Neurosteroids and GABA-A receptors and their roles in neuropsychiatric disorders

The brain receives and sorts information by individual nerve cells communicating with each other using a process known as chemical neurotransmission. This involves networks of cells becoming broadly excited or inhibited to control key aspects of mental function such as memory and the overall behaviour of individuals. The basis for this is the release of chemicals (neurotransmitters) from one nerve cell that binds to specific protein receptors on another, to cause excitation or inhibition. One major neurotransmitter in the brain is GABA. It is responsible for dampening nerve cell excitation by causing inhibition - it does this by interacting with a class of receptors known as GABA-A receptors (GABA-ARs). Most nerve cells in the brain will possess GABAARs underlining the pivotal role that they play in fundamental aspects of brain function including the following examples of - cognition, memory, sleep, sensation, control of muscle movement and response to stress. Indeed, so important are these receptors that their dysfunction is directly implicated in numerous and serious neurological and mental health disorders, such as anxiety and depression, epilepsy, schizophrenia, pain, substance addiction, autism, Down's and Fragile-X syndromes, to name only a few. As a consequence, GABAARs are widely recognised as molecular targets for therapeutic drug classes that include the benzodiazepines (BDZs) and general anaesthetics.

Of equal significance, GABA-ARs are also subject to regulation by a number of endogenous agents in the brain. Prime amongst these are the neurosteroids, which are released from cells and can potentiate GABA-AR function at very low concentrations. Dysfunctional neurosteroid production is also implicated in neurological diseases, such as anxiety and depression, pain and catamenial epilepsy, and this is thought to involve altered GABA-AR function. It is therefore critical to understand how neurosteroids regulate GABA-ARs, what neural circuits are involved, how this contributes to disease, and how neurosteroids may be used as therapeutic agents. Our recent breakthrough in identifying the molecular basis for neurosteroid binding sites on GABA-ARs enables us to address these questions and determine precisely which types of GABA-ARs are involved in particular actions of the neurosteroids.

Our aims are to use a combination of techniques, some involving genetic engineering, to determine how neurosteroids regulate selected GABA-ARs and to broaden our understanding of neuropsychiatric disease when neurosteroid regulation becomes dysfunctional. We will use the neurosteroid modulatory site on GABA-ARs to facilitate the development of novel therapeutics for the treatment of neurological disorders. Our aim is to improve understanding of mechanisms that may precipitate mental illness and to provide new approaches to its prevention and treatment.

Specifically, we have four goals - (i) to examine what actions of the neurosteroids are mediated by a specific isoform of the GABA-AR that is involved in mediating anxiety and pain; (ii) to understand how modification of GABA-ARs by naturally-occurring enzymes in the brain affects neurosteroid modulation; (iii) to probe these interactions with neurosteroids at another important GABA-AR that is involved in dampening excitation and is implicated in substance addiction; and (iv) to examine how we can develop new neurosteroid agents for therapeutic benefit.

The outcomes from our programme of work will have many implications. Chiefly, it will provide much needed information on the basis by which neurosteroids work in the brain, how they regulate GABA-ARs under physiological conditions and what happens during dysfunction to precipitate disease. By exploring the neurosteroid binding site, we now have for the first time, an opportunity to provide new impetus for the treatment of neurological diseases.

We will examine how neurosteroids exert their effects via specific GABA-AR isoforms and the consequences for neuropsychiatric disease when this becomes dysfunctional. Our aim is to better understand some of the mechanisms that precipitate mental illness and to provide new approaches for its prevention and treatment by exploiting our knowledge of the neurosteroid modulatory site on GABA-ARs. A combination of cellular, molecular and genetic techniques will examine the roles of neurosteroids using recombinant GABA-ARs in expression systems and native GABA-ARs in cell cultures or brain slices.

Whole-cell patch clamp recording methods will measure GABA synaptic and tonic inhibition and associated kinetic parameters. Biochemical (Western blots, phosphorylation) and immunocytochemical techniques (antibody labelling, phosphorylation-specific antibodies, Tirf microscopy) will measure changes to GABA-AR composition, and single particle tracking with quantum dots and the bungarotoxin binding site will monitor GABA-AR mobility. New transgenic animals will help to explore the interactions between GABA-AR phosphorylation and neurosteroid modulation with regard to synaptic and tonic inhibition.

Behavioural techniques will examine how neurosteroids affect intact brain function and the consequences that follow when this becomes dysfunctional. We will use structure-function approaches to probe the neurosteroid binding site and transplant it into other receptors to gain valuable information on key structural determinants. We will design (collab. with UCL Chemistry and Industry) new neurosteroid molecules that could be therapeutically beneficial by targeting specific isoforms of the GABA-AR.

Our research programme will contribute to our understanding of brain function. It will help us to understand how neurological diseases can be precipitated by dysfunctional neurosteroid signalling, as well as providing new insight into novel therapeutic strategies for mental health.

For Academic, Economic and Societal Impact, our proposed programme is likely to have broad-ranging effects on several groups. These include academics, and in the longer-term, healthcare professionals associated with mental health. We also expect the general/lay public who are directly or indirectly affected by poor mental health to benefit, as well as pharmaceutical companies concerned with drug development for mental health disorders.

Academics will gain valuable insight into precisely how neurosteroids function in the brain as well as achieving a better understanding of their role in mental illness. We will engender Capacity Building Impact by training new generations of scientists to use innovative technologies and methods, as well as providing clinicians with potentially new therapeutic targets for treating mental illness. Our previous collaborative studies of inhibitory synaptic transmission provide direct evidence of translation relevant to inherited neurological disorders by providing molecular diagnostics and potential therapeutic remedies (Rees et al 2006, Nature Genet 38, 801-806). In the longer term, we predict our programme has the potential to lead to improved quality of life for patients, family members, carers and indeed medical staff.

Our research programme is designed to enable a greater understanding of brain function by providing new knowledge regarding GABA-mediated synaptic and tonic inhibition and how modulation by neurosteroids can cause and affect neuropsychiatric disease. These findings are expected to be at least partly-translated across scientific disciplines during the time-course of the programme. We will ensure promulgation of our findings by presentations at major international conferences, by engaging in invited lectures in the UK and abroad to scientific and lay audiences, by publishing in internationally-recognised high quality scientific journals, and by engaging with national/international press, television and radio as appropriate.

If patentable results arise from our research programme, and this is potentially likely, UCL has excellent support structures to enable us to proceed and exploit our discoveries. Indeed, with regard to the discovery of neurosteroid binding sites, a patent has been issued with a view to encouraging interactions with the pharmaceutical industry.

In the longer term, understanding how neurosteroids and GABA receptors contribute to neurological disease could improve our identification of new components involved in disease pathology. This is particularly relevant for an ageing population where the prevalence of neural diseases is projected to increase and is in urgent need of new therapeutic initiatives.