Role of Intracellular Divalent Cations on the Adenylate Cyclase Activation by Human LH in mLTC-1 Leydig Cells

The Luteinizing Hormone (LH) regulates Leydig cell activities through LHR occupation, promoting Gs protein and/or β-arrestin activation. The activated GsαGTP subunit stimulates Adenylyl Cyclase (ACs) and, therefore, intracellular cyclic Adenosine Monophosphate (cAMP) accumulation from the AC substrate Adenosine Triphosphate (ATP). The divalent cations, magnesium (Mg2+) or manganese (Mn2+) associate with ATP to form the real substrates of ACs. In addition, ACs are sensitive to calcium (Ca2+) but in a different way than Mg2+ or Mn2+. Indeed, LH increases the cytoplasmic calcium ion concentration ([Ca2+]cyt) but only when Ca2+ is present in the extracellular medium. In the present study, the effects of intracytoplasmic Ca2+, Mg2+, and Mn2+ on the cyclic AMP response to human LH in mLTC-1 Leydig tumor cells were investigated. The mLTC-1 cells were incubated at 37 °C in media supplemented with and without 5 µM Ca2+, 5 µM Mg2+, or 5 µM Mn2+. The intracellular cyclic AMP accumulation was then monitored under LH stimulation. Our findings revealed that only Mg2+ and Mn2+ in the extracellular medium potentiate the cAMP response to hLH, in contrast to Ca2+. In addition, we also showed that HCO3- increased the stimulation of the adenylyl cyclase enzyme by Ca2+, Mg2+ or Mn2+. In mLTC-1 cells, extracellular Mg2+ and Mn2+ might potentiate LH-stimulated ACs activity by favoring LH interaction with its receptor, whereas Ca2+ from internal stores might be mobilized towards the cytoplasm to increase ACs activity, possibly through the soluble isoform.