associated with transplant-based therapy. Supporting glial cells of the CNS can be reprogrammed to neurons by targeted viral delivery of transcription factors and small molecules. Previous studies have demonstrated the effectiveness of a single proneural transcription factor NeuroD1 to drive reprogramming in the reactive glial lesions of Alzheimer’s disease and stroke. However, the ability of NeuroD1 (ND1) to promote a similar benefit in models of Parkinson’s disease (PD) has yet to be demonstrated. The current study aims to test the hypothesis that NeuroD1 delivered via an Adeno-Associated Virus (AAV) can promote reprogramming in striatal astrocytes to neurons in an in vivo PD model. The FDA-approved, clinically employed AAV-9 gene delivery platform was used to transduce non-dividing cells with minimal off-target effects. A two-part AAV-9 viral system was designed to express ND1 in astrocytes, driven by the GFAP promoter. First, ND1-mediated direct neuronal reprogramming was tested using a simple and scalable in vitro culture system. Primary astrocytes in vitro transduced with the AAV9-ND1 dual virus system display characteristics of immature neuroglial precursor stage, suggesting successful reprogramming. Second, the potential of ND1 to drive reprogramming was assessed in vivo in mice. Intracranial and intravenously delivered AAV-9 dual virus system driven by the GFAP promoter targets astrocytes and surprisingly, mature resident neurons in vivo. Finally, ND1-mediated in vivo reprogramming was assessed in a well-established chemically-induced 6-hydroxydopamine (6-OHDA) Parkinson’s disease mouse model. The 6-OHDA injury model provides insight into the novel application of the AAV-9 dual virus system to target astrocyte-to-neuron reprogramming as well as to target resident neurons for potential neuronal repair in Parkinson’s disease.