Paneth cells (PCs) are terminally differentiated, highly specialized secretory cells located at the base of the crypts of Lieberkühn in the small intestine. Besides their antimicrobial function, PCs serve as a component of the intestinal stem cell niche. By secreting granules containing bactericidal proteins like defensins/cryptdins and lysozyme, PCs regulate the microbiome of the gut. Here we study the control of PC degranulation in primary epithelial organoids in culture. We show that PC degranulation does not directly occur upon stimulation with microbial antigens or bacteria. In contrast, the pro-inflammatory cytokine Interferon gamma (IFN-γ) induces rapid and complete loss of granules. Using live cell imaging, we show that degranulation is coupled to luminal extrusion and death of PCs. Transfer of supernatants from in vitro stimulated iNKT cells recapitulates degranulation in an IFN-γ-dependent manner. Furthermore, endogenous IFN-γ secretion induced by anti-CD3 antibody injection causes Paneth loss and release of goblet cell mucus. The identification of IFN-γ as a trigger for degranulation and extrusion of PCs establishes a novel effector mechanism by which immune responses may regulate epithelial status and the gut microbiome.