The human colonic stem cell niche provides a safe harbour for long-lived stem cells. Intestinal stem cells are responsible for driving tissue renewal and cellular differentiation that underpins intestinal barrier function.  Neuronal and non-non-neuronal (i.e. crypt autonomous) neurotransmitters are emerging as key regulators of physiological function in the stem cell niche.  Similarly, downstream calcium signals are emerging as central integrators of signals that regulate the physiology of the stem cell niche.  The aim of this study was to determine the mechanism of purinergic coupled calcium signals in the human colonic stem cell niche and investigate their physiological consequences. Methods: Human colonic crypts were isolated from colorectal tissue samples obtained at surgical resection (NREC approval) and cultured as crypts in the short term (or propagated as crypt-like organoids over the long-term (5 years). The spatio-temporal characteristics of intracellular calcium was monitored by Fura-2/Fluo-4/Calbryte-630 imaging and the mechanism of receptor-mediated calcium mobilisation was characterised by pharmacological and knockdown gene approaches. Calcium signalling toolkit expression was investigated by bulk RNAseq and visualised by fluorescence immunolabelling and super-resolution imaging. Mucus secretion was visualised by Muc2 immunofluorescence depletion assays and real-time imaging of fluorescently tagged mucin-2, using MUC2::mNEON crypt-like organoids generated by CRISPR-HOT. An organoid swelling assay was used as a proxy for fluid secretion. Results: Bulk RNA seq demonstrated mRNA expression of metabotropic P2Y1 (P2RY1), P2Y2 (P2RY2), P2Y11 (P2RY11) and P2Y13 (P2RY13) in native human colonic mucosa, isolated crypts, and cultured organoids.  A focus on P2Y2 immunolocalisation revealed expression on basolateral membranes lining the colonic crypt-axis.  P2Y2 co-labelled with LGR5+ stem cells along their basolateral membranes at the crypt-base, in addition to labelling MUC2-positive goblet cells, and  sparse numbers of chromogranin-positive enteroendocrine cells. Both ATP and UTP, a P2Y2-selective ligand, stimulated intracellular calcium levels in the absence of extracellular calcium, and was inhibited by the InsP3R receptor antagonists 2-APB (100 mM), but not by inhibitors of two pore channels, NED19 (250 μM or tetrandrine (20 μM). UTP induced a reduction of intracellular MUC2 immunofluorescence intensity in colonic crypt-bases and crypt-like organoids. Similarly, UTP stimulated a decrease in intracellular MUC2-mNEON fluorescence and a concomitant increase in  luminal MUC2-mNEON fluorescence.  UTP also promoted an increase in the cross-sectional area of closed organoids. UTP effects on intracellular calcium, intracellular and luminal MUC2 levels and organoid cross sectional area were abolished by the P2Y2 antagonist, AR-C118925XX (5 μM). These functional observations of fluid secretion are supported by basal expression of NKCC1 and apical expression of ANO1. Conclusions: Puringeric input stimulates P2Y2 receptor mediated calcium mobilisation from intracellular stores, mucus secretion and fluid secretion.  These effects suggest that purinergic input regulates the microenvironment of the human colonic stem cell niche.