Lacrimal and submandibular gland fluid secretion is regulated by changes in [Ca²⁺]_i. We have shown previously, in both mouse and human submandibular acinar cells, that NO has a biphasic effect on Ca²⁺ signalling (Caulfield et al., 2009). The first phase of the response is a cyclic-GMP mediated amplification of the CCh-evoked Ca²⁺ signal, most likely via cyclic-ADP ribose production and activation of ryanodine receptors. The second, chronic, phase of the response to NO is a slow inhibition of the CCh-evoked signal which was not blocked by ODQ and is therefore unlikely to be mediated by cGMP. We have speculated that modulation of the Ca²⁺ signal by NO could have a role in the pathophysiology of Sjögren’s syndrome, a disease of salivary hypofunction. Sjögren’s syndrome affects both salivary and lacrimal glands and therefore we have hypothesized that, if NO is important in the pathophysiology of this condition, it should also modulate Ca²⁺ signaling in lacrimal gland acinar cells. We have tested our hypothesis using isolated acinar cells, obtained by collagenase digestion of mouse lacrimal glands and maintained for 24-48 hours in primary culture. Changes in [Ca²⁺]_i were measured using Fura-2 microfluorimetry. We have measured the effects of exposure of the lacrimal acinar cells to the NO releasing agent SNAP on the magnitude of the Ca²⁺ signal elicited by repetitive CCh stimulation. Like submandibular cells, lacrimal cells showed a pattern of first amplification and then inhibition of the Ca²⁺ signal in the presence of NO. Unlike submandibular cells, lacrimal acinar cells also showed a similar pattern of response to repetitive CCh stimulation in the absence of NO. These data show a “use dependence” to repetitive stimulation by CCh in lacrimal cells that mirrors the effect of NO seen previously in submandibular cells. One possible mechanism for the “use dependence” is endogenous production of NO by lacrimal acinar cells in vitro. Preliminary experiments using L-NAME to block NO production do not support this mechanism.