Effects of isoquinolonesulfonamides on action potential secretion coupling in pituitary cells

Background: Pituitary lactotrophs fire action potentials spontaneously and the associated voltage-gated calcium influx is sufficient to maintain high and steady prolactin release. Several intracellular proteins can mediate the action of calcium influx on prolactin secretion, including calmodulin-dependent protein kinases. Here, we studied effects of isoquinolonesulfonamides KN-62 and KN-93, calmodulin-dependent protein kinase inhibitors, and KN-92, an inactive analog, on spontaneous electrical activity, voltage-gated calcium influx, cyclic nucleotide production, and basal prolactin release. Methods: The effects of these compounds on electrical activity and calcium signaling was measured in single lactotrophs and cyclic nucleotide production and prolactin release were determined in static culture and perifusion experiments of anterior pituitary cells from postpubertal female rats. Results: KN-62 and KN-93 blocked basal prolactin release in a dose- and time-dependent manner, suggesting that calmodulin-dependent protein kinase could mediate the coupling of electrical activity and secretion. However, a similar effect on basal prolactin release was observed on application of KN-92, which does not inhibit this kinase. KN-93 also inhibited cAMP and cGMP production, but inhibition of prolactin release was independent of the status of cyclic nucleotide production. Single cell measurements revealed abolition of spontaneous and depolarization-induced electrical activity and calcium transients in KN-92/93-treated cells, with a time course comparable to that observed in secretory studies. Conclusions: The results suggest that caution should be used when interpreting data from studies using isoquinolonesulfonamides to evaluate the role of calmodulin-dependent protein kinases in excitable endocrine cells, because inactive compounds exhibit comparable effects on action potential secretion coupling to those of active compounds.