Engineering vasoactive probes for brain-wide imaging of molecular signaling

Informazioni relative al progetto

AVATar

ID dell’accordo di sovvenzione: 101040901

DOI

10.3030/101040901

Data della firma CE 8 Marzo 2022

Data di avvio 1 Maggio 2023

Data di completamento 30 Aprile 2028

Finanziato da

• European Research Council (ERC)

Costo totale

€ 1 492 968,00

Contributo UE

€ 1 492 968,00

Coordinato da

MAX-PLANCK-GESELLSCHAFT ZUR FORDERUNG DER WISSENSCHAFTEN EV
Germany

Periodic Reporting for period 1 - AVATar (Engineering vasoactive probes for brain-wide imaging of molecular signaling)

Periodo di rendicontazione: 2023-05-01 al 2025-10-31

Sintesi del contesto e degli obiettivi generali del progetto

Brain function depends on spatiotemporally defined brain-wide signalling via molecules such as neurotransmitters. No current technology can measure signalling molecules throughout the brain with sufficient spatial and temporal resolution in living mammals. This poses a major roadblock for understanding how molecular neuronal communication coordinates whole-brain function.

Magnetic resonance imaging (MRI) currently provides the highest brain-wide resolution. Dynamic imaging of blood flow and oxygenation in the finely arborized vasculature, so-called functional MRI (fMRI), can visualize whole-brain function in mammals and humans. However, MRI is inherently insensitive, which precludes it from accessing neurotransmitter signalling that occurs at (sub)micromolar concentrations and fMRI does not provide molecular information underlying measured hemodynamic signals.

This project aims develop protein-based vasoactive sensors, called AVATar, that convert neurotransmitter signalling into hemodynamic contrast to visualize brain-wide molecular signalling dynamics that shape healthy and pathological brain function. The project is structured along three objectives:

1) Engineering AVATars that convert neurotransmitter signalling into hemodynamic signals.
2) Brain delivery via non-invasive routes.
3) Application for fMRI of brain-wide neurotransmitter signalling in rodents.

Lavoro eseguito dall’inizio del progetto fino alla fine del periodo coperto dalla relazione e principali risultati finora ottenuti

Objective 1 aims to develop neurotransmitter-sensing AVATars along two different designs featuring competitive or allosteric functional mechanisms. We succeeded in generating an allosteric AVATar that acts at nanomolar concentrations and responds to physiological neurotransmitter concentrations and generated all key building blocks of the competitive AVATar design and produced all parts for a serotonin-sensing competitive AVATar.

In objective 2, we achieved genetic encoding and virus-mediated expression of AVATars, which facilitates wide-field, minimally invasive brain delivery. In situ expression circumvent two challenges of alternative delivery through the bloodstream, namely homogenous brain delivery and potential peripheral effects of AVATars outside the brain.

Progress towards objective 3 was largely hampered by delayed approvals of animal protocols. We are currently establishing the fMRI methodology for imaging AVATars in vivo.

Progressi oltre lo stato dell’arte e potenziale impatto previsto (incluso l’impatto socioeconomico e le implicazioni sociali più ampie del progetto fino ad ora)

The AVATar technology is still at an early development stage, so assessing its impact is still speculative. That said, our achievements from the first 2-year reporting period represent major progress for the field of brain imaging and understanding of molecular brain function in several ways:

1) The allosteric neurotransmitter-sensing AVATar we developed shows an unprecedented combination of detectability at low doses and sensitivity towards its target neurotransmitter. This paves the way towards fMRI detection of neurotransmitter release at physiological levels, which has not been possible with current technologies. Upcoming in vivo experiments will assess the full potential of AVATars for such applications.

2) Our progress in making AVATars genetically encodable and express them in situ through adeno-associated viruses facilitates brain-wide targeting and functionality of these probes. With this, we circumvent a major roadblock of existing molecular MRI probes, namely delivering sufficient doses of the probe into the brain in a non-disruptive and homogenous manner. Genetic encoding has been the key advantage of fluorescent sensors and was the main driver behind the massive success of GFP-derived tools. Still, in situ expression of molecular MRI sensors has not been achieved so far.

Together, these achievements will, address a major challenge in neuroscience: enabling imaging of neurotransmitters throughout the entire mammalian brain will address long-standing research questions into the fundamental role of long-range molecular signalling patterns in brain function.