Pepsinogen secretion from perifused frog peptic glands: rapid transients detected with a modified pepsin assay

In order to define the early transient responses of peptic cells to stimulation, we have modified the acidified hemoglobin substrate digestion method for pepsin to provide a very sensitive, linear (10-250 ng/ml), reproducible, inexpensive (less than 3 cents/sample) and simple semi-automated assay. Using a specially designed perifusion chamber, this method was used to accurately measure secretion at 1 min intervals from approximately 3 mg of isolated peptic glands from the esophageal peptic organ of R. catesbeiana, containing approximately 150 micrograms pepsinogen. Responses to carbachol applied for 1, 2 and 4 min could be described in discrete 1 min intervals. Secretion stimulated by carbachol (1-2 min) peaked at 200-300% of basal and upon withdrawal decayed with t1/2 approximately 3 min. Atropine added to continuous carbachol stimulation inhibited secretion with t1/2 approximately 4 min, indicating rapid metabolism of activated messengers.