Stimulus-secretion coupling in the exocrine pancreas of the rat: Interactions between secretagogues

Abstract

1. Investigations have been carried out in two parts (a) in vitro studies into the roles of Mg2 in ACh and CCK3-evoked Ca 2 mobilisation assàciated with the secretory process and (b) in vitro and in vivo studies into the effects and mechanisms of action of phorbol esters (and related compounds) on secretagogueevoked 3H-protein release (ACh & CCK8) and pancreatic juice secretion (CCK8 & Secretin) in the rat pancreas.
2. Modifications in extracellular Mg 2 affected ACh-evoked 45Ca2 influx, efflux and intracellular free C a2+ mobilisation. Elevated extracellular Mg2+ (10 mM) depressed whereas, zero extracellular Mg2+ (0 mM) augmented these parameters
in comparison to normal extracellular Mg2+ (1.13 mM). ACh also elicited simultaneous transmembrane movement of Ca2+ (influx) and Mg2+ (efflux). Parallel secretory effects under the same conditions were observed when monitoring ct-aniylase and total protein outputs. Similar results were obtained for CCK8. Mg2+ facilitates control over the secretory process which occurs primarily through modifications in Ca 2 mobilisation.
3. The phorbol ester TPA (12-0-tetradecanoyl phorbol-l3-acetate) alone induced 3 H-protein release from permeabilised pancreatic acini but when combined with ACh a synergistic secretory response resulted at an intracellular free [Ca 2+]i = 10
4M. Lowering intracellular free [Ca 2+]i to 10 7M removed the potentiatory action of TPA but did not abolish AC!, induced secretion. Whereas, at [Ca2t] of 104M combining TPA with CCK8 resulted in complete depression of the CCK8-induced 3H-protein release. The action of TPA on CCK8-evoked 311-protein release was mediated through protein kinase C activation as Polymyxin B, an inhibitor of protein kinase C, reversed the effects of TPA. However, the potentiatory action of TPA on ACh-elicited 3 H-protein secretion was not mediated through protein kinase C activation or increases in cellular cyclic GMP. Hence, another cellular mechanism or mediator independent of protein kinase C activation possibly exists to account for both ACh-evoked secretion and the potentiatory action of TPA.
4. ACh and CCK8-evoked dose-dependent increases in intracellular free [Ca 2 ]. The dose-dependency of the Ca2+ transients elicited by these secretagogues showed a degree of correlation with their known secretory activity. Incubation of
the pancreatic acini with TPA inhibited CCK8-induced but NOT ACh-induced intracellular Ca2+ transients. Therefore, protein kinase C may facilitate to some extent its secretory modulatory activity through changes in intracellular C a2+ mobilisation.
5. In vivo administration of TPA to the anaesthetised rat resulted in increases in pancreatic juice secretion. Secretin administration caused marked increases in pancreatic juice flow but not of protein or u-amylase. Whereas, CCK8 administration induced marked increases in juice flow, protein and ct-amylase secretion. TPA combined with secretin or CCK8 differentially suppressed the secretory activity of these secretagogues. Inhibition of protein kinase C reversed to
some extent the inhibitory activity of TPA. Hence, it is believed that protein kinase C can physiologically initiate and modulate the quantity and quality of secretagogue-evoked pancreatic juice secretion irrespective of the intracellular pathway utilised in the pancreas of the rat.
6. In conclusion, the present study has demonstrated that Mg2+ plays an important role in stimulus-secretion coupling and therefore requires greater consideration when evaluating secretory events. Protein kinase C has been shown to possess an
essential modulatory role in the whole secretory process. Furthermore, the possibility exists that an as yet uncharacterised intracellular pathway may be responsible for the protein kinase independent actions of TPA.