Immediate early genes (IEGs) such as c-fos and Arc play important roles in translating synaptic inputs into transcriptional events, thus providing a mechanism for encoding neural plasticity and information storage. In addiction neuroscience, IEG expression is widely used to identify brain regions that become active in response to specific stimuli or during certain addiction associated behaviors. Thus, accurate brain-wide monitoring of IEG response is essential to discovering neural impact of drug exposure and molecular bases of addiction related behavioral patterns. However, the technology for noninvasive monitoring of IEG activity throughout large regions of the living brain is still incipient. To this end, our goal is to engineer genetically encoded metal-free MRI reporters for brain-wide monitoring of IEG activity in vivo. The technology we propose is based on aquaporins, which we recently introduced as genetic reporters for diffusion based MRI. By expressing aquaporins in cells, it becomes possible to create a mismatch in water diffusion between aquaporin-expressing cells and the surrounding milieu, which permits detection using diffusion weighted MRI. Here, we propose to adapt this mechanism for monitoring evoked c-fos and Arc activity in neuronal cultures. Our work will be accomplished via two aims. In Aim 1, we will harness aquaporin biodiversity and intracellular trafficking mechanisms to identify and engineer new reporter architectures that exceed the diffusion enhancement obtained with our first generation reporter, AQP1. Increasing the diffusion gain will enable neural responses to be detected with high molecular sensitivity and across a wider dynamic range of molecular inputs. In Aim 2, we will adapt the resulting technology for imaging pharmacologically evoked induction of c-fos and Arc activity in primary neurons. The accuracy of our new measurement technique will be validated by comparing with parallel measurement of c-fos and Arc dynamics using an established luciferase reporter. Ultimately, the MRI reporters developed in this project will be useful for revealing brain-wide IEG response profiles that are inaccessible to existing methods such as fMRI, MEMRI, and reporter gene techniques. In addition, the imaging methods described here can be readily adapted in larger animals (e.g., nonhuman primates) for investigating complex addiction associated behavioral patterns.