Abstract
Salivary fluid secretion involves an intricate choreography of membrane transporters to result in the trans-epithelial movement of NaCl and water into the acinus lumen. Current models are largely based on experimental observations in enzymatically isolated cells where the Ca2+ signal invariably propagates globally and thus appears ideally suited to activate spatially separated Cl and K channels, present on the apical and basolateral plasma membrane, respectively. We monitored Ca2+ signals and salivary secretion in live mice expressing GCamp6F, following stimulation of the nerves innervating the submandibular gland. Consistent with in vitro studies, Ca2+ signals were initiated in the apical endoplasmic reticulum. In marked contrast to in vitro data, highly localized trains of Ca2+ transients that failed to fully propagate from the apical region were observed. Following stimuli optimum for secretion, large apical-basal gradients were elicited. A new mathematical model, incorporating these data was constructed to probe how salivary secretion can be optimally stimulated by apical Ca2+ signals.
Highlights
Ca2+ signal have been described in preparations of enzymatically isolated preparations of salivary gland acinar cells or lobules [30, 31] loaded with fluorescent Ca2+ indicators and imaged by time resolved wide field or confocal microscopy
421 An increase in [Ca2+] following neurotransmitter release from parasympathetic neurons is fundamentally important for the stimulation of salivary fluid secretion
It is recognized that 423 the spatiotemporal properties of intracellular Ca2+ signals in exocrine cells are pivotal for appropriately activating ion channels necessary for the underlying process [57,58,59]
Summary
To generate mice expressing a fluorescent Ca2+ indicator in exocrine acinar cells, homozygous mice expressing the fast genetically encoded Ca2+ indicator GCaMP6f A standing apical-basal [Ca2+] gradient was established, as shown in the SD image of the field and the topographical representation of the SD of the highlighted region (Fig. 9C/D), together with the analysis of manually positioned representative apical (red ROI), cytoplasmic (blue ROI) and basal regions (green ROI) (Fig. 9E) to illustrate the magnitude of the gradient To further investigate this spatial heterogenicity we performed MP line scans following stimulation at 3-10 Hz stimulation where the scanned line was orientated to interrogate changes in fluorescence originating in the apical region extending through the nucleus to the base of an individual cell. Optimal fluid secretion occurs when the apical and basal KCa densities are approximately equal
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