Spatiotemporal organization of prefrontal norepinephrine influences neuronal activity.

Norepinephrine (NE), a neuromodulator released by locus coeruleus (LC) neurons throughout cortex, influences arousal and learning through extra-synaptic vesicle exocytosis. While NE within cortical regions has been viewed as a homogenous field, recent studies have demonstrated heterogeneous axonal dynamics and advances in GPCR-based fluorescent sensors permit direct observation of the local dynamics of NE at cellular scale. To investigate how the spatiotemporal dynamics of NE release in the prefrontal cortex (PFC) affect neuronal firing, we employed in vivo two-photon imaging of layer 2/3 of PFC in order to observe fine-scale neuronal calcium and NE dynamics concurrently. In this proof of principle study, we found that local and global NE fields can decouple from one another, providing a substrate for local NE spatiotemporal activity patterns. Optic flow analysis revealed putative release and reuptake events which can occur at the same location, albeit at different times, indicating the potential to create a heterogeneous NE field. Utilizing generalized linear models, we demonstrated that cellular Ca2+ fluctuations are influenced by both the local and global NE field. However, during periods of local/global NE field decoupling, the local field drives cell firing dynamics rather than the global field. These findings underscore the significance of localized, phasic NE fluctuations for structuring cell firing, which may provide local neuromodulatory control of cortical activity. Significance Statement NE is a neuromodulator which plays a critical role in learning and arousal, but understanding its spatial scale has been limited by technical barriers. Here, we utilized two-photon imaging of GPCR-based sensors, light sheet imaging, and computational modeling to gain insight into the fine scale organization of NE in PFC. We found that NE can influence neuronal activity at a local scale within cortex, which has not been shown before, and we developed new computational approaches to analyzing two-photon imaging of GPCR based fluorescent sensors. This insight will facilitate improved understanding of NE's role in motivated behaviors, as well as new approaches for understanding local neurotransmitter function.