Nitric oxide‐induced signalling in rat lacrimal acinar cells

The aim of the present study was to investigate the physiological role of nitric oxide (NO) in mediating secretory processes in rat lacrimal acinar cells. In addition, we wanted to determine whether the acinar cells possess endogenous nitric oxide synthase (NOS) activity by measuring NO production using the fluorescent NO indicator 4,5‐diaminofluorescein (DAF‐2). We initiated investigations by adding NO from an external source by means of the NO‐donor, S‐nitroso‐N‐acetyl‐penicillamine (SNAP). Cellular concentrations of cyclic guanosine 5′‐phosphate (cGMP) (cGMP]) were measured by radioimmunoassay (RIA), and we found that SNAP induced a fast increase in the cGMP], amounting to 350% of the cGMP] in resting cells. Moreover, addition of SNAP and elevating cGMP] in fura‐2 loaded lacrimal acinar cells, resulted in a cGMP‐dependent protein kinase‐mediated release of Ca²⁺ from intracellular stores, leading to a rise in the intracellular free Ca²⁺ concentration (Ca²⁺]_i). The Mn²⁺ quenching studies revealed that the Ca²⁺ release was not accompanied by Ca²⁺ influx. Finally, we demonstrate that lacrimal acinar cells possess endogenous NOS activity, which is activated by β‐adrenergic stimulation and not by a rise in Ca²⁺]_i alone. We show that in rat lacrimal acinar cells, NO and cGMP induce Ca²⁺ release from intracellular stores via G kinase activation. However, the changes in Ca²⁺]_i are relatively small, suggesting that this pathway plays a modulatory role in Ca²⁺ signalling, thus not by itself causing fast transient increases in Ca²⁺]_i. In addition, we suggest that endogenously produced NO activated by β‐adrenergic receptor stimulation, plays an important role in signalling to the surrounding tissue.