Endoplasmic Reticulum–Mitochondria Calcium Drives Salivary Bioenergetics

Saliva secretion requires continuous energy supply throughout the day. Mitochondria dynamically adapt to fluctuating energy demands, yet the mechanisms underlying the adaptions remain poorly understood. Here, we employed real-time intravital imaging and fluorescence lifetime imaging microscopy (FLIM) to monitor mitochondria functions in submandibular glands. We revealed distinct mitochondrial distribution patterns; in acinar cells, mitochondria were predominantly distributed near the cell membranes or scattered throughout the cytoplasm with extensive endoplasmic reticulum (ER)-mitochondria contact sites, whereas in ductal cells, mitochondria were densely packed within the cytoplasm. At resting states, mitochondria exhibited larger volumes, fewer numbers, and higher oxidative phosphorylation activity in acinar cells compared with those in ductal cells. Upon stimulation with pilocarpine, mitochondrial motility, NAD(P)H levels, NAD(P)H enzyme-bound fractions, and mitochondrial adenosine triphosphate (ATP) production were significantly elevated. Pilocarpine-induced secretion, mediated by both aquaporin 5 translocation and the opening of paracellular pathway, was markedly attenuated by oligomycin A, an ATP synthase inhibitor. Notably, pilocarpine increased mitochondria-ER contact sites to 1.7 times the control level (from 18% to 31%), and blocking mitochondrial calcium uptake significantly suppressed pilocarpine-induced NAD(P)H and ATP production. These findings highlight the critical role of ER-mitochondria calcium transfer in sustaining bioenergetics required for salivary secretion, providing new insights into mitochondrial functional adaptation and its physiological significance in intact secretory systems.