Severe acute pancreatitis(AP)has a significant associated mortality and morbidity rate and despite extensive studies the initial trigger for the disease is poorly understood. In recent years an impairment of [Ca2+ ]i signalling and an irreversible increase in [Ca2+ ]i (Ca2+ overload) has emerged as a possible unifying mechanism. Intra-peritoneal injections of high doses of L-arginine (Larg) is commonly used to experimentally induce AP in rodents(1),however, the underlying molecular mechanism remains controversial. The most notable explanation is that Larg is metabolised to nitric oxide (NO) by nitric oxide synthase (NOS) which elevates oxidative or nitrosative stress. However, studies using NOS inhibitors and NO donors have cast doubt on the role of NO during AP. An alternative explanation is that Larg activates a calcium-sensing receptor (CaSR) or the related GPRC6A receptor, resulting in Ca2+ overload and a consequent necrosis. In addition, L-ornithine (Lorn), a NOS-independent arginase-mediated metabolite of Larg, has been used to induce AP(2) and is also a potent activator of GPRC6A(3). The overall aim of the project was to explore the effects of Larg and Lorn on[Ca2+ ]i resting [Ca2+ ]i and CCK-evoked oscillations in pancreatic acinar cells. Larg and Lorn were used at concentrations that were equivalent or lower than those used to induce pancreatitis experimentally (100-500 mM). Pancreatic acinar cells were isolated from male Sprague-Dawley rats by collagenase digestion and [Ca2+ ]i was measured in fura-2-loaded cells using an epifluorescence imaging system. Larg and Lorn (100mM) induced a reversible spike-like increase in [Ca2+ ]i in 46% and 74% of cells respectively, and Ca2+ overload in 11% and 26% of cells respectively. At higher concentrations (500 mM) both Larg and Lorn induced Ca2+ overload in all cells. However, glycine (used as a negative control in experimental AP) and mannitol (used as a hyperosmotic control) also induced Ca2+ overload in 40% and 60% cells respectively at 500 mM. Isosmotic Larg and Lorn (100 mM)induced a reversible increase in [Ca2+ ]i in 74% and 87% cells respectively and failed to induce Ca2+ overload in any cells. Furthermore, isosmotic Larg and Lorn had no significant effect on the frequency of CCK-evoked [Ca2+ ]i oscillations, but caused a small increase in the area under the curve. Collectively these data suggest that Larg and Lorn-induced Ca2+ overload are most likely due to non-specific hyper-osmotic effects. Moreover, while isosmotic Larg and Lorn are capable of inducing reversible changes in [Ca2+ ]i, these effects are unlikely to mediate the cytotoxicity during experimental pancreatitis. Further studies using DCF and DAF-2-imaging will determine whether Larg and Lorn induced cytotoxicity is due to oxidative stress and NO respectively.