I am fascinated with how much we have learned about the function and architecture of inhibitory synapses at the molecular level and how much we still need to discover to understand how deficits in synaptic inhibition in our brain lead to neurological conditions.My research focuses on GABA-A and Glycine receptors, the major constituents of inhibitory synapses in the central nervous system. These receptors are pentameric ligand-gated ion channels that allow the passage of Cl- ions across the neuronal membrane. Their proper function is critical for the maintenance of appropriate neuronal excitability and consequently, changes in the inhibitory system are implicated in a range of neurological conditions including epilepsy, addiction, alcohol withdrawal syndrome, anxiety disorders, chronic pain, autism, spasticity, and autoimmune encephalitis. In most of these disorders we are only able to treat symptoms rather than the underlying cause of the disease and this is largely because we do not fully understand the relationship between inhibitory neuro-receptors and their interaction partners at inhibitory synapses.

Goals to achieve

I would like to better understand the molecular basis of neurological diseases by examining the relationships between function, cellular localisation and organisation of GABA-A and Glycine receptors, and how these properties change in neurological disorders. The overreaching aim is to better understand the underlying deficits at a molecular level to enable the identification of novel pharmacological targets for the development of clinically relevant strategies.

The approach

We use a combination of quantitative super-resolution microscopy and electrophysiology to gain a quantitative understanding of how molecules found in synapses drive neurological processes. The microscopy techniques include stochastic optical reconstruction microscopy (STORM), photoactivated localization microscopy (PALM), single particle tracking (SPT) and single step photobleaching. We also use various confocal microscopy approaches. With these methods, we can directly visualize proteins that are involved in different cellular processes, accurately measure the absolute number of molecules in protein clusters, follow molecular interactions on relevant time scales (10 ms to 1 s), and reconstruct synaptic architecture with localisation precision comparable to the size of a single inhibitory neuro-receptor (~ 10 nm). As new tools facilitate new biology, our efforts also go towards the development of new methods that aim to overcome the limitations of current techniques and help us visualize action of molecular complexes in the cell in real time.

GABA-A a Glycine receptors are targets of many clinically important drugs including neurosteroids, barbiturates, benzodiazepines, and general anaesthetics. Electrophysiology allows us to test channel function and the effects of drugs. We are able to characterise inhibitory synaptic currents mediated by GABA-A and Glycine receptors with desired subunit composition using synapses formed between HEK293 cells and neuronal presynaptic terminals ("artificial synapses"). This system is particularly useful when testing the impact of genetic mutations on channel function as neuronal postsynaptic terminals contain many neuroreceptor subtypes and the properties of synaptic currents in neurons reflect that diversity. The "artificial synapse" system allows the recording of inhibitory synaptic currents mediated by the receptors containing disease-associated subunits in isolation from other subtypes. We routinely use this technique to understand the functional properties of Glycine and GABA-A receptor variants found in hereditary neurological disorders and to test how clinically relevant drugs modulate their properties.

Biography

Nela Durisic obtained a PhD degree from McGill University in Canada where she used quantum dots and fluorescence fluctuation techniques to show that the fluorescent emission of quantum dots can be used to measure intracellular oxygen content. Upon completion of her PhD, she joined the Laboratory of Melike Lakadamyali (currently University of Pennsylvania) where she developed a technique for direct counting of proteins in small clusters using quantitative PALM microscopy and used this technique to count the number of α1 and β subunits in Glycine receptors. For her second postdoctoral training, she joined the laboratory Joe Lynch (Emeritus Professor, Queensland Brain Institute) to study the functional properties of inhibitory neuro-receptors. Since June 2021, Dr Durisic is running an independent research program at Queensland Brain Institute.