Involvement of tetrodotoxin-resistant Na+ current and protein kinase C in the action of growth hormone (GH)-releasing hormone on primary cultured somatotropes from GH-green fluorescent protein transgenic mice.
GHRH depolarizes the membrane of somatotropes, leading to an increase in intracellular free Ca2+ concentration and GH secretion. Na+ channels mediate the rapid depolarization during the initial phase of the action potential, and this regulates Ca2+ influx and GH secretion. GHRH increases a tetrodotoxin-sensitive somatotrope Na+ current that is mediated by cAMP. TTX-resistant (TTX-R) Na+ channels are abundant in sensory neurons and cardiac myocytes, but their occurrence and/or function in somatotropes has not been investigated. Here we demonstrate expression of TTX-R Na+ channels and a TTX-R Na+ current, using patch-clamp method, in green fluorescent protein-GH transgenic mouse somatotropes. GHRH (100 nm) increased the TTX-R Na+ current in a reversible manner. The GHRH-induced increase in TTX-R Na+ current was not affected by the cAMP antagonist Rp-cAMP or protein kinase A inhibitors KT5720 or H89. The TTX-R current was increased by 8-bromoadenosine-cAMP (cAMP analog), forskolin (adenylyl-cyclase activator), and 3-isobutyl-1-methylxanthine (phosphodiesterase inhibitor), but the additional, GHRH-induced increase in TTX-R Na+ currents was not affected. U-73122 (phospholipase C inhibitor) and protein kinase C (PKC) inhibitors, Gö-6983 and chelerythrine, blocked the effect of GHRH. PKC activators, phorbol dibutyrate and phorbol myristate acetate, increased the TTX-R Na+ current, but GHRH had no further effect on the current. Na+-free extracellular medium significantly reduced GHRH-stimulated GH secretion. We conclude that GHRH-induced increase in the TTX-R Na+ current in mouse somatotropes is mediated by the PKC system. An increase in the TTX-R Na+ current may contribute to the GHRH-induced exocytosis of GH granules from mouse somatotropes.
Yang
Seung-Kwon
SK
Prince Henry's Institute of Medical Research, Melbourne, Victoria, Australia.
Wang
Kun
K
Parkington
Helena
H
Chen
Chen
C
D000818:Animals; D018929:Cell Culture Techniques; D002478:Cells, Cultured; D000242:Cyclic AMP / Q000502:physiology; D004351:Drug Resistance / Q000502:physiology; D005089:Exocytosis / Q000187:drug effects; D049452:Green Fluorescent Proteins / Q000235:genetics* / Q000378:metabolism; D013006:Growth Hormone / Q000235:genetics* / Q000378:metabolism; D013007:Growth Hormone-Releasing Hormone / Q000494:pharmacology*; D015640:Ion Channel Gating / Q000187:drug effects; D051379:Mice; D008822:Mice, Transgenic; D018408:Patch-Clamp Techniques; D011493:Protein Kinase C / Q000502:physiology*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology; D015222:Sodium Channels / Q000187:drug effects / Q000502:physiology*; D052683:Somatotrophs / Q000187:drug effects* / Q000378:metabolism / Q000502:physiology; D013779:Tetrodotoxin / Q000494:pharmacology*; D010738:Type C Phospholipases / Q000502:physiology
D011993:Recombinant Fusion Proteins; D015222:Sodium Channels; D049452:Green Fluorescent Proteins; D013779:Tetrodotoxin; D013006:Growth Hormone; D013007:Growth Hormone-Releasing Hormone; D000242:Cyclic AMP; D011493:Protein Kinase C; D010738:Type C Phospholipases