Regulation of mucin secretion in T84 adenocarcinoma cells by forskolin: relationship to Ca2+ and PKC
Forstner, G.; Zhang, Y.; McCool, D.; Forstner, J.
American Journal of Physiology 266(4 Pt 1): G606-G612
1994
ISSN/ISBN: 0002-9513 PMID: 8178999 Document Number: 438705
The relationship between the adenosine 3',5'-cyclic monophosphatemediated protein kinase A (PKA)-dependent stimulatory pathway for mucin secretion and Ca-2+-mediated and protein kinase C (PKC)-mediated secretion was studied in T84 cells, using the postreceptor secretagogues forskolin, A-23187, and phorbol 12-myristate 13-acetate (PMA), the protein kinase inhibitors staurosporine and 1-(5-isoquinolinylsulfonyl)-2-methylpiperazine (H-7), high- and low-Ca-2+ media, and the Ca-2+ chelator 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid (BAPTA). Staurosporine (10-5 M) inhibited both PMA and forskolin at their maximally effective concentrations, whereas H-7 (5 times 10-5 M) inhibited only PMA. Stimulation of mucin secretion by forskolin (5 times 10-5 M) was not significantly affected by the reduction of medium Ca-2+ to 47 and 129 nM, equivalent to published values for intracellular Ca-2+ concentration ((Ca-2+)-i). Stimulation by forskolin was reduced by preloading cells with BA-PTA, but to a much smaller extent than Ca-2+-dependent Stimulation by A-23187. A-23187-mediated mucin secretion from BAPTA-loaded cells was augmented by high doses of forskolin. Similar concentrations of forskolin had no effect on A-23187-stimulated secretion in calcium-replete cells. Our results indicate that forskolin does not stimulate mucin secretion by increasing Ca-2+ entry or releasing Ca-2+ from intracellular stores. Forskolin can stimulate mucin secretion in a Ca-2+-independent manner but is apparently inhibited by high levels of intracellular Ca-2+ induced by Ca-2+ ionophores in 1.0 mM Ca-2+ media. PKA and PKC pathways stimulate mucin secretion independently and supportively in the presence and absence of Ca-2+. The Ca-2+-independent protein kinase-dependent response to calcium ionophores is more susceptible to H-7 than secretion mediated by forskolin, making it unlikely that calcium ionophores stimulate secretion by activating PKA.