Possible roles of calcium and calmodulin in mammary gland differentiation in vitro.

In response to insulin, cortisol and prolactin, mammary gland explants from virgin mice will undergo differentiation, as measured by lactose synthetase activity and casein synthesis. W-13 (N-(4-aminobutyl)-5-chloro-2-naphthalenesulphonamide), a calmodulin inhibitor, reduced prolactin-induced differentiation by 80% while inhibiting insulin- and cortisol-induced RNA synthesis only 40%. However, tissue levels of calmodulin did not change during a 3-day culture, with or without prolactin, suggesting that the regulatory component of the calcium-calmodulin system was the intracellular calcium concentration. In support of this hypothesis, verapamil, a calcium channel blocker, also preferentially inhibited prolactin-induced differentiation and prolactin did stimulate calcium accumulation (from a control value of 3.8 +/- 1.3 (S.E.M.) pg calcium/mg wet tissue per 15 min to 61.4 +/- 7.0). On the other hand, A23187, a calcium ionophore which mimicked prolactin actions on calcium accumulation, was not able to induce differentiation in the absence of prolactin; in fact, high concentrations of either A23187 or extracellular calcium actually inhibited lactose synthetase activity, although casein synthesis was relatively unaffected. Therefore, it appears that the calcium-calmodulin system is involved in prolactin-induced differentiation of mammary gland explants but that it cannot be the only mediator of prolactin actions; i.e. it is necessary but not sufficient. There also appears to be another, separate action of calcium on casein synthesis which involves increasing the mammary epithelial sensitivity toward prolactin with respect to casein synthesis but not lactose synthetase activity.