Abstract
Hormone secretion form anterior pituitary cells is known to be regulated by G protein coupled receptors (GPCRs) that regulate intracellular cAMP. The objective of this study was to determine mechanisms by which elevated cAMP control intracellular calcium. We have used fura‐2 measurements of intracellular calcium concentration along with both population biochemical cAMP measurements and single cell FRET‐based cAMP measurements to investigate the relationship between [Ca2+] and cAMP in clonal rat MMQ lactotrophs. Activation vasoactive intestinal peptide receptors that are coupled to Gs led to Ca2+ influx as evident by increase in Ca2+‐oscillations. Bypass of GPCRs with the adenylyl cylclase activator forskolin also resulted in strong in increase in Ca2+ oscillations consistent with measured forskolin‐induced increase in [cAMP]. Pharmacological activation of PKA using 6‐Bn‐cAMP or activation of EPAC using 8‐cpt‐cAMP each resulted in increase in Ca2+ indicating a role for both cAMP binding proteins in control of calcium dynamics although with differing lag times likely reflecting distinct signaling pathways. The broad‐spectrum cAMP phosphodiesterase (PDE) inhibitor IBMX caused an increase in Ca2+ influx without delay. The PDE3 inhibitor milrinone caused a strong increase in Ca2+ after a delay whereas the PDE4 inhibitor rolipram caused an immediate increase in Ca2+. Roliparam also triggered apparent store release. FRET‐based measurements of [cAMP] are being used to further analyze these differences. These results suggest a possible different relationship between the various PDE sensitive pools of cAMP and Ca2+ ‐channels as well as distinct roles for PKA and EPAC in control of Ca2+ dynamics and secretion.
Published Version
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