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  1. Oxidative stress - reactive oxygen species as novel plasticity signals in the nervous system
  2. Embryonic critical periods determine synaptic function and animal behaviour for later life
  3. Mechanisms of change – how critical period experiences specify and then maintain cellular properties
  4. Mechanisms of network adjustment during critical periods of nervous system development
  5. Modelling network adjustment during critical periods of nervous system development

Oxidative stress - reactive oxygen species as novel plasticity signals in the nervous system

High levels of reactive oxygen species (ROS) are a hallmark of aging and neurodegenerative conditions, leading to cellular damage and, eventually, cell death. Most ROS are generated as metabolic by-products of mitochondrial respiration. Given that the nervous system is the most energetically demanding organ, we wondered whether neurons might utilise ROS as a readout of their activity. Indeed, we discovered that ROS are both necessary and sufficient for adaptive changes, of synaptic terminal size and transmission

This was possible by being able to study identified neurons in their entirety, revealing that adjustments of presynaptic (neuromuscular junctions/NMJs) and postsynaptic (dendrites) terminals are coordinated. We further discovered the highly conserved redox-sensitive DJ-1b protein acting as a ROS sensor, and PI3Kinase signalling as the critical downstream pathway Oswald et al., 2018).

We are now investigating other sources of activity-induced ROS generation and the molecular pathways by which these bring about distinct structural changes to presynaptic NMJs and postsynaptic dendrites.

This is a longstanding BBSRC funded collaboration with Sean Sweeney, University of York, currently translated to a vertebrate system.

Image:Top: 3D reconstructions of aCC motoneurons from control and after progressive over-activation, leading to progressively smaller dendritic arbors. Below: the opposite growth phenotype (more but smaller boutons) occurs in the periphery at the neuromuscular junction (NMJ) [Matt Oswald].

 

Embryonic critical periods determine synaptic function and animal behaviour for later life

Neurons, and the networks in which they exist, function within a physiologically appropriate activity range, often referred to as ‘homeostatic set-point’. During development, after much of the network architecture and basic connectivity has been established, a temporally restricted phase of heightened plasticity sets in, referred to as the Critical Period, during which this activity set-point is specified.

Critical period experience, including the temperature that embryos are exposed to, nerve cell activity or levels of reactive oxygen species experienced, all provoke differential set-point adjustments in response. This occurs both at the single-cell level, as well as across the network. We are exploring how critical period experience is translated into lasting adjustment at single-cell level, focusing upon the morphology, connectivity and physiology of a model synaptic terminal, the Drosophila neuromuscular junction (NMJ) in the larva (see below). Using transcriptomics we are exploring critical period-induced changes in gene expression that underpin changes in cellular properties (e.g. as measured by imaging, electrophysiology and behavioural analysis).

Image: Set-point adjustment at the NMJ. Following 18ºC critical period experience the eEJP (single evoked action potential) amplitude is reduced. A suction electrode (brown) is used to stimulate the nerve axon to induce single eEJPs, the amplitude of which is recorded using a sharp electrode (blue) in the muscle.

 

Importantly, transient experiences during the embryonic critical period have lasting effects on nervous system function, which manifest in dramatic changes of animal behaviour. In this case, how larvae crawl is significantly affected by the experience of the embryo – below: a critical period experience of 4 degrees warmer than control (lower animal) leads to larvae having calibrated on a faster crawling speed...but also a loss of several key plasticity mechanisms, making those animals inflexible to environmental change.

 

Mechanisms of change – how critical period experiences specify and then maintain cellular properties

How do transient experiences, during the critical period of nervous system development, lead to lasting changes of cellular properties? Critical periods are phases of nervous system development, when neuronal properties are specified and network function emerges. Their importance lies in the fact that transient disturbances during a critical period lead to lasting defects, or suboptimal nervous system function. In humans, epilepsy and several neuro-developmental psychiatric conditions are increasingly seen as caused by sub-optimal critical period experiences.

Working with the fruit fly as an experimental model system that has clearly defined critical period in late embryogenesis, we focus on two basic questions: 1. How are neuronal properties established during the critical period? 2. How are those subsequently maintained?

We find that during the critical period bio-physical aspects of the environment, such as temperature, are used as information. A few hours of being exposed to slightly cooler or warmer temperatures leads to changes in neuronal properties and animal behaviour. We think that the changes in gene expression we see might be maintained by changes in epigenetics marks. We are also investigating how those changes are initiated in the first place, looking at metabolic by-products as candidate instructive signals.

Image: Epigenetic marks at the neuromuscular junction - motoneuron terminal (grey) on a muscle with large nuclei (blue), including pattern of histone modification (white).

 

Mechanisms of network adjustment during critical periods of nervous system development

At the end of development of the Drosophila embryo, the nervous system is particularly plastic, sensitive to activity levels, and changes in the internal and external environment. We are investigating how such transient embryonic experiences, during the critical period, lead to morphological changes in nerve cell structure and connectivity. Our experimental approach is to subject embryos to changes during the critical period, for example manipulating temperature, activity or reactive oxygen species (ROS). We then use expansion microscopy (ExM) to achieve super-resolution that allows us to visualise and quantify numbers of connections (synapses) between specific interneurons and motor neurons.

Our project is a collaboration with the Richard Baines group at the University of Manchester, who specialise in electrophysiology. We envisage that the combination of high-resolution imaging to reveal structure and connectivity, paired with electrophysiology to assay function, will provide clarity on how networks adjust during this critical time window.

Image: Expansion microscopy (ExM) image of a neuron (magenta) taken on a spinning disk confocal microscope, showing presynaptic specialisations (Brp - cyan) in apposition to postsynaptic sites (Drep2 – yellow).

 

Modelling network adjustment during critical periods of nervous system development

Using computational modelling of the locomotor network of Drosophila larvae, we are investigating how cellular excitability, connectivity, and synaptic transmission might be changed during the critical period of nervous system development. If such periods of heightened/altered plasticity are needed for normal network function to emerge, then why are they closed during later life? How come errors that occur during the critical period lead to lasting network defects, in that subsequent homeostatic plasticity mechanisms cannot correct these?

Collaborators

Richard Baines (Faculty of Life Sciences, University of Manchester)

Jimena Berni (University of Sussex)

Albert Cardona (Department of Physiology, Development and Neuroscience, University of Cambridge)

Jan Felix Evers (Centre for Organismal Studies Heidelberg, Ruprecht-Karls-Universität, Heidelberg)

Gregory Jefferis (MRC-LMB, Cambridge; Department of Zoology, University of Cambridge)

Timothy O’Leary (Department of Engineering, University of Cambridge)

Tony Southall (Imperial College London)

Sean Sweeney (Department of Zoology, University of York)

Jelle van den Ameele (MRC Mitochondrial Biology Unit, Cambridge)

Marta Zlatic (Department of Zoology, University of Cambridge)

Positions

If you are interested in joining our group, please e-mail Matthias Landgraf to discuss research projects and positions/lab space. We have been successful in obtaining support for promising scientists and are happy to help.