Simultaneous monitoring of Ca 2+ responses and salivary secretion in live animals reveals a threshold intracellular Ca 2+ concentration for salivation

New FindingsWhat is the central question of this study? The effects of Ca 2+ responses on salivary fluid secretion have been studied indirectly by monitoring ion channel activities and other indices. Therefore, Ca 2+ responses during salivary secretion remain poorly understood.What is the main finding and its importance? Herein, we developed a simultaneous monitoring system for Ca 2+ responses and salivary secretion in live animals using a YC‐Nano50‐expressing submandibular gland and a fibre‐optic pressure sensor. This new approach revealed a clear time lag between the onset of Ca 2+ responses and salivary secretion. We also estimated the [Ca 2+ ] i and provided direct evidence for the regulation of salivary secretion by small increases in [Ca 2+ ] i in submandibular gland acinar cells.Abstract We monitored changes in [Ca 2+ ] i during salivary secretion in the rat submandibular gland in live animals using a combination of intravital Ca 2+ imaging with the ultrasensitive Ca 2+ indicator YC‐Nano50 and a fibre‐optic pressure sensor. Intravenous infusion of ACh (10–720 nmol min −1 ) increased [Ca 2+ ] i and salivary flow rate in a dose‐dependent manner. Repetitive stimulation with ACh induced equivalent Ca 2+ responses and salivary secretion in the same individual animals. The accurate ACh stimulation experiments revealed a clear time lag between the onset of the increase in [Ca 2+ ] i and salivary secretion. The time lag with the lowest dose of ACh (30 nmol min −1 ) was 106 s, which shortened to 19 s with the dose used for maximal salivary secretion (360 nmol min −1 ). This time lag might reflect the time required for [Ca 2+ ] i to reach the level required to activate molecules for fluid secretion. The resting [Ca 2+ ] i in submandibular gland was 37 n m , and [Ca 2+ ] i at the onset of salivary secretion was 45–57 n m , irrespective of ACh dose. These results indicate that low [Ca 2+ ] i is sufficient to trigger fluid secretion in the rat submandibular gland in vivo .