The pancreatic secretagogue cholecystokinin (CCK) is widely thought to stimulate enzyme secretion by acinar cells indirectly via activation of the vagus nerve. We postulate an alternative pathway for CCK-induced pancreatic secretion. We hypothesize that neurally related pancreatic stellate cells (PSCs; located in close proximity to the basolateral aspect of acinar cells) play a regulatory role in pancreatic secretion by serving as an intermediate target for CCK and secreting the neurotransmitter acetylcholine (ACh), which, in turn, stimulates acinar enzyme secretion. To determine whether PSCs (i) exhibit CCK-dependent ACh secretion and (ii) influence acinar enzyme secretion, primary cultures of human and rat PSCs were used. Immunoblotting and/or immunofluorescence was used to detect choline acetyltransferase (ACh synthesizing enzyme), vesicular ACh transporter (VAChT), synaptophysin, and CCK receptors 1 and 2. Synaptic-like vesicles in PSCs were identified by EM. ACh secretion by PSCs exposed to 20 pM CCK was measured by LC-MS/MS. Amylase secretion by acini [pretreated with and without the muscarinic receptor antagonist atropine (10 μM) and cocultured with PSCs] was measured by colorimetry. PSCs express ACh synthesizing enzyme, VAChT, synaptophysin, and CCK receptors; exhibit CCK-dependent ACh secretion; and stimulate amylase secretion by acini, which is blocked by atropine. In conclusion, PSCs express the essential elements for ACh synthesis and secretion. CCK stimulates ACh secretion by PSCs, which, in turn, induces amylase secretion by acini. Therefore, PSCs may represent a previously unrecognized intrapancreatic pathway regulating CCK-induced pancreatic exocrine secretion.acinar cells | cholecystokinin | pancreas T he pancreas is the major source of the body's digestive enzymes (1). The functional unit of the exocrine pancreas that produces these enzymes is the acinus, composed of individual acinar cells surrounding a central lumen. Acinar cells are capable of synthesizing and secreting as many as 10 million protein (enzyme) molecules per day. The process of enzyme synthesis and secretion at the acinar cell level is highly regulated and involves enzyme synthesis on the rough endoplasmic reticulum, posttranslational changes in the Golgi apparatus, and packaging into zymogen granules. Enzyme secretion is accomplished by the movement of zymogen granules to the apical pole of the acinar cell, fusion of the zymogen granule membrane with the apical plasma membrane followed by discharge of zymogen granule contents into the lumen, a process termed exocytosis (2).Postprandial secretion of pancreatic enzymes is predominantly mediated by regulatory peptides such as cholecystokinin (CCK) and by the vagovagal reflex, which activates cholinergic postganglionic neurons in the gland (3). CCK is known to be a major secretagogue and a trophic factor for the pancreas (4). It is synthesized in I cells of the small intestine and is released into the circulation following a meal. CCK is also present in neurons of the gastrointest...