Enhancement of Ca2+ currents by GHRH and its relation to PKA and [Ca2+]i in human GH-secreting adenoma cells.

Abstract

The effects of human growth hormone-releasing hormone (hGHRH) on Ca2+ channels were examined in human growth hormone-producing adenoma cells using the perforated whole cell clamp technique. These cells exhibited T- and L-type Ca2+ channel currents, and application of 10(-8) M hGHRH increased the amplitude of both currents. Application of 10(-5) M 8-bromoadenosine 3',5'-cyclic monophosphate also increased T- and L-type currents. Additional application of 10(-8) M hGHRH did not further increase the current amplitudes. Treatment with the Rp diastereomer of adenosine 3',5'-cyclic monophosphothioate (10(-5) M) or H-89 (10(-5) M) inhibited the enhancement of Ca2+ channel currents by hGHRH, as did intracellular injection of protein kinase A (PKA) inhibitor peptide [PKI-(5-24)], indicating that hGHRH increased the amplitude of Ca2+ channel currents through the activation of the adenosine 3',5'-cyclic monophosphate (cAMP)-PKA system. When intracellular Ca2+ concentration ([Ca2+]i) was chelated to < 30 nM with 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid acetoxymethyl ester (BAPTAAM), hGHRH failed to increase the Ca2+ channel currents. In this condition, hGHRH activated nonselective cation channels, which revealed that the cAMP-PKA system operated after treatment with BAPTA-AM and that the site of low [Ca2+]i-induced inhibition of hGHRH effects on Ca2+ channels was at a step after PKA activation.