Biosensors bring together the fields of protein engineering, fluorescence imaging and cell biology to offer a rich and colourful toolbox capable of probing complex biological systems in order to understand the molecules and mechanism(s) underlying complex processes such as learning and memory. Glutamate has been at the centre of fluorescent biosensor development in the study of neural systems; however neurotransmission occurs as a closely regulated interplay of multiple neurotransmitters. While the identity of these molecules is known, we lack the spatial and temporal knowledge required to truly define their role in these signalling processes. In our lab we are working to create a robust neuro-sensor toolbox using a combination of protein engineering techniques such as ancestral reconstruction, circular permutation and rational design with fluorescence techniques such as FRET. These fluorescent sensors are intended to be specific for a range of neurotransmitters such as D-serine, Glycine, GABA and Arginine utilising the conserved scaffold of periplasmic binding proteins. In defining the role of these molecules the sensors provide a platform with which to create a general model of signalling dynamics allowing for the study and better interpretation of aberrant systems that could potentially occur in neurological disorders.