Neural Activation of Human Primary Intestinal Enteroendocrine L-cells

Intestinal enteroendocrine L-cells play a central role in metabolic homeostasis by secreting the gut hormones GLP-1 and PYY, but whether and how neural signals regulate human L-cell function remains poorly understood. This gap reflects the limited availability of human model systems enriched for mature L-cells. Here, we establish a differentiation strategy that generates human donor-derived ileal organoids with markedly increased L-cell abundance, enabling functional interrogation of L-cell physiology in their basal state. Using this system, we show that cholinergic and adrenergic stimuli activate L-cells, triggering calcium influx and secretion of GLP-1 and PYY. Pharmacological analysis reveals that cholinergic regulation of human L-cells is mediated predominantly through M3 muscarinic receptors, while adrenergic regulation is mediated through the β1 adrenergic receptor. Neurotransmitter-evoked activation also initiates gap junction-dependent calcium waves that propagate across the intestinal epithelium; however, disruption of epithelial coupling by abolishing calcium propagation does not impair hormone secretion. Together, these findings identify L-cells as integrative hubs that couple neural signals to gut hormone secretion and epithelial calcium waves, providing mechanistic insight into the gut-brain axis and how the nervous system regulates intestinal endocrine function in human intestine.