M3 muscarinic receptor mediates regulation of protein secretion in rabbit lacrimal gland

Purpose. To identify which muscarinic receptor subtypes mediate protein secretion in isolated rabbit lacrimal glands. To compare protein secretory profiles in vitro with those in rabbit tears. Methods. Rabbit lacrimal gland slices were incubated with carbachol in the presence and absence of different muscarinic antagonists. Protein secretion into the incubation medium was characterized with a Bio-Rad protein assay dye reagent in conjunction with Laemmli's method of SDS-PAGE. The medium protein profile was compared to that in tear samples. Results. Dose dependent increases in protein secretion were elicited by carbachol between 10(-7) and 10(-4) M. During the first 20 min period, a maximal increase of 64% above the basal level was seen at the highest concentration. With 10(-4) M carbachol, the response was transient because after 100 min it decreased to its basal level. The increases in protein secretion caused by 10(-4) M carbachol were completely suppressed in the presence of 10(-5) M atropine. On the other hand, the relatively selective M1 antagonist, pirenzepine, at concentrations from 10(-6) M to 10(-4) M, had no effect on either the basal levels or the stimulatory effects of carbachol. Similarly, the relatively selective M2 antagonist, gallamine, at concentrations from 10(-6) M to 10(-4) M, had no effect on either of these levels. In contrast, the relatively selective M3 antagonist, 4-DAMP at concentrations from 10(-6) M to 10(-4) M, progressively suppressed the stimulated level of protein secretion elicited by 10(-4) M carbachol without affecting the basal level. The protein profiles found in the tears and in the incubation medium were similar to one another. Conclusion. In vitro rabbit lacrimal gland protein secretion is comparable to that in vivo. Cholinergic-mediated control of rabbit lacrimal gland protein secretion occurs through stimulation of the M3 muscarinic receptor subtype.