Calcium (Ca²⁺) plays an important physiological role in mediating salivary gland secretion (Ambudkar, 2000). Cytosolic Ca²⁺ is derived from two sources, intracellular stores such as the endoplasmic reticulum and from extracellular medium. Secretagogue-evoked Ca²⁺ influx is referred to as capacitative Ca²⁺ entry (CCE) since it is the driving force which is responsible for prolonged and sustained salivary secretion (Putney, 1986). The precise mechanism involved in the regulation of CCE is still not fully understood (Ambudkar, 2000). This study investigated the effect of perturbation of extracellular magnesium [Mg²⁺]_o on CCE in isolated rat parotid acinar cells. Donor rats were humanely killed by rapid cervical dislocation, and parotid glands were surgically removed. An established method was used to prepare parotid acinar cells and load them with fura-2 (Baum et al. 1990). [Ca²⁺]_i was measured in single cells using microspectrofluorimetry (Lajas et al. 2000) in the absence and presence of 10^-5 M ACh during perturbation of [Mg²⁺]_o (0, 1.1, 5 and 10 mM). Initially, cells were perfused with a Ca²⁺ free medium containing the different [Mg²⁺]_o and 1 mM EGTA and then stimulated with ACh. After peak recovery, cells were perfused with a medium containing 2 mM Ca²⁺ in different [Mg²⁺]_o. [Ca²⁺]_i values in 35Ð40 experiments are expressed as mean (± S.E.M.) 340/380 ratio. All experiments were undertaken at room temperature.

Basal [Ca²⁺]_i in zero, normal (1.1 mM) and elevated (5 mM and 10 mM) [Mg²⁺]_o were 0.252 ± 0.006, 0.227 ± 0.009, 0.209 ± 0.013 and 0.211 ± 0.009 (n = 35Ð40), respectively. Stimulation of fura-2-loaded acinar cells with 10^-5 M ACh resulted in significant (ANOVA plus post-hoc test; P < 0.05) increases in the initial peak Ca²⁺ transient (Ca²⁺ released from intracellular stores) in zero and normal [Mg²⁺]_o compared with the respective controls. In contrast, elevated (5 mM and 10 mM) [Mg²⁺]_o attenuated the ACh-evoked Ca²⁺ transient compared with the responses obtained in zero and normal (1.1 mM) [Mg²⁺]_o. Typically, [Ca²⁺]_i in zero, normal and elevated [Mg²⁺]_o were 0.562 ± 0.016, 0.511 ± 0.016, 0.310 ± 0.018 and 0.293 ± 0.021 (n = 35Ð40), respectively. Perfusion of fura-2-loaded acinar cells with a physiological salt solution containing 2 mM Ca²⁺ resulted in significant (P < 0.05) increases in Ca²⁺ uptake (CCE) in zero and normal [Mg²⁺]_o compared with their respective basal values. The peak CCE was much more rapid (35 ± 5 s) in normal [Mg²⁺]_o compared with a slower uptake (226 ± 6 s) in zero [Mg²⁺]_o. In both 5 mM and 10 mM [Mg²⁺]_o CCE was significantly (P < 0.05) reduced and slower compared with the response obtained in normal (1.1 mM) [Mg²⁺]_o. The results indicate that Mg²⁺ can regulate secretagogue-evoked Ca²⁺ mobilization (both its release from intracellular stores and CCE) in parotid acinar cells.

All procedures accord with current national and local guidelines.