combining-optogenetics-with-artificial-micrornas-to-characterize

Combining Optogenetics with Artificial microRNAs to Characterize the Effects of Gene Knockdown on Presynaptic Function within Intact Neuronal Circuits

The purpose of this protocol is to characterize the effect of gene knockdown on presynaptic function within intact neuronal circuits. We describe a ...

Istituto Italiano di Tecnologia

At synapses, pre- and post-synaptic cells get in direct contact with each other via cell adhesion molecules (CAMs). Several CAMs have been identified at the neuromuscular junction and at central synapses, where they regulate synaptic strength, by recruiting scaffolding proteins, neurotransmitter receptors and synaptic vesicles in response to the binding of counter-receptors across the synaptic cleft. Many synapses are also surrounded by astrocytic processes and embedded in conspicuous extracellular matrix (ECM). It is now widely recognized that astrocytes play a central role in regulating the synaptic machinery by exchanging information with the neuronal elements via diffusible molecules and direct physical interactions; this has lead to the concept of the 'tri-partite synapse'. More recently, the term 'tetra-partite synapse' has been introduced to underlie the importance of ECM in shaping synaptic function by mediating interaction and signaling between neurons and astrocytes. Here, we will review how this integrated view of the synapse can help us understand homeostatic synaptic plasticity at the neuromuscular junction and in the central nervous system. We will explore how synaptic CAMs regulate two forms of homeostatic plasticity: (i) postsynaptic scaling of synaptic currents to counteract changes in neuronal network activity and (ii) the compensatory modulation of presynaptic neurotransmitter release in response to changes in postsynaptic efficacy. We will discuss recent findings on activity-dependent trans-synaptic signaling events and the role of cell adhesion in the feedback control of network activity. This article is part of the Special Issue entitled 'Homeostatic Synaptic Plasticity'.

Cell adhesion and homeostatic synaptic plasticity.

α-Synuclein is thought to regulate neurotransmitter release through multiple interactions with presynaptic proteins, cytoskeletal elements, ion channels, and synaptic vesicles membrane. α-Synuclein is abundant in the presynaptic compartment, and its release from neurons and glia has been described as responsible for spreading of α-synuclein-derived pathology. α-Synuclein-dependent dysregulation of neurotransmitter release might occur via its action on surface-exposed calcium channels. Here, we provide electrophysiological and biochemical evidence to show that α-synuclein, applied to rat neurons in culture or striatal slices, selectively activates Cav2.2 channels, and said activation correlates with increased neurotransmitter release. Furthermore, in vivo perfusion of α-synuclein into the striatum also leads to acute dopamine release. We further demonstrate that α-synuclein reduces the amount of plasma membrane cholesterol and alters the partitioning of Cav2.2 channels, which move from raft to cholesterol-poor areas of the plasma membrane. We provide evidence for a novel mechanism through which α-synuclein acts from the extracellular milieu to modulate neurotransmitter release and propose a unifying hypothesis for the mechanism of α-synuclein action on multiple targets: the reorganization of plasma membrane microdomains.

Exogenous α-synuclein decreases raft partitioning of Cav2.2 channels inducing dopamine release.

Alternative splicing of pre-mRNAs is prominent in the mammalian brain, where it is thought to expand proteome diversity. For example, alternative splicing of voltage-gated Cachannel (VGCC) αsubunits can generate thousands of isoforms with differential properties and expression patterns. However, the impact of this molecular diversity on brain function, particularly on synaptic transmission, which crucially depends on VGCCs, is unclear. Here, we investigate how two major splice isoforms of P/Q-type VGCCs (Ca2.1[EFa/b]) regulate presynaptic plasticity in hippocampal neurons. We find that the efficacy of P/Q-type VGCC isoforms in supporting synaptic transmission is markedly different, with Ca2.1[EFa] promoting synaptic depression and Ca2.1[EFb] synaptic facilitation. Following a reduction in network activity, hippocampal neurons upregulate selectively Ca2.1[EFa], the isoform exhibiting the higher synaptic efficacy, thus effectively supporting presynaptic homeostatic plasticity. Therefore, the balance between VGCC splice variants at the synapse is a key factor in controlling neurotransmitter release and presynaptic plasticity.

Article	Year
Cell adhesion and homeostatic synaptic plasticity. Neuropharmacology\| PubMed ID: 23542441	2014
Exogenous α-synuclein decreases raft partitioning of Cav2.2 channels inducing dopamine release. The Journal of neuroscience : the official journal of the Society for Neuroscience\| PubMed ID: 25100594	2014
Alternative Splicing of P/Q-Type CaChannels Shapes Presynaptic Plasticity. Cell reports\| PubMed ID: 28700936	2017

Agnes Thalhammer

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