Mapping and specific viral targeting of peripheral pancreatic innervation

Neuronal inputs from central nervous system (CNS) are essential in regulating pancreatic endocrine function. It has been shown that perturbation of CNS activity alters pancreatic hormone release. CNS and peripheral organs constantly communicate through the peripheral nervous system (PNS), which can be divided into autonomic efferent (parasympathetic and sympathetic) and afferent sensory nervous systems. While we know that the pancreas is densely innervated by the PNS, we lack a comprehensive map of peripheral ganglia providing inputs to the pancreas. These data could provide crucial information about pancreatic function. Published studies suggest parasympathetic inputs to the pancreas lower blood glucose by enhancing insulin secretion. On the other hand, sympathetic action results in increased blood glucose levels by stimulating glucagon secretion and inhibiting insulin. However, these studies are based on electrical stimulation or transection of peripheral splachnic and vagal nerves that may affect the activity of other metabolically active organs (liver, stomach, intestine,). Highly targeted modulation of specific pancreatic nerve activity in health and pancreatic diseases requires new approaches. In the present project, we provide a detailed map of the distribution of peripheral pancreas-innervating neurons using retrograde tracing, tissue clearing and 3D imaging techniques (Idisco+). We show that a substantial pancreatic innervation (Fig 1b) with significant neuronal populations in sympathetic, parasympathetic and sensory ganglia projecting to pancreatic tissue (Fig 1 c,d). In addition, we achieved targeted gene delivery to specific pancreatic efferent and afferent pathways by optimizing adeno-associated viral serotypes (AAV), promoters, titers and delivery (Fig2 a-d). By utilizing our optimized approach, we show that highly targeted chemogenetic neural activation of intrapancreatic parasympathetic cholinergic neurons increased plasma insulin and significantly improved glucose tolerance in male mice (Fig 2 e,f). Additionally, using a dual viral strategy to specifically target pancreas-projecting sympathetic neurons, we demonstrated that targeted activation of pancreas-projecting sympathetic neurons impaired glucose tolerance in male mice (Fig 2g,h). Our innovative viral approaches allow highly targeted gene expression and neuromodulation of defined pancreatic nerves. These methodologies will allow examination of functional roles of pancreatic parasympathetic and sympathetic innervation in glucose metabolism as well as in digestion (exocrine function). Our work also allow for future studies to determine the specific contribution of pancreatic innervation to metabolic diseases, pancreatic inflammation and cancer.