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Intracellular calcium changes in mice Leydig cells are dependent on calcium entry through T-type calcium channels

By Roberta Ribeiro Costa and Wamberto Antonio Varanda

Abstract

Luteinizing hormone (LH) regulates testosterone synthesis in Leydig cells by inducing an intracellular increase in cAMP concentration. LH also increases the intracellular calcium concentration ([Ca2+]i), dependent on the presence of Ca2+ in the extracellular medium ([Ca2+]e) for its effect. Despite these evidences, the identity of a pathway for calcium entry has remained elusive and the relationship between cAMP and [Ca2+]i has been questioned. Here we show that mice Leydig cells do have an inward Ca2+ current carried by T-type Ca2+ channels. In 10 mm [Ca2+]e, the currents start to be activated at −60 mV, reaching maximal amplitude of 1.8 ± 0.3 pA pF−1 at −20 mV. Currents were not modified by Ba2+ or Sr2+, were suppressed in Ca2+-free external solution, and were blocked by 100 μm nickel or 100 μm cadmium. The Ki for Ni2+ is 2.6 μm and concentrations of Cd2+ smaller than 50 μm have a very small effect on the currents. The calcium currents displayed a window centred at −40 mV. The half-voltage (V0.5) of activation is −30.3 mV, whereas the half-voltage steady-state inactivation is −51.1 mV. The deactivation time constant (τdeactivation) is around 3 ms at −35 mV. Confocal microscopy experiments with Fluo-3 loaded cells reveal that both LH and dibutyryl-cAMP (db-cAMP) increase [Ca2+]i. The db-cAMP induced calcium increase was dependent on Ca2+ influx since it was abolished by removal of extracellular Ca2+ and by 400 μm Ni2+. [Ca2+]i increases in regions close to the plasma membrane and in the cell nucleus. Similar effects are seen when Leydig cells are depolarized by withdrawing K+ from the extracellular solution. Altogether, our studies show that Ca2+ influx through T-type Ca2+ channels in the plasma membrane of Leydig cells plays a crucial role in the intracellular calcium concentration changes that follow binding of LH to its receptor

Topics: Cellular
Publisher: Blackwell Science Inc
OAI identifier: oai:pubmedcentral.nih.gov:2375479
Provided by: PubMed Central
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