Intravital microscopy is a powerful technique to observe dynamic processes with single-cell resolution in live animals. No intravital window has been developed for imaging the colon due to its anatomic location and motility, although the colon is a key organ where the majority of microbiota reside and common diseases such as inflammatory bowel disease, functional gastrointestinal disorders, and colon cancer occur. Here we describe an intravital murine colonic window with a stabilizing ferromagnetic scaffold for chronic imaging, minimizing motion artifacts while maximizing long-term survival by preventing colonic obstruction. Using this setup, we image fluorescently-labeled stem cells, bacteria, and immune cells in live animal colons. Furthermore, we image nerve activity via calcium imaging in real time to demonstrate that electrical sacral nerve stimulation can activate colonic enteric neurons. The simple implantable apparatus enables visualization of live processes in the colon, which will open the window to a broad range of studies.

Epidemiological and histopathological findings have raised the possibility that misfolded alpha-synuclein protein might spread from the gut to the brain and increase the risk of Parkinsons disease (PD). While past experimental studies in mouse models have relied on gut injections of exogenous recombinant alpha-synuclein fibrils to study gut to brain alpha-synuclein transfer, the possible origins of misfolded alpha-synuclein within the gut have remained elusive. We recently demonstrated that sensory cells of the gut mucosa express alpha-synuclein. In this study, we employed mouse intestinal organoids expressing human alpha-synuclein to observe the transfer of alpha-synuclein protein from gut epithelial cells in organoids co-cultured with vagal nodose neurons that are otherwise devoid of alpha-synuclein expression. In intact mice that express pathological human alpha-synuclein, but no mouse alpha;-synuclein, alpha-synuclein fibril templating activity emerges in alpha-synuclein seeded fibril aggregation assays in tissues from the gut, vagus nerve, and dorsal motor nucleus. In newly engineered transgenic mice that restrict pathological human alpha-synuclein expression to intestinal epithelial cells, alpha-synuclein fibril-templating activity transfers to the vagus nerve and to the dorsal motor nucleus. Subdiaphragmatic vagotomy prior to the induction of alpha-synuclein expression in the gut epithelial cells effectively protects the hindbrain from the emergence of alpha-synuclein fibril templating activity. Overall, these findings highlight a novel potential non-neuronal source of fibrillar alpha-synuclein protein that might arise in gut mucosal cells.